Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1-4

The Drug Discovery Process Hugo Kubinyi Weisenheim am Sand, Germany

HISTORY AND CURRENT SITUATION Early drug discovery was dominated by natural products and their derivatives [1–3]. Accumulated experience from folk medicine and subsequent systematic search uncovered the active principles of many plants. These sources were complemented by active principles from microorganisms, with antibiotics being the Þrst. Recently, microorganisms turned out to be a source for other important drugs. About half of all therapeutics are natural products, their derivatives, or synthetic compounds that were derived from natural products [4]. The father of the Bayer chemist Felix Hoffmann suffered from rheumatic disease. The prescribed salicylic acid had a bitter taste and unpleasant side effects. In 1897, Hoffmann synthesized acetylsalicylic acid (ASA) for the Þrst time as a pure, chemically stable compound. Today, we would call this a prodrug approach, and indeed the goal had been to produce a better and more tolerable salicylic acid analog. Heinrich Dreser, the Bayer pharmacologist at that time, was not happy about the new compound. Whereas he favored heroin, the diacetyl derivative of morphine, as an “effective and well tolerated antitussive,” he tried to stop the development of ASA because in animal experiments he had seen some cardiac side effects. Being clinically tested without his consent, the excellent antipyretic and analgesic activities of ASA immediately convinced clinicians and patients. The drug, then called Aspirin, was introduced into therapy in 1899. Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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ASA is one of the greatest success stories in drug discovery and most probably is the leader in the number of pills consumed in the past 108 years. In the 1970s the mechanism of action was elucidated: it irreversibly blocks cyclooxygenase, the enzyme responsible for the biosynthesis of pain-mediating prostaglandins. At that time, it was also recognized that ASA prevents thrombocyte aggregation via the same mechanism, by inhibition of thromboxane biosynthesis, because platelet cyclooxygenase cannot be regenerated during the lifetime of a thrombocyte. Thus, ASA has been a serendipitous drug discovery, as so many others in the history of drug research [1, 5, 6]. A period of thirty years, from about 1950 to 1980, marks the “golden time” of drug research. Many highly valuable drugs were discovered in a few decades, applying rational approaches to develop neurotransmitter, steroid, and peptide analogs [7]. Nonselective as well as selective receptor agonists and antagonists resulted (e.g., the antihistamines). Despite the fact that these early H1 antagonists had strong sedative side effects, they were celebrated as “miracle drugs.” The many new technologies developed in the past twenty years should increase the number of new drugs that can be used in human therapy. Presently, however, the opposite is the case. Since 1995 there has been a steady decline in the number of new drugs entering the market (Table 1). The year 2005 showed a marked decline after a short recovery in 2004 [8]. In contrast, research costs in the pharmaceutical industry doubled about every seven years, from 1991 to 1998 and again from 1998 to 2005, up to about $40 billion US per year—constituting the “productivity gap” in drug discovery. Research costs for a new drug are now estimated to be in the range of $800 million US [9]. However, considering all failures in drug research and comparing worldwide research and development costs with the number of NCEs, this Þgure becomes even higher, approaching almost $2 billion US per new drug. Several reasons have been advanced for the decline in new drugs. We already use safe and efÞcient drugs to treat (most often symptomatically) the “simple” diseases. Our demands for the safety of new drugs are increasing; today,

TABLE 1. New Drugs Approved by the FDA in the Years 1996–2005 [8] Year

NCEsa

Biologicals

1996 1997 1998 1999 2000 2001 2002 2003 2004 2005

53 39 30 35 27 24 17 21 31 18

3 6 7 3 2 5 7 6 4 2

a NCEs, small molecule new chemical entities.

Total 56 45 37 38 29 29 24 27 35 20

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several established drugs, from ASA to corticosteroids and other active agents, would have problems getting FDA approval. In many diseases, especially cancer, the development goes from nonspeciÞc cytotoxic agents to drugs that speciÞcally interact with certain signaling chains (e.g., the kinase inhibitors). However, kinases are a group of about 500 homologs with very similar binding sites, thus obstructing the design of speciÞc inhibitors. Drugs for the treatment of chronic diseases have to be given for many years and drugs for the prevention of a certain disease have to be given to healthy people. Such therapeutics must be absolutely safe, whereas more severe side effects can and have to be accepted in the case of anticancer drugs. As discussed later, early combinatorial chemistry and high-throughput screening produced large numbers of “hits” that were not suited as lead structures—they were too large and too lipophilic. In lead optimization, afÞnity to a target was often overemphasized, neglecting selectivity, bioavailability, other ADME parameters, and toxicity. Another reason for the poor yield in new drugs might be their long development time. Several new technologies in drug discovery, like structure-based and computer-aided design, virtual screening, and fragment-based design, are just starting to produce interesting results. On the other hand, molecular modeling goes back about twenty-Þve years and QSAR has existed for more than forty years. Although both approaches had no direct impact on drug discovery, they stimulated drug research in indirect ways: QSAR helped medicinal chemists to understand the inßuence of lipophilicity and dissociation constants [10] and molecular modeling forced chemists to consider molecules as threedimensional (sometimes chiral), ßexible objects [11]. The Þrst drugs that resulted from structure-based design are already on the market; many more will (hopefully) follow. The pipelines of the pharmaceutical and biotech companies are full of drug candidates from such rational approaches. The decline in new drug approvals is only one facet of the problem. The other one is that several drugs were approved and introduced into the market but had to be withdrawn shortly afterward, due to insufÞcient efÞcacy or toxicity problems. Well-known examples from recent years are terfenadine, cisapride, mibefradil, cerivastatin, rofecoxib, and ximelagatran, some of which were already blockbuster drugs or at least bearing the potential to become blockbusters [12]. In all cases, the reason for withdrawal was an unfavorable balance between beneÞcial and adverse toxic effects. The decline in drug approvals is in strong contrast to the hope that resulted from genomics, proteomics, system biology, and many other experimental and computer-aided new technologies. Whereas the FDA and other agencies considered lowering the barrier for clinical phase I studies, even to the point of accepting pre-phase I studies in humans, a strong question mark was put on such a procedure by the severe toxic reactions experienced by some healthy volunteers who were treated with a monoclonal antibody in a phase I study in early 2006 [13]. Since the most important steps in drug discovery are lead structure search and optimization, the focus is mainly on these two issues.

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DRUG TARGETS Gene technology and especially information on the human genome sequence created a lot of hype. Originally, there were estimates of about 100,000 or more human genes, which after 2001 had to be corrected to only about 30,000–35,000; recent estimates are even closer to 20,000–25,000. Drews counted 483 targets of current therapies and he speculated that in total there might be about 5,000–10,000 drug targets [14]. Hopkins and Groom arrived at a much smaller number of 600–1,500 drug targets in their estimate of the “druggable genome” [15]. However, the term “druggable genome” is misleading: the relatively small number of genes within our genome codes for hundreds of thousands of different proteins, due to alternative splicing, post-translational modiÞcations, and protein complex formation. If only a small percentage of them are involved in disease, a much larger number of potential targets results [16]. Gene technology contributes to drug research in different ways. Sequence determination of the coding regions of a gene provides the sequence of the corresponding protein, which often allows the prediction of its function, sometimes even its fold and three-dimensional (3D) structure. Proof of a new therapeutic approach can be performed in knock-out or other transgenic animal models or by silencing genes with siRNA technology. One of the major beneÞts of gene technology is the fact that almost any protein can be produced in cell culture. Thus, instead of animals or animal proteins, human proteins are used in screening. Production of larger amounts of a human protein enables its crystallization and 3D structure determination by protein crystallography or structure determination by NMR. In this way, gene technology supports drug research, without a patient having to take any genetically produced or modiÞed products: this is probably the greatest beneÞt of gene technology for humankind. There is an ongoing discussion on the extent to which a certain drug should be speciÞc for just one target or should act on several targets (in earlier times belittled as “dirty drugs,” but today praised as compounds with “rich pharmacology”). There is no answer from theory; experience shows that in many cases high selectivity might be favorable, whereas successful CNS-active drugs often modulate some to many targets. Even kinase inhibitors for tumor therapy are not completely selective. The recently approved drug sunitinib (Sutent, SU 11248; Sugen, PÞzer) for the treatment of renal and gastrointestinal tumors is a highly promiscuous inhibitor of a large number of different kinases [17]; possibly its therapeutic value results just from this lack of selectivity. A related question is: Do we lose too many potential drugs by target-based screening? The very Þrst antibacterial sulfonamide, sulfamidochrysoidine, would not have been discovered in vitro because it is a prodrug; clopidogrel has to be activated by CYP3A4 oxidation; acyclovir is monophosphorylated in virusinfected cells by a viral thymidine kinase and only afterward do cellular kinases convert the monophosphate to the biologically active triphosphate; and omeprazole acts only in acid-producing cells, after acid-catalyzed rearrangement.

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So are whole animal experiments better suited than target-based screening? There is no going back to mice and guinea pigs as primary test models. Chemical biology, which aims to discover new leads by searching for phenotypic changes in cells, for example, embryonic stem (ES) cells or Saccharomyces cerevisiae, or small animals, like Caenorhabditis elegans, Drosophila, and the zebraÞsh, Danio rerio, is a step in the right direction. Some results from chemical biology approaches are synthetic small molecules that induce the differentiation of embryonic stem cells. A high-throughput phenotypic cell-based screen identiÞed a pyrrolopyrimidine that differentiates ES cells to neurons by modulating glycogen synthase kinase-3β (GSK-3β) [18]. Cardiac muscle cell formation from stem cells is induced by cardiogenol C, which was discovered in a 100,000-member heterocycles library [19]. The same group described the dedifferentiating agent reversine that (seemingly) converts adult murine myotube cells to mesenchymal progenitor cells, which can then be induced to form either bone or fat cells [20, 21].

THE SEARCH FOR NEW LEADS One reason for the decline in the number of new drugs, which has not been mentioned so far, is a shortage of new lead structures that can be optimized into therapeutically useful drugs. A lead structure must have some biological activity, albeit weak and even nonselective, and there must be chemically related analogs, which indicate that structural modiÞcation modulates biological activity as well as other properties. The compounds should not react irreversibly with its biological target (although some important drugs, like acetylsalicylic acid, the penicillins and cephalosporins, and omeprazole, are irreversible enzyme inhibitors). Drug candidates most often become larger in size and more lipophilic. Thus, a lead should have a molecular weight <350 and a lipophilicity, expressed by log P (P = n-octanol/water partition coefÞcient), smaller than 3 [22, 23]. As already mentioned, natural products have been the primary source of drugs and lead structures [1–4]. Besides the classical examples of salicylic acid, quinine, morphine, papaverine, ephedrine, atropine, cocaine, cardiac glycosides, and curare alkaloids, some recent plant-derived drugs are the antitumor drugs taxol and camptothecin, the anti-Alzheimer natural product huperzine, and the antimalarial drug artemisine. Whereas originally microorganisms were only investigated as producers of antibiotics (e.g., penicillin, cephalosporin, and streptomycin), several other important classes of drugs resulted from microorganisms. In the Þrst place, the cholesterol-lowering statins have to be mentioned but also the hallucinogenic lysergic acid diethylamide (LSD) from ergot, the immunosuppressants cyclosporin A and tacrolimus, and the antitumor drug epothilone. Even the anticoagulants phenprocoumon and warfarin had their origins in the microbial degradation product dicoumarol, which was isolated from rotten hay after discovering cattle bled to death after consuming the hay [1, 3, 6].

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Some of the very Þrst drugs were discovered by serendipity [1–3, 5, 6]: for example, the narcotic gases and nitroglycerin as an antiasthmatic agent. Some other drugs resulted from incorrect working hypotheses: for example, the sedative urethane, which was supposed to release ethanol but is a hypnotic itself. The story of serendipitous discoveries continues with penicillin, the benzodiazepines, LSD, and clonidine [1, 3, 6]. It should also be mentioned that all major artiÞcial sweeteners (i.e., saccharin, cyclamate, and aspartame) were serendipitous discoveries; chemists experienced the sweet taste when licking their Þngers or smoking a cigarette [5]. A large number of therapeutically useful receptor agonists and antagonists resulted from the understanding of the biochemical mechanisms of nerve impulse transmission and hormone action. This phase of drug research may be considered its golden age [7]. Mimics of the transition state of an enzyme substrate serve as lead structures for enzyme inhibitors. In this manner, protease inhibitors [24] start from cleavage-site peptides, where the relevant amide bond is converted into another functionality, either into a group that binds to or reacts with a catalytic amino acid (serine and cysteine proteases), a group that coordinates a metal ion (metalloproteases), or an enzymatically stable isostere of the peptide bond that is attacked by the protease (aspartyl proteases). Another important source of new lead structures is screening. Tests in animals or isolated organs are now almost exclusively replaced by high-throughput screening (HTS) in vitro [25], using human targets. There is no question that animal tests contributed to the discovery of many valuable leads but it is no longer acceptable, because of animal protection issues and the huge number of tested compounds. With automated high-throughput screening, the situation originally worsened: mass screening of in-house compound collections, any commercially available compounds, or combinatorial libraries produced many hits that could not be further optimized. Poor solubility, unknown concentration, compound decomposition in the storage solution, as well as colored impurities and ßuorescence of some compounds, produced many false positives; retesting often did not conÞrm the primary hits. On the other hand, false negatives resulted. Tests of analogs that are similar to conÞrmed hits uncovered their activity, although they were initially not found to be active. Some structural classes seemingly bind with micromolar afÞnity to various enzymes; their promiscuous behavior is not caused by binding to certain sites but by aggregation of the ligands and clumping of these aggregates to the protein, in this manner mimicking biological activity [26–28]. Even more disappointing than HTS results with historical compound collections was the success rate of combinatorial libraries, especially in the early years. Syntheses were driven by chemical accessibility and huge libraries of ill-deÞned mixtures were tested, without reproducible results. The situation changed only after introduction of the Lipinski rule of Þve [29], which deÞnes that drug candidates should have a molecular weight <500, a lipophilicity range of log P < 5, no more than Þve hydrogen bond donors, and no more than ten N and O atoms (as a rough estimate of the number of hydrogen bond acceptors) in the molecule. Poor bioavailability is to be expected if two or more of these conditions are

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violated [29]. Originally designed only as a warning ßag for PÞzer chemists, the rule of Þve made the scientiÞc community aware of the importance of certain compound properties, especially appropriate molecular weight and balanced lipophilicity. Other groups explored polar surface area as a factor determining bioavailability and blood–brain barrier penetration [30, 31], as well as the number of rotatable bonds [32]. As alternative to HTS, NMR turned out to be well suited for the discovery of small, low-afÞnity ligands. Such small molecules have a better chance to bind to a certain target, due to their lower complexity. The concept of combining low-afÞnity ligands to a high-afÞnity ligand has been used by Fesik in the SAR by NMR strategy [33]. This experimental method searches for relatively small ligands that bind to adjacent pockets of a binding site of small proteins. In the next step, the fragments are connected to a high-afÞnity ligand [33–35]. Several other NMR-based techniques have been developed to search for fragments, which later can be combined to new lead structures [36–40]. Soaking of protein crystals with a mixture of small ligands that differ in size and shape, in combination with high-throughput crystallography, is another promising new approach in lead discovery [41, 42]. Another experimental method, the dynamic assembly of ligands [43–45], generates ligands from fragments that reversibly react with each other. In the presence of a target protein, those ligands that Þt the binding site bind to the protein. Afterward they are trapped by a reaction that freezes the equilibrium. Some other fragment-based approaches have recently been reviewed [46, 47]. In addition to these experimental techniques, there are several computerassisted techniques for the combination of fragments to new leads. TOPAS [48, 49] is a program that dissects lead structures and drugs into fragments and assembles them into new molecules; cleavage and assembly of the molecules follow chemical reactions that are deÞned by the RECAP procedure [50]. In this manner, a “scaffold hopping” [51] is achieved, leading to new chemistry. One of the greatest achievements in molecular modeling is the pharmacophore searches [52–55]. By deÞnition, a pharmacophore is “the ensemble of steric and electronic features that is necessary to ensure the optimal supramolecular interactions with a speciÞc biological target structure and to trigger (or to block) its biological response” [56]. Neither the generation of a pharmacophore from a series of active and less active molecules nor pharmacophore 3D searches are an easy task. Both have to consider the pharmacophore properties of the different atoms and groups and the ßexibility of the molecules. The computer program GRID [57–59] inspects the binding site of a protein with different chemical probes, to search for “hot spots” where a certain functionality of a ligand should favorably interact. Alternatives to GRID are IsoStar [60] and SuperStar [61–63], programs that extract the statistics of nonbonded intermolecular interactions from the Cambridge Crystallographic Database [64]. CATALYST is used for the generation of pharmacophore hypotheses and for 3D database searches [65–68]. The new program LigandScout is an attractive option to automatically generate a ligand pharmacophore from the 3D structure of the ligand–protein complex [69].

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The FTrees (feature trees) and Mtree (multiple feature trees) approaches enable extremely fast similarity searches with huge numbers of molecules, by comparing their pharmacophore similarity [70–72]. Virtual screening is now used extensively in lead structure search. A toolbox of methods is available, from simple Þltering and pharmacophore searches to docking and scoring. The goal is to enrich libraries and to discover promising lead candidates, starting from a 2D or 3D structural database of real compounds or from a virtual database [73–79]. As the input are chemical structures and/or estimated properties of compounds, virtual screening can be applied to virtual libraries of almost any size. Most important is a proper preprocessing of the database, deÞning the right protonation state, for example, by a set of rules (a problem that still awaits a satisfactory solution), and deÞning the most prominent tautomer of a compound or all possible tautomers. If 3D searches are performed, correct constitutions and correct or alternative enantiomers (diastereomers) must be generated, as well as reliable 3D structures or even multiple 3D structures. For similarity searches, superposition of molecules, pharmacophore searches, and docking, a correct deÞnition of hydrogen bond donor and acceptor properties is of the utmost importance [16]. The Lipinski rule of Þve [29] should be applied to select compounds with the potential for oral bioavailability, whereas neural nets have been trained for the identiÞcation of compounds with drug-like structures [80, 81]. Filters for cytotoxicity, toxicity, mutagenicity, and cancerogenicity have to be applied with extreme care; too many different Þlters generate too many false positives—that is, they eliminate nontoxic molecules that are estimated to be toxic. Unfortunately, some of these Þlters have only poor test set predictivity, coming close to chance prediction.

LEAD STRUCTURE OPTIMIZATION Lead structure optimization is a trial-and-error procedure, dominated by medicinal chemistry know-how and creativity. One guideline for chemical variation of a lead is the so-called isosteric replacement, the exchange of a certain atom or group (e.g., a halogen atom) by more or less equivalent groups (e.g., a trißuoromethyl or methyl group). Whereas in most cases only minor modiÞcations of biological properties result, sometimes a signiÞcant increase or decrease of activity or a change in selectivity is observed [82]. Other modiÞcations are the elimination of substituents or groups, introduction of new substituents, cleavage or formation of rings, chain length or ring size variation, and scaffold hopping [83]. “Me too” research has a negative image but often an improvement of certain biological properties results. Superior analogs are, for example, the bioavailable, broad-spectrum, and lactamase-resistant penicillins, highly speciÞc mineralo- and glucocorticoids, polar H1 antihistaminics without sedative side effects, or β1 speciÞc antagonists as well as partial agonists [84]. Many peptide substrates of enzymes and many enzyme and receptor ligands (e.g., the serpins, enkephalins, neurokinins, somatostatin, Þbrinogen, and

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vitronectin) are either peptides or proteins. The interaction of these ligands with their target is often mediated by only a few amino acid side chains. Peptide analogs can easily be synthesized in large numbers; thus high-afÞnity ligands can be discovered in a short time. However, the chemical conversion of such a peptide into a nonpeptidic (“peptidomimetic”) bioavailable drug is difÞcult. With the exception of the benzodiazepines, most other peptide mimics have not yet produced active compounds. Some examples where the conversion of (cyclic) peptides into peptidomimetics has been successful are the integrin ligands [85–87]. Other success stories are neurokinin-1 receptor ligands [88] and somatostatin receptor ligands with pronounced receptor subtype selectivity [89]. Several peptidomimetic HIV protease inhibitors could be derived from the sequence around the cleavage site [90]. The Þrst anti-HIV drugs, saquinavir, ritonavir, and indinavir, are still peptide-like molecules, whereas nelÞnavir and amprenavir, derived by structure-based design, as well as tipranavir and the DuPont inhibitors [91, 92] are real peptidomimetics. These success stories demonstrate that the conversion of peptides into peptidomimetics is indeed possible. On the other hand, peptidomimetic design is not a straightforward, generally applicable strategy; billions of dollars have been spent on renin and thrombin inhibitor research, without any drugs currently being on the market. Prodrug design is a common strategy in lead optimization, by converting drug candidates with good in vitro but insufÞcient in vivo properties (e.g., poor bioavailability) into appropriate derivatives that are orally available [93]. Sometimes just lipophilicity is enhanced by masking a polar group (e.g., in oseltamivir); sometimes the derivatives are actively transported (e.g., enalapril, the monoester of the active metabolite enalaprilate). In other cases, absorption and blood–brain barrier penetration are achieved by active transport (e.g., L-dopa). Some prodrugs exert their activity only in certain cells (e.g. acyclovir, after monophosphorylation in virus-infected cells, and omeprazole, after acid-catalyzed activation in acid-producing cells). Soft drugs are active derivatives of inactive drug analogs, for example esters of corticosteroid 21-acids, which are topically active but are immediately metabolically degraded to the biologically inactive 21-acids, after dermal absorption. Sometimes toxic compounds can be “rescued” by switching to an active metabolite. Paracetamol, the active metabolite of phenacetin, is better tolerated and has replaced phenacetin; only recently, the mechanism of action of both drugs could be elucidated [94]. Similarly, the nonsedative H1 antagonist terfenadine had to be replaced by fexofenadine. Terfenadine, a hERG (human ether-a-go-go-related gene) channel inhibitor, is nontoxic when it is rapidly oxidized to its well-tolerated active metabolite fexofenadine; however, it causes fatal toxic effects if metabolism is inhibited by comedication of a CYP3A4 inhibitor [95, 96]. In the past, chiral drugs were often developed as racemates or as diastereomeric mixtures, neglecting the effect that, in general, optical isomers have different biological effects quantitatively and qualitatively. In recent years, companies

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have started to extend the lifetime of their chiral drugs, if originally developed as a racemate, by a “chiral switch,” that is, by replacing the racemate by its biologically active enantiomer [97]. The poor start of combinatorial chemistry has already been mentioned. In the meantime, combinatorial chemistry developed into automated parallel synthesis of much smaller libraries of single, pure compounds of biological interest. Today, its main application is not so much in lead structure search but in lead validation and in early phases of lead optimization [98]. In chemogenomics, congeneric chemical libraries are screened in certain target families, for example, the G protein-coupled receptors (GPCRs), a subsection of GPCRs or certain GPCR subtypes, nuclear receptors, different protease families, kinases, phosphodiesterases, ion channels, and transporters [99–103]. Although strategies to screen compounds against related targets had been developed earlier, chemogenomics aims to discover highly potent, selective ligands against functionally and evolutionary related targets in a much more systematic manner, with the least effort. Several examples show that surprising activity and selectivity differences may result by following such a strategy [104]. From the observation that certain classes of compounds tend to produce more active analogs than others, for different targets, Evans and co-workers formulated the concept of “privileged structures” [105]. Examples of such privileged structures are benzodiazepines, phenethylamines, diphenylmethanes, diphenylamines, tricyclics, steroids, arylpiperazines, arylspiropiperidines, and biphenyltetrazoles [106]. In a continuation of this concept, Wermuth proposed the SOSA (selective optimization of side activities) approach [107–109]. The best-known historic examples for an optimization of side activities are the diuretic and antidiabetic sulfonamides, which resulted from antibacterial sulfonamides after the clinical observation of corresponding side effects, and the development of tricyclic neuroleptics and antidepressants from antihistaminics. Some recent reviews list many more examples [104], including also the work of Wermuth and his group, who performed chemical variations to optimize some weak side effects of the antidepressant minaprine to nanomolar acetylcholinesterase inhibitors, corticotrophin releasing factor (CRF) receptor antagonists, and muscarinic M1 agonists, and from these M1 agonists further to 5-HT3 antagonists [107–109]. Another striking example is the antileukemic drug imatinib (Gleevec®, Novartis), a bcr-abl inhibitor, which resulted from PKC inhibitors via a nonspeciÞc PKC/abl inhibitor [110]. As with chemogenomics, the SOSA principle is not new but its systematic application may yield more drugs, with less effort than blind screening. Sometimes it is not so important to optimize a chemical structure as to apply a biochemical approach. This was realized decades ago by the rational design of a drug combination to treat Parkinson’s disease. Since this disease is caused by a loss of dopamine-producing cells in a certain brain area, the substantia nigra, a logical therapy is dopamine substitution. However, dopamine has poor bioavailability, does not permeate the blood–brain barrier (BBB), and produces peripheral side effects after intravenous application. On the other hand, its precursor L-dopa is actively transported through the intestinal wall and through the BBB. In the

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brain it is decarboxylated to substitute the lacking dopamine. As decarboxylation of L-dopa in the cardiovascular system would cause an increase in heart rate and blood pressure, a polar, non-BBB-permeable decarboxylase inhibitor is given in combination. A further comedication is a nonpolar, BBB-permeable monoamine oxidase inhibitor, to prolong the action of dopamine in the brain. A big step forward in drug discovery could be achieved by the use of protein 3D structures in ligand design. The very Þrst drug that resulted from structurebased design was the angiotensin-converting enzyme (ACE) inhibitor captopril; its lead structure was modeled from the 3D structure of a carboxypeptidaseinhibitor complex [111]. Other drugs followed, for example, dorzolamide [112] and the HIV protease inhibitors nelÞnavir and amprenavir [113]; many more are in clinical development. A most impressive result of structure-based design resulted from the application of the program GRID to the enzyme neuraminidase of the inßuenza virus. A pocket of the binding site was discovered, where a positively charged substituent of a low-afÞnity transition state mimic should enhance biological activity. This was indeed the case: exchange of a hydroxy group against a guanidinium group increased afÞnity by four orders of magnitude, leading to the drug zanamivir [114]. This success of structure-based design also shows the shortcomings of this approach: zanamivir is a highly active ligand but it is orally unavailable—it has to be applied by inhalation. After discovery of the less polar neuraminidase inhibitor GS 4071, marketed as the orally active prodrug oseltamivir [115], zanamivir lost about 90% of its market share. About 35,000 3D structures of proteins and protein–ligand complexes are currently deposited in the Protein Databank (PDB) and many more will be available in the near future, due to high-throughput techniques in protein crystallization and structure determination [116]. With the increasing number of experimental 3D structures, homology modeling is also becoming more and more important. Some problems in the application of X-ray crystallographic data in drug design were discussed by Davis et al. [117]. Structure-based design is supported by computer-aided techniques. DOCK was the Þrst computer program for a geometric docking of ligands into a binding site [118]. Further progress resulted from the program LUDI [119, 120], which deÞnes interaction sites and uses a scoring function [121] to evaluate the docking results. Programs for ßexible docking of ligands into a rigid binding site include DOCK in its newer versions [122], GOLD [123], FlexX [124, 125], and the public domain program AutoDock [126, 127]; the FlexX modiÞcations FlexE [128] and Flex-Pharm [129] allow ßexible ligand docking into an ensemble of different binding site conformations and the deÞnition of pharmacophore constraints, respectively. AfÞnity estimations of ligands, to rank geometric poses and different ligands, are still a major problem, which is illustrated by recent comparisons of the performance of different scoring functions [130–132]. The combination of various virtual screening strategies and docking and scoring are now the most powerful tools in computer-aided design of new leads and their stepwise optimization [75–79].

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CONCLUSIONS What are the main problems in drug discovery? New strategies and technologies speed up drug discovery, once a target is deÞned. But is the target the right one? Will it work not only in animals but also in humans? Will it be druggable, that is, is there a possibility to modulate its activity by a small molecule? For most drug–drug interactions this is currently not the case. Is the focus on one target appropriate or should a chemical biology approach be followed to study the inßuence of compounds in the biological system, be it cells or animals? Currently, none of these questions can be answered in a sufÞcient manner. The discovery process itself—that is, the search for a lead structure and its optimization—is often straightforward and faster than it was in the past. But then again problems with unpredictable consequences arise. Will there be sufÞcient absorption (in humans), no Þrst-pass effect, no signiÞcant plasma protein binding, no interaction with the hERG channel, no CYP inhibition or induction, and no formation of toxic metabolites? Can animal toxicity results be extrapolated to humans? Will the drug itself or its metabolites have some rare side effects that cannot be tolerated, considering the therapeutic beneÞt? Can the drug be applied for longer time or even life-long? Several of these questions (e.g., bioavailability, hERG and CYP inhibition) are already being investigated in the optimization phase. But not all potential problems can be considered at the same time. It’s like a chess game. Black and white positions have been developed into a highly complex interaction; touch one Þgure and the whole strategy may break down. The same applies to a CNS-active drug. The compound may already have a favorable spectrum of activities against several targets, as well as some other favorable properties. You, as a medicinal chemist, try to improve a minor problem but after a slight modiÞcation of the chemical structure, all other properties change. There are certain rules about what to do when, for example, if bioavailability is too low and/or biological half-life is too short, then the introduction of ßuorine atoms in certain positions helps. The reverse, however, introduction of an aromatic methyl group, should be performed to achieve a signiÞcant decrease in biological half-life. Many other recommendations follow from medicinal chemistry experience: for example, how to improve BBB penetration, how to increase solubility, and how to reduce hERG channel toxicity. Unfortunately, in our time of high-throughput philosophy, there is the risk that this accumulated medicinal chemistry know-how [83] may fade away. Overreliance on technologies and computer methods worsens this situation. Poor ADME properties are often cited as the most common reason for failure in clinical development, creating a hype in the need for ADME prediction tools. However, this conclusion is based on old data [133] and is not even generally supported by these data [16, 134]. With the exception of antiinfectives, it was never a major reason for clinical failure, neither in the early period of 1964–1985 (7% attrition rate due to ADME, not including antiinfectives) [16, 134], nor in the years 1992–2002 (11% attrition rate due to inappropriate ADME) [12]. Bioavailability problems can now be minimized at a very early stage by applying

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Clinical Trials, Overview Curtis L. Meinert

Trial is from the Anglo–French trier, meaning to try. Broadly, it refers to the action or process of putting something to a test or proof. Clinical is from clinic, from the French cliniqu´e and from the Greek klinike, and refers to the practice of caring for the sick at the bedside. Hence, narrowly, a clinical trial is the action or process of putting something to a test or proof at the bedside of the sick. However, broadly it refers to any testing done on human beings for the sake of determining the value of a treatment for the sick or for preventing disease or sickness. The broad deÞnition of clinical trial includes deÞnitions allowing for use of the term in references to studies involving a single treatment (e.g., as in most Phase I trials and some Phase II drug trials) and for studies involving use of an external control (e.g., studies involving historical controls) (Meinert CL, 1996). However, use herein will be in the stricter sense of usage; that is, to refer to trials involving two or more treatment groups comprised of persons enrolled, treated, and followed over the exact same time frame. The treatment can be anything considered to hold promise in caring for the sick, in the prevention of disease, or in the maintenance of health. The term, in the context of a trial, refers to the experimental variable—the variable manipulated by the trialist. The variable may have just two states (e.g., as in a trial involving a single test treatment and single control treatment) or three or more states (e.g., as in a trial involving several different test treatments and one or more control treatments). The variable, in the case of drug trials, may serve to designate different drugs, different doses of the same drug, or different forms or Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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routes of administration of the same drug. In other contexts, it may variously refer to different kinds or forms of surgery, different kinds or forms of care or management regimens, different kinds or forms of diagnostic tests, different kinds or forms of medical devices, different kinds or forms of counseling regimens to achieve some desired end, or combinations of the above. The clinical trial, in its simplest form, involves the application of the experimental variable—treatment to a person or group of persons—and observation during or following application of the treatment to measure its effect. That measure (outcome measure) may be death, occurrence or recurrence of some morbid condition, or a difference indicative of change (e.g., difference in blood pressure measured for each person just prior to the start of treatment and again at some point during or after treatment). There is no way to “test” a treatment or to “prove” its effectiveness in the absence of some absolute or relative measure of success. Trials are said to be controlled if the effect of a treatment is measured against a comparison treatment administered over the same time period and under similar conditions. That comparison treatment may be another test treatment or, depending on circumstances, a control treatment consisting of an accepted standard form of therapy, a placebo or sham treatment, or observation only (no treatment). A trial is said to be uncontrolled if it does not have a comparison treatment or if the enrollment to and administration of the test and comparison treatments is not concurrent (e.g., as with use of historical controls for evaluation of a treatment). The Book of Daniel (Chapter 1, verses 12–15) provides an account of what amounts to an uncontrolled trial involving a diet of pulse—edible seeds of certain pod-bearing plants, such as peas and beans (American Bible Society, 1816): Prove thy servants, I beseech thee, ten days; and let them give us pulse to eat, and water to drink. Then let our countenances be looked upon before thee, and the countenance of the children that eat of the portion of the King’s meat: and as thou seest, deal with thy servants. So he consented to them in this matter, and proved them ten days. And at the end of ten days their countenances appeared fairer and fatter in ßesh than all the children which did eat the portion of the King’s meat. Fortuitous events can produce conditions reminiscent of the features of a trial. One such account is that given by Ambroise Par´e (surgeon, 1510–1590) during the battle in 1537 for the castle of Villaine. The treatment for gunshot wounds in Par´e’s time was boiling oil poured over the wound. Because of the intensity of the battle, Par´e ran out of oil and resorted to using an ointment made of egg yolks, oil of roses, and turpentine. The result of his “trial” is summarized by his observation the morning after the battle (Packard FR, 1921): I raised myself very early to visit them, when beyond my hope I found those to whom I had applied the digestive medicament, feeling but little pain, their wounds neither swollen nor inßamed, and

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having slept through the night. The others to whom I had applied the boiling oil were feverish with much pain and swelling about their wounds. Then I determined never again to burn thus so cruelly the poor wounded by arquebuses. Many of the essential elements of the modern day controlled trial are contained in Lind’s account of a trial performed aboard the Salisbury at sea in 1747 (Lind J, 1753): On the 20th of May 1747, I took twelve patients in the scurvy, on board the Salisbury at sea. Their cases were as similar as I could have them. They all in general had putrid gums, the spots and lassitude, with weakness of their knees. They lay together in one place, being a proper apartment for the sick in the fore-hold; and had one diet common to all, viz., watergruel sweetened with sugar in the morning; fresh mutton-broth often times for dinner; at other times puddings, boiled biscuit with sugar, etc; and for supper, barley and raisins, rice and current, sago and wine, or the like. Two of these were ordered each a quart of cyder a day. Two others took twenty-Þve gutts of elixir vitriol three times a day, upon an empty stomach; using a gargle strongly acidulated with it for their mouths. Two others took two spoonfuls of vinegar three times a day, upon an empty stomach; having their gruels and their other food well acidulated with it, as also the gargle for their mouth. Two of the worst patients, with the tendons in the ham rigid, (a symptom none of the rest had), were put under a course of seawater. Of this they drank half a pint every day, and sometimes more or less as it operated, by way of gentle physic. Two others had each two oranges and one lemon given them every day. These they eat with greediness, at different times, upon an empty stomach. They continued but six days under this course, having consumed the quantity that could be spared. The two remaining patients, took the bigness of a nutmeg three times a-day, of an electuary recommended by an hospital surgeon, made of garlic, mustard-seed, rad raphan, balsam of Peru, and gum myrrh; using for common drink, barley-water well acidulated with tamarinds; by a decoction of which, with the addition of cremor tartar, they were gently purged three or four times during the course. . . . the most sudden and visible good effects were perceived from the use of oranges and lemons, one of those who had taken them being at the end of six days Þt for duty. TREATMENT PROTOCOL The treatment protocol (the general term, study protocol or trial protocol has broader meaning and refers to the constellation of activities involved in conducting a trial) of the trial speciÞes the treatments being studied, the manner and

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method of usage and administration, and conditions under which other treatments are called for when needed for the well-being of those enrolled. The treatment may be administrated in one application or multiple applications. The period of treatment may be short (e.g., as in trials involving a single application of treatment such as surgery) or extended (e.g., as in trials involving the treatment of a chronic condition with drugs) over a period of weeks, months, or years. The treatment, in the case of drug trials, may involve a Þxed dose administered according to some schedule or dose titration in which each person ultimately receives the amount needed to achieve a desired effect (e.g., the amount of a hypoglycemic agent needed to bring blood glucose levels to within the normal range). Protocols for all research involving human beings are subject to review and approval by institutional review boards (IRBs) or ethics review boards (ERBs) before implementation and at periodic intervals thereafter until the research is Þnished. Therefore, investigators undertaking trials have the obligation and responsibility to obtain IRB or ERB review and approval prior to initiation of a trial, and to seek its review and approval prior to implementing amendments to the protocol of the trial. They also have a responsibility to inform IRBs and ERBs of record of any untoward events in the conduct of the trial and to report to such boards any conditions or events believed to change the risk–beneÞt ratio for persons enrolled into the trial or still to be enrolled. Only patients judged eligible (as determined by speciÞed eligibility criteria) may be enrolled, and among those, only those who consent to participate in the trial. Persons are under no obligation to enroll or to continue once enrolled, and must be so informed prior to being enrolled. A person must be informed, as well, of what is entailed by enrollment, of the risks and beneÞts that may accrue by enrollment, and of such matters and details that might cause a reasonable person to decline enrollment when so informed (e.g., that treatments are randomly assigned and that they will be administered in masked fashion). All trials involve data collection at various time points over the course of enrollment and follow-up of persons. The amount collected per person depends on the nature of the disease or condition being treated and on the nature of the treatment process implied by the study treatments being used. The requirement for repeated observation of a person, as a rule (except for trials done in hospital or other settings involving resident populations or in which enrollment, follow-up, and treatment is directed or managed by telephone or mail), obligates a person to a series of visits to the study site. Usually, the purpose of the Þrst visit or series of visits will be to determine eligibility, collect necessary baseline data, obtain consent, and initiate treatment. Visits thereafter will be to Þne-tune or continue treatment and to collect necessary follow-up data. The schedule of follow-up visits will be timed from the point of randomization or initiation of treatment and, as a rule, will be on a deÞned time schedule (e.g., once every week) with provisions for interim (unscheduled) visits when necessary for the care of those enrolled. Comparison of the different treatments tested for effect is done in different ways depending on the outcome measures used to assess effect. The comparison,

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in the case of an event, such as death or occurrence of a morbid event, will be based on the event rate (or the raw percentage of persons experiencing the event) as seen for the different treatment groups. In the case of a continuous variable, such as weight or blood pressure, the change from entry to some deÞned point after enrollment will be determined for each person studied and then summarized in some fashion (e.g., by calculating a mean or median). The treatment effect will be estimated by the difference obtained by subtracting the summary measure for the comparison treatment from the indicated test treatment. Judging the safety or efÞcacy of a treatment is problematic in trials not involving a designed comparison group—often the case in Phase I, II, and I/II trials (see below for deÞnitions). The problem is compounded by the typically short duration and small size of these trials. The problem is most acute in the testing of drugs in people having a life-threatening disease when the drugs themselves carry their own morbidity and increased risk of death. Are the morbid events observed the result of the disease or the drug? Even deaths become difÞcult to interpret in the presence of a high background death rate from the disease. Was a death the natural outcome of the disease, or was it induced by the treatment? The issue is rarely clear until sufÞcient information has accumulated to cause one to discount natural causes as the likely explanation, or to allow one to recognize an unusual clustering of deaths and morbid events, as with the case of a trial of Þaluridine (FIAU) (McKenzie R, 1995).

CLASSES OF TRIALS Most clinical trials involve parallel treatment designs, that is, designs where an assignment unit (usually a person) is assigned to receive only one of the treatments under study. The word parallel indicates that two or more groups of assignment units are proceeding through the trial side by side, with the only ostensible difference (other than baseline differences in the composition of the groups) being the treatment administered. The goal in trials with parallel treatment designs is for each person enrolled to receive the assigned treatment and to have no exposure to any of the other treatments under study in the trial (except where the requirements for proper care are overriding and make such exposure necessary). The assignment unit (randomization unit in randomized trials), in the case of parallel treatment designs, is usually a person but can be an aggregate of persons (e.g., members of the same household) or a subpart of a person (e.g., an eye, as in the Glaucoma Laser Trial Research Group, 1991). The treatment design in crossover trials is different. In this class of designs a person or treatment unit receives two or more study treatments in a speciÞed order. Crossover trials are classiÞed by the number of treatments to be administered to a person or treatment unit and by whether a given person or treatment unit receives all (complete or full crossover) or just some (partial or incomplete crossover) of the study treatments. For example, a two-period crossover design

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is one in which each person or treatment unit receives two study treatments in some order, usually random. An n-way crossover design is one in which a person or treatment unit receives n of the treatments represented in the design. The utility of crossover designs is limited to settings in which it is feasible to administer different treatments to the same person or treatment unit, each for a short period of time, and in which it is possible to measure the effect at the end of each treatment period. They are not useful in settings in which the outcome of interest is a clinical event that can occur at any time after enrollment. In a trial with a parallel treatment design, assignment determines the treatment to be administered (except to the extent that other treatments are needed for proper care) whereas, in a crossover trial, assignment determines the order of treatments to be used. Typically, each treatment is administered for a designated period of time (e.g., 4 weeks). Often the last administration of one treatment and the Þrst administration of the next treatment are separated in time (e.g., 1 week) to allow the effect of the preceding treatment to “wear off” (“washout period”) before administering the next treatment. Imagine a trial involving three study treatments (A, B, and C) with the same (uniform) assignment probabilities and 54 people. In a trial with a parallel treatment design, 18 people would be assigned to receive treatment A, 18 would be assigned to receive treatment B, and 18 would be assigned to receive treatment C. In a trial involving a complete (full) crossover of treatments, each of the 54 people would receive treatments A, B, and C. Assuming treatments are arranged in all possible orderings, there would be six different orderings of the treatments (ABC, ACB, BAC, BCA, CAB, and CBA), and nine patients would be randomly assigned to receive a given ordering. While the goal of the two designs is the same, to Þnd the most effective treatment, the methodology differs. With the parallel treatment design, the treatment is evaluated in comparable groups of treatment units (usually persons), and with the crossover treatment design, the treatment effect is evaluated within the same treatment unit (usually a person). Trials involving parallel treatment designs are of two general types with regard to sample size design—Þxed or sequential. The majority are of the Þxed type. That is, the sample size is speciÞed at the outset, as determined by pragmatic considerations (e.g., by the amount of money available for the trial) or by a formal sample size calculation. Trials are considered to have a Þxed sample size even if they do not proceed to the desired sample size (e.g., are stopped early because of a treatment difference). The sample size is Þxed in the sense that the intent is to enroll and follow the speciÞed number of assignment units unless indicated otherwise by events transpiring during the course of the trial. In sequential trials (also of two types—open and closed), enrollment and observation continue until a stopping boundary, constructed for the outcome of primary interest (usually a binary “success” or “failure” type event), is crossed. Open sequential designs involve two boundaries, one indicative of superiority and the other indicative of inferiority of a test treatment relative to a com-

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parison treatment. Enrollment continues until the observation function for the outcome measure of interest crosses one of the two boundaries. The design has the advantage of providing a test of the null treatment hypothesis for given type I and II error levels that, on average, requires a smaller sample size than that for a Þxed sample size design. However, the actual sample size required for a boundary crossing can be larger (in theory, sometimes much larger) than that for a Þxed sample size design. The possibility of the Þnal sample size being much larger is ruled out with the closed sequential design. That design, in addition to the two boundaries mentioned above, involves a third boundary serving to place an upper bound on enrollment. If that boundary is crossed, because neither of the other two boundaries is crossed (signifying a difference in favor of one of the treatments), then the treatments being compared are considered to be of equivalent value as measured by the outcome observation function. Sequential designs have limited utility in the context of clinical trials, partly because they require rigid adherence to a stopping rule. Use is limited to instances where the “success” or “failure” of a treatment can be determined shortly after administration. They are not useful in settings involving long-term treatment and with outcome measures requiring weeks, months, or years of observation. In general, more ßexible methods of monitoring trials are more appropriate.

DRUG TRIALS Compounds, no matter how promising or impassioned the pleas for use, have to go through a series of tests in animals before they can be tested in humans. Those considered to lack promise after animal testing do not come to testing in humans. Typically, the testing in humans is done in a time-ordered sequence, as suggested by the phase label afÞxed to trials as deÞned below. However, in truth, adjoining phases overlap in purpose. Hence, the label, at best, serves only as a rough indicator of the stage of testing, especially when, as is often the case, drug sponsors, at any given point in time, may have several trials underway carrying different phase labels. The deÞnitions of the different phase labels follow: Phase I : Usually the Þrst stage of testing performed in anticipation of an Investigational New Drug Application (INDA or NDA); done to generate preliminary information on the chemical action and safety of the indicated drug and to Þnd a safe dose; usually not randomized. Phase II : Usually the second stage of testing; generally carried out on persons having the disease or condition of interest; done to provide preliminary information on efÞcacy of the drug and additional information on safety; may be designed to include a control treatment and random assignment of patients to treatment.

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Phase I/II : A trial having some of the features of Phase I and II trials; designed to provide preliminary information on safety and efÞcacy. Phase III : Usually the third and Þnal stage in testing, prior to submission of an NDA; concerned with assessment of dosage effects, efÞcacy, and safety; usually designed to include a control treatment and random assignment to treatment. When the test is completed (or nearly completed), the drug manufacturer or sponsor may request permission to market the drug for the indication covered in the testing by submission of an NDA. Phase II/III : A trial having some of the features of Phase II and III trials; designed to provide information on safety and efÞcacy. Phase IV : A fourth stage of testing, sometimes carried out. Usually controlled and performed after approval of the NDA. Typically done under circumstances approximating real-world conditions; usually has a clinical event as a basis for sample-size calculation and provides for extended treatment (where appropriate) and long-term follow-up, with efÞcacy and safety of the drug being measured against a control treatment. Drugs, after marketing approval, remain under surveillance for serious adverse effects. The surveillance—broadly referred to as postmarketing surveillance —involves the collection of reports of adverse events via systematic reporting schemes and via sample surveys and observational studies. Sample size tends to increase with the phase of the trial. Phase I and II trials are likely to have sample sizes in the 10s or low 100s compared to 100s or 1,000s for Phase III and IV trials. The focus shifts with phase. The aim in the early phases of testing is to determine whether the drug is safe enough to justify further testing in human beings. The emphasis is on determining the toxicity proÞle of the drug and on Þnding a proper, therapeutically effective dose for use in subsequent testing. The Þrst trials, as a rule, are uncontrolled (i.e., do not involve a concurrently observed, randomized, control-treated group), of short duration (i.e., the period of treatment and follow-up is short), and conducted to Þnd a suitable dose (usually via some traditional or Bayesian dose escalation design) for use in subsequent phases of testing. Trials in the later phases of testing, for the most part, involve traditional parallel treatment designs, randomization of patients to study treatments, a period of treatment typical for the condition being treated, and a period of follow-up extending over the period of treatment and beyond. Most drug trials are done under an investigational new drug application (INDA or IND) held by the sponsor of the drug. The “sponsor” in the vernacular of the Food and Drug Administration (FDA) is typically a drug company, but can be a person or agency without “sponsorship” interests in the drug. Regulations require investigators to report adverse events to the FDA. The general guidelines regarding consent are similar, but not identical, to those promulgated by the OfÞce for the Protection from Research Risks (OPRR) for IRBs.

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RANDOMIZED TRIAL A randomized trial is a trial having a parallel treatment design in which treatment assignment for persons (treatment units) enrolled is determined by a randomization process similar to coin ßips or tossings of a die. The trialist’s purpose in randomization is to avoid selection bias in the formation of the treatment groups. The bias is avoided because the treatment to which a person is assigned is determined by a process not subject to control or inßuence of the person being enrolled or those responsible for recruiting and enrolling the person. The comparison of one group to another for treatment effect will be biased if, for whatever the reason, one group is “healthier” or “sicker” on entry than the other. Schemes in which one knows or can predict treatment assignments in advance of issue are open to such bias. Clearly, that is the case with assignment schemes posted in a clinic and open for all to see prior to issue. The bias is likely as well with systematic schemes, such as those in which every other person is assigned to the test treatment or in which persons seen on odd-numbered days receive the test treatment and those seen on even-numbered days receive the control treatment. The goal is to create groups that provide a valid basis for comparison. To achieve that end one has to ensure that the groups are similar (within the range of chance) and to avoid bias in the assignment process. The usual method for achieving both ends is randomization. Randomization does not guarantee comparability of the treatment groups with regard to the various entry characteristics of interest. Indeed, one can, by chance, have differences among the treatment groups. A large difference (one yielding a small P value) can arise by chance and, hence, cannot be taken as prima facie evidence of a “breakdown” (e.g., “peeking” or other purposeful acts aimed at determining assignment before issue) of the randomization process, unless supported by other evidence of a “breakdown.” The hallmarks of a sound system of randomization are: reproducible order of assignment; documentation of methods for generation and administration of assignments; release of assignments only after essential conditions are satisÞed (e.g., only after a person has been judged eligible and has consented to enrollment); masking of assignments to all concerned until needed; inability to predict future assignments from past assignments; clear audit trial for assignments; and the ability to detect departures from established procedures (Meinert CL and Tonascia S, 1986). The randomization may be simple (complete) or restricted. The purpose of restriction is to force the assignments to satisfy the speciÞed assignment ratio at intervals during enrollment. Those restrictions are typically referred to as blocking. For example, suppose a trial involves two treatments, A and B, and the desired assignment ratio is one-to-one. A simple (unrestricted) randomization scheme would involve the equivalent of repeated ßips of an unbiased coin with a head leading to assignment to treatment A and a tail to treatment B. The design would, on average, yield the desired assignment ratio, but allows for wide departures from the desired mix, depending on the “luck” of the ßips.

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33

If such departures are of concern, then the randomization scheme can be restricted by blocking so as to ensure the desired mix after a speciÞed number of assignments. For example, imposition of a blocking requirement after every eighth assignment would have the effect of “forcing” the randomization to yield the desired mix of one-to-one after every eighth assignment. The purpose of the blocking is to ensure a near desired assignment ratio so as to protect treatment comparisons against secular trends in the mix of patients as the trial proceeds. The randomization also may be stratiÞed. The purpose of stratiÞcation is to provide treatment groups comprised of persons or treatment units having identical (within the limits of the stratiÞcation) distributions of the stratiÞcation variable. It is useful only insofar as the variable used for stratiÞcation serves to inßuence or moderate the outcome of interest. The stratiÞcation has the effect of “controlling” the inßuence of the stratiÞcation variable on outcome by ensuring the same distribution of the variable across the different treatment groups. For example, suppose one wishes to stratify on gender in the trial described above (because, perhaps, of a belief that the treatment effect will be different in women than in men). The stratiÞcation would be achieved by creating two randomization schedules, each with a one-to-one assignment ratio and with blocking to satisfy the assignment ratio after enrollment of the 8th, 16th, 24th, and so on, person in each stratum. The effect of the stratiÞcation would be to ensure the same gender mix (within the limits of the blocking) for the two treatment groups, regardless of the underlying gender mix of the population to be studied. For example, suppose 96 patients are to be enrolled from a population with a 1:2 mix of males to females. In that case, one would expect to enroll 32 males and 64 females, and to have 16 males and 32 females in each treatment group. If the underlying mix is one-to-one, then there would be 24 males and 24 females in each of the two treatment groups. Clearly, the number of variables that can be controlled by stratiÞcation is limited. The more variables, the more subgroups for randomization and the less useful the process is as a reliable means of variance control (Grizzle JE, 1982). In addition, there are logistic difÞculties associated with use of variables whose values have to be determined by performing laboratory tests or other diagnostic procedures during the enrollment process. Even if one stratiÞes on a few selected variables, other variables may well be considered to be important determinants of outcome. Hence, the experienced trialist strives to “remove” the effect of such differences via analysis procedures, for example, by assessing the treatment effect within deÞned subgroups (subgroup analysis); or by providing estimates of treatment effect that are adjusted for differences in the distribution of important demographic or baseline variables via regression procedures (University Group Diabetes Program Research Group, 1970b). MASKING Masking is the purposeful concealment of some fact or condition and is done to keep knowledge of that fact or condition from inßuencing the behavior,

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observation, or reporting of persons so masked. Masking, in the context of trials, is imposed to reduce the likelihood of a treatment-related bias due to knowledge of treatment assignment. That bias, after a person is enrolled, occurs whenever knowledge of that person’s treatment assignment serves to color the way he or she is treated, followed, or observed. One way of reducing it is by masked treatment administration. In one form of such administration (single-masked), only one member of the subject—treater pair is masked to treatment, usually the subject. Another form of masking is one in which both members of the pair are masked—doublemasked treatment administration. As a rule, double-masked treatment administration means that all persons in a clinic are masked and, therefore, that those responsible for data collection and generation are masked to treatment as well. Generally, it is not possible or prudent to mask treatment administration in trials involving treatments requiring different routes or modes of administration (e.g., as in a trial involving a medical vs. a surgical form of treatment), where knowledge of treatment assignment is part of the effect being tested (e.g., as in trials aimed at modiÞcation of one’s eating habits via different modes of dietary consulting), or where the masking carries risks for those enrolled. Therefore, the opportunities for double-masked treatment administration are limited largely to trials of drugs considered safe and that are reasonably free of side effects and that can be administered at Þxed dose levels. It is usually not wise or practical to administer treatments in a double-masked fashion when treatment doses are to be titrated to achieve desired effects. Masked treatment administration has been used as a mark of “quality” for trials. There is, therefore, a tendency to view results from masked trials as more reliable than those from unmasked trials. In truth, however, masked treatment administration is rarely 100% effective. All forms of treatment, and especially those involving drugs, produce side effects and tell-tale signs that may serve to unmask treatment. Hence, the protection provided by masking can be illusory. As a result, it is better to make assessments of “quality” in terms of the risk of treatment-related bias and the likely effect of such bias, if present, on the results reported. The risk of treatment-related bias is low for “hard” outcome measures and with explicitly deÞned treatment protocols, even in the absence of masked treatment administration. The second line of defense, in the absence of double-masked treatment administration, is to mask as many groups of persons involved in the trial as is possible within the limits of practicality and safety. Hence, even if it is not possible to mask patients or those who treat them, it may be possible to mask those responsible for data collection or data generation (e.g., as with an arrangement as in the Glaucoma Laser Trial Research Group (1991), where intraocular pressure was measured by masked readers, or as with laboratory personnel or readers of X-rays, ECGs, or fundus photographs masked to treatment assignment). With or without treatment masking, trialists strive for objectively deÞned treatment and data collection procedures and for outcome measures as free from observer or respondent bias as is humanly possible. In addition, they are inclined

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toward continuing effort over the course of a trial aimed at maintaining the training and certiÞcation of study personnel in regard to required study procedures, and toward establishing and maintaining standards of performance via ongoing monitoring and quality control surveillance.

ANALYSIS The protection provided against treatment-related bias by the assignment process is futile if the analysis is biased. Treatment comparisons, to be valid, must be based on analyses that are consistent with the design used to generate them. In the case of the randomized trial, this means that the primary analyses of the outcomes of interest must be by assigned treatment (also known as analysis by intention-to-treat). It means, for example, that observations relating to a morbid event are counted to a patient’s assigned treatment regardless of whether or not the patient was still on the assigned treatment when the event occurred. Analyses involving arrangements of data related to treatment administered may be performed, but only as supplements to the primary analyses. They should not and cannot serve as replacements for those analyses. Analyses by treatment assignment, as a rule, serve to underestimate the treatment effect. Usually, analyses in which the requirement is relaxed will yield a larger estimate of the treatment difference than seen when evaluated under the intention-to-treat mode of analysis (e.g., as in the case of the University Group Diabetes Program (UGDP ) trial) (University Group Diabetes Program Research Group, 1970b). Designs allowing for termination of data collection when a person can no longer receive or be maintained on the assigned treatment are open to treatmentrelated bias. The goals of the primary analyses cannot be met when data collection for a person ceases when that person experiences a nonfatal “endpoint” or when the person’s treatment is stopped or changed. The analysis requirement implies continued follow-up of all persons enrolled into a trial to the scheduled close of follow-up regardless of their treatment or outcome status.

MONITORING TREATMENT EFFECTS The randomized trial depends on a state of equipoise—a state of legitimate doubt regarding the test treatment relative to the control treatment(s) (Beecher HK, 1966; Freedman B, 1987; Lilford RJ and Jackson J, 1995). It cannot be undertaken without a proper ethical climate characterized by such a state of doubt. It does not matter whether that state has been dispelled by observation and data, by declaration, or in other ways. For example, it would not be possible to assess the value of coronary care units for persons appearing to be having a myocardial infarction (MI), even if their value has not been demonstrated by controlled trials. They are considered to be required for good care and, hence, the window

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of opportunity for testing via designed randomized trials has closed. Once closed, it may remain closed, or may open again years later if people start questioning the merits of the treatment. When the oral hypoglycemic agents appeared on the scene in the early 1950s, they were widely regarded as safe and effective and, hence, became a part of the armamentarium for care of the adult-onset diabetic. However, doubts raised in the late 1950s as to their value led to a climate of doubt suitable for initiation of the UGDP trial (University Group Diabetes Program Research Group, 1970a). Trials are done because of the prospect of beneÞt associated with a new treatment, or to test the efÞcacy of an existing treatment. They are not undertaken to prove a treatment to be useless or harmful. Indeed, a trialist is obligated to stop a trial prior to its scheduled completion if the accumulated data indicate that the treatment of interest is inferior to the control or comparison treatment. In fact, some argue that there is an obligation to stop if it becomes clear that the test treatment is no better than the comparison treatment, even if one is uncertain whether it is harmful. Hence, for example, investigators in the UGDP opted to stop use of tolbutamide in that trial once they were certain it was no better than the control treatment—the usual antidiabetic dietary recommendations and placebo medication. The need for ongoing monitoring exists for any trial in which the treatments carry risk of harm, and in which it is possible to reduce that risk by timely monitoring. That need makes it necessary for the trialist to aim for an orderly and timely ßow of data from the site of generation or collection to the processing and analysis site. Clearly, the best systems in this regard are those having realtime or near-real-time ßows (e.g., as with systems requiring transmission of data related to a patient visit on completion of the visit or on occurrence of an outcome of interest). Typically, treatment effects monitoring is entrusted to a group of people that together have the necessary skills and expertise to monitor effectively (Hawkins BS, 1991; Meinert CL and Tonascia S, 1986; NIH Clinical Trials Committee, 1979; Wittes J, 1993). The group is usually comprised of 5–12 people with expertise in the disease under treatment, in the design, conduct, and analysis of clinical trials, or in other specialty areas. When the group comprises a mix of people from within the trial (e.g., the ofÞcers of the trial, such as the chair and vice chair, the director of the coordinating center, etc.) and outside the trial, the votes concerning recommendations for change, generally, are vested in those outside the study. The restriction is imposed, typically, because of concerns that persons associated with the trial may have conßicts of interest that could serve to inßuence their votes (Chalmers TC and Amacher P, 1982). Monitoring proceeds under different constructs, depending on the philosophy of those doing the monitoring. Some constructs require stopping rules and restrictions on the type of data that may be monitored and the number of interim “looks” that can be made in relation to the monitoring. Other groups consider such restrictions unnecessary and rely instead on the collective judgment of the monitoring group (Ellenberg S, 1993).

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The OfÞce for Protection from Research Risks (OPRR) (an ofÞce within the National Institutes of Health responsible for the promulgation and administration of regulations regarding institutional review boards) and the set of rules relating to research on human beings obligates IRBs to be satisÞed that risk to subjects is minimized. As part of this assurance in regard to clinical trials, investigators must have “adequate provision for monitoring the data collected to ensure the safety of subjects” (Section 46.111) (OfÞce for Protection from Research Risks, 1991) and must provide participants with information on . . . “signiÞcant new Þndings developed during the course of the research which may relate to the subject’s willingness to continue participation will be provided to the subject” (Section 46.116) (OfÞce for Protection from Research Risks, 1991). This requirement makes it necessary to inform patients of results during the trial that bear on their willingness to continue. This requirement pertains to information from inside or outside the study, if the information is likely to cause patients to reconsider their decision to be enrolled in the trial. Formal reconsent procedures may be required if the treatment effects monitoring committee recommends changes to the treatment protocol (e.g., as discussed in Meinert CL and Tonascia, 1986).

REPRESENTATIVENESS, VALIDITY, AND GENERALIZABILITY Representativeness, in the context of a trial, refers to the degree or extent to which those enrolled can be considered representative of the general population of persons to whom the treatment may be applied, if shown to be useful. Validity, in the context of a treatment difference, refers to the extent to which that difference can be reasonably attributed to treatment assignment. Generalizability refers to the degree to which the Þndings of the trial can be extended to the general population of eligible persons. The concepts of validity and generalizability are different. Validity derives from the design of the trial and from the way it is carried out, whereas generalizability is largely a matter of judgment. A treatment comparison is valid if it is based on comparable groups of persons treated and observed in such a way so as to make treatment assignment the most likely explanation of the result observed. “Representativeness” is deduced by comparison of the demographic and other host characteristics of the study population to that of the general population of eligible persons (or by comparison with all persons screened for enrollment). The desire for representativeness arises from the belief that conclusions from a trial will be strengthened by having a broadly “representative” study population. The drive for demographic representativeness has been propelled in recent years by the belief that women and persons of ethnic minorities have been “underrepresented” or “understudied” relative to men and the prevailing ethnic majority in trials and other areas of clinical research. Those concerns have been sufÞcient to cause the US Congress, in the NIH Revitalization Act of 1993, to impose requirements on trials aimed at ensuring adequate numbers of women and ethnic minorities to determine whether the treatments being studied in a trial work

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differently in men than in women or in an ethnic minority than in the ethnic majority (United States Congress, 1993). There is no way to ensure “representativeness” in the absence of a sampling frame for the eligible study population and a related sampling scheme aimed at providing a representative sample of that population. However, even if one were able to develop a sampling frame (usually impossible because to do so one would have to screen the general population to identify persons eligible for study), the population ultimately enrolled, even if selected by sampling, would, at best, be representative only of those able and willing to be enrolled, because of the requirements of consent. Hence, trials, by nature of their design, involve select, nonrepresentative populations. Even if a treatment is found effective in a trial, one has no direct way of knowing if it would be effective for those patients not agreeing to be studied. If the issues of consent and lack of a sampling frame were overcome, then one would still be left with the fact that most clinics, for practical and ethical reasons, have to rely on those who come to them. They do not have the ability or moral authority to go and seek out suitable patients for study, especially if doing so means that those who routinely come to them would be turned away. Such a “selective” approach would be viewed as violating the principle of justice as set forth in the Belmont Report (National Commission for the Protection of Human Subjects of Biomedical and Behavioral Research, 1979). That one needs to generalize is obvious. The need arises in regard to the route of treatment, amount of treatment, type of treatment, and type of patients. For example, if a trial involved a single Þxed dose of a drug and failed to Þnd a difference (e.g., as in the UGDP trial, regarding tolbutamide) (University Group Diabetes Program Research Group, 1970b) does one conclude that use of the same drug, under a different, more ßexible dosing scheme would produce a more favorable result? Similarly, if one compound produces a beneÞt, does one conclude that other sister compounds will show the same effect? Or conversely, if one member of a drug family has a bad effect (e.g., Þaluridine) or fails to show a beneÞt (e.g., tolbutamide), does one shy away from other related compounds? Also, if a trial involves mildly diseased people and shows a beneÞcial effect for the test treatment, does one conclude that the test treatment will have a similar effect in sicker people? Last, if the drug tested works for the disease or condition being treated, is it not likely that it would be useful as well for a related condition or disease? So-called “off label” use (from the fact that drugs are approved for designated indications) accounts for a large number of treatment prescriptions (Brosgart CL, 1996; Grossman E, 1996; US General Accounting OfÞce, 1996; Winker MA, 1996). Whenever one generalizes, whatever the nature or direction, one is in effect answering one or more of the above questions. If as a treater, one chooses to use a sister compound of a drug shown to be ineffective in a trial, then one is in effect saying that the result from the trial, for whatever reason, is not generalizable. Generalizations depend on judgments regarding the trials and on prior beliefs regarding the treatment in question.

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39

A trial can provide a valid basis for comparing one treatment to another if the differences in outcomes for the treatment groups being compared can be attributed reasonably to treatment. The general “laws of science” and “principles of parsimony” require that one defaults to the simplest explanation—usually the one requiring the fewest assumptions. Hence, in the case of the trial in which treatments are selected by the patient or physician, one is as a rule more inclined to attribute the difference to selection factors than to the test treatment. By the same principle, one should be more inclined to attribute a treatment difference to bias on the part of the observer rather than to the treatment when the opportunity for such bias exists. The degree of “reasonableness” of such an explanation will depend on the nature of the outcomes and whether one can reasonably ascribe it to biased observation. It becomes progressively more difÞcult to do so, even if the observer is not masked, the “harder” the outcome measure. For example, it is not reasonable to expect that one’s opinion regarding the merits of a treatment will inßuence one’s ability to report reliably whether a person is alive or dead, but such opinion may inßuence how one sees or reports on a person’s quality of life. There is a responsibility on the part of trialists to rule out other lesser explanations of results before ascribing them to treatment. Contrary to lay perceptions, trials and comparisons of treatments within the trial are made robust to selection bias and the consequences of “nonrepresentative” study populations by randomization. The assessment of treatment effect is achieved by having comparable groups of patients in the different treatment groups and by having procedures for observing and following patients that are independent of treatment assignment. The comparison is valid regardless of the study population and provides information on the relative value of one treatment to another. Hence, from the perspective of the trialist, it is far more important to have comparable treatment groups than to have “representative” treatment groups. The drive for “representativeness,” while perhaps of some social value, does little to make generalizations less risky or to increase the validity of trials. There are sound practical reasons to design trials with as few exclusions to enrollment as possible. The fewer the restrictions the easier and faster it is to recruit. Any effort to make them more “representative” by selective recruitment and enrollment will make them more costly and will increase the time required to enroll them. The imposition of recruitment quotas to achieve a desired sample size for gender, age, and ethnic origin groups poses a far more complicated and costly recruitment effort than one involving the enrollment of all comers regardless of gender, age, or ethnic origin. The goal of the trialist should be to strive for demographic neutrality in enrollment: that is, the trialist should not exclude potential participants on the basis of gender, ethnic origin, or age unless justiÞed on scientiÞc grounds. ScientiÞc grounds include the knowledge or expectation of a qualitative treatment by demographic interaction (i.e., where treatment is believed to be beneÞcial for one demographic group and harmful for another).

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Another reason for exclusion is contraindication of treatment in a particular demographic group. If any one treatment is contraindicated in a trial involving multiple treatments, then the restriction has to apply to all treatments. For example, this requirement was one of the reasons why the Coronary Drug Project (CDP) involved only men. Two of the Þve test treatments in the trial could not have been administered to premenopausal women; thus this demographic group could not be included without making the trial much more complex (Coronary Drug Project Research Group, 1973). As a rule, an anticipated low number in a speciÞed demographic group is not a reason to exclude. Disease and extent of disease are much more likely to affect the response to treatment than are “demographic” characteristics. Analyses of treatment effects across the various demographic subgroups represented in a trial can help determine whether there are treatment by demographic interactions. In general, interactions, when noted in the context of treatment trials, are more likely to relate to disease characteristics than to demographics (Freedman LS, 1995; Holbrook JT, 1994; Institute of Medicine, Committee on the Ethical and Legal Issues Relating to the Inclusion of Women in Clinical Studies, 1994a,b; Meinert CL, 1995). The mind-set regarding selection is different in prevention trials, where the goal is to determine whether a proposed prevention strategy works. One has to Þnd a population suitable for testing the proposed strategy. Hence, unlike the treatment trial, risk factors predisposing to a disease and risk of an event are important. In this setting, one has to pay attention to both factors in trying to design a cost-effective trial. Considerations of this sort led, for example, the designers of MRFIT (Multiple Risk Factor Intervention Trial Research Group, 1977) to exclude females from enrollment. The risk factors targeted (high blood pressure, high cholesterol, and smoking) occur less frequently in women than in men. Consequently, the effort required to Þnd women for study would have been much greater than that required to Þnd men. Furthermore, for the age range studied, women have a markedly lower myocardial infarction rate (the outcome of primary interest) than do men. This lower event rate would have meant that the planned sample size with women included would have to have been considerably larger to detect the same relative difference at the power level speciÞed for the trial. As it was, the trial required a sample size of 12,866 men.

READINGS The literature on the design, conduct, and analysis of clinical trials is ever expanding. Students of trials need to monitor the literature of reported trials as they appear in medical journals and to read specialty journals, such as Biometrics, Statistics in Medicine, Controlled Clinical Trials, Applied Clinical Trials, and Statistical Methods in Medical Research. The list of citations given in Table 1 is but a snapshot of selected references dealing with the methods and procedures of trials.

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41

TABLE 1. References on Methods and Procedures of Clinical Trials Topic

References

Specialty journals Applied Clinical Trials (Applied Clinical Trials, 1992–1997) Controlled Clinical Trials (Hawkins BS, 1991) Statistical Methods in Medical Research (Statistical Methods in Medical Research, 1992–1996) Statistics in Medicine (Colton T, 1991a,b) Textbooks Clinical trials (Buyse ME, 1984; Friedman LM, 1985; Hulley SB, 1988; Johnson FN and Johnson S, 1977; Meinert CL and Tonascia S, 1986; Pocock SJ, 1983; Smith PG and Morrow RH, 1991) Data analysis (Hedges L and Olken I, 1987; Hoaglin DC, 1983) Ethics (Katz J, 1972; Levine RJ, 1986) History (Stigler SM, 1986) Dictionaries/Encyclopedias Clinical trials (Meinert CL, 1996) Epidemiology (Last JM, 1995) Statistics (Kotz S, 1982–1988) Journal articles Analysis (Colton T, 1989; Cox DR, 1972; D’Agostino RB, 1992; Kaplan EL and Meier P, 1958; Peto R, 1977; Souhami RL and Whitehead J, 1994) Bayesian methods (Berry DA, 1985; Cornfield J, 1969; Freedman LS and Spiegelhalter DS, 1989; Spiegelhalter DS and Freedman LS, 1986) Cost and efficiency (Wittes J, 1990) Design (Colton T, 1989) Equipoise (Beecher HK, 1966; Freedman B, 1987; Lilford RJ and Jackson J, 1995; Pocock SJ, 1993) Ethics (Ashby D, 1993; Beecher HK, 1966; National Commission for the Protection of Human Subjects of Biomedical and Behavioral Research, 1979) Forms design and data management (Hawkins BS, 1995; Singer SW and Meinert CL, 1995; Wright P and Haybittle J, 1979a–c) History (Bull JP, 1959; Lind J, 1753) Meta-analysis and overviews (Berlin JA, 1989; Boissel JP 1989; Buyse M and Ryan LM, 1987; Hedges L and Olken I, 1987; Jones DR, 1995; Lee YJ, 1996; Stewart LA and Clarke MJ, 1995; Yusuf S, 1987) Philosophy (Sackett DL and Gent M, 1979; Schwartz D and Lellouch J, 1967) Randomization and stratification (Brown BW Jr, 1980; Grizzle JE, 1982; Kalish LA and Begg CB, 1985; Lachin JM, 1988; Meier P, 1981; Peto R, 1976; Senn SJ, 1989; Wei LJ, 1986) Sample size (Blackwelder WC, 1982; Blackwelder WC and Chang MA, 1984; Bristol D, 1989; Donner A, 1984; Lachin JM, 1981; Lachin JM and Foulkes MA, 1986; Lakatos E, 1988; Rubinstein LV, 1981)

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TABLE 1. (continued) Topic Subgroup analyses Treatment effects monitoring

References (Berry DA, 1985; Coronary Drug Project Research Group, 1980; Yusufs, 1991) (Ashby D. 1993; Canner PL, 1977; DeMets DL, 1987; Ellenberg S, 1993; Fleming TR and DeMets DL, 1993; Hawkins BS, 1991; Hughes MD and Pocock SJ, 1988; Kiri A and Meinert CL, 1995; Lee YJ, 1996; Pocock SJ, 1993)

Source: Susan Tonascia, ScM, Johns Hopkins School of Public Health, Department of Epidemiology.

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Aggressive Non-Hodgkin’s Lymphomas

ETIOLOGY AND PATHOPHYSIOLOGY Overview Lymphoma is a general term for cancers of the lymphatic system. The two main groups of lymphoma are Hodgkin’s disease (HD) and non-Hodgkin’s lymphomas (NHLs). The latter term encompasses a heterogeneous set of lymphomas grouped only by their lack of the giant histocytes (Reed-Sternberg cells) that characterize HD. ClassiÞcation of NHLs is extremely complex and has undergone extensive revisions in recent years, incorporating morphological, immunophenotypical, genetic, and clinical characteristics. This diverse set of diseases is characterized by varied pathogeneses and clinical outcomes that make the management of NHLs clinically challenging. NHLs are a disease of the elderly and are more common in males than females. Incidence increases with age and peaks between 80 and 85; the median age at diagnosis is 65. NHLs can be considered broadly as indolent or aggressive. This report focuses on the aggressive NHLs, which account for approximately 60% of NHLs in the seven markets covered. Pathophysiology The lymphatic system, an integral part of the body’s immune system, comprises a network of vessels (lymph ducts) in which a milky-colored ßuid (lymph) circulates. Hundreds of tiny, bean-shaped lymph nodes are present along this network; Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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they are found in clusters in the axilla (underarms), groin, neck, and abdomen. The bone marrow, spleen, tonsils, and thymus—a small gland behind the upper sternum—are also associated with the lymphatic system. Nonlymphatic organs that contain lymph tissues and can therefore be affected by lymphoma include the skin, lungs, liver, kidneys, stomach, and intestines. Lymph is made up primarily of lymphocytes. The two major types of lymphocytes are B lymphocytes (known as B cells), which produce antibodies, and T lymphocytes (known as T cells), which attack and destroy foreign antigens. Both types of lymphocytes originate from hematopoietic stem cells in the bone marrow and differentiate through a series of complex—and incompletely understood—interactions with growth factors, cytokines, and other cells as they develop. B cells leave the bone marrow and circulate throughout the blood and lymph systems as small resting cells until they encounter a foreign pathogen. When activated by such a pathogen, they travel to lymph nodes, where they undergo a period of intense proliferation in lymph node follicles. During this time, they adapt so that they are more speciÞc for the pathogen and can efÞciently clear the host of infection. The B cells leave the lymph nodes and begin another phase of recirculation to sites of infection and through the blood, bone marrow, spleen, and lymph systems. T cells leave the bone marrow and travel to the thymus. Following a series of maturation steps here, they circulate as small resting cells through the blood and lymph until they too encounter pathogenic organisms. Their pathway of activation is similar to that of B cells, and they proliferate in lymph nodes and recirculate to clear the infection. Both lymphomas and leukemias originate when a lymphocyte undergoes malignant transformation. This transformation can occur at any point along a B or T cell’s complex differentiation pathway; the point at which the transformation occurs determines the type and clinical course of the malignancy. Leukemias occur at an early stage of differentiation, when T and B cells are still in the bone marrow or have just left it. Lymphomas occur later, often while the lymphocytes are proliferating in lymph nodes. Classification. Over the past 40 years, numerous classiÞcation systems have been developed that divide NHLs into separate clinical subtypes (e.g., Rappaport, Gail and Mallory, Kiel). To standardize these systems, the Working Formulation (WF) was developed in 1982. It divides NHL subtypes according to clinical prognosis, grouping them as low-grade (indolent), intermediate-grade (aggressive), or high-grade (highly aggressive) (Table 1). These three categories subdivide depending on the morphological appearance of the lymphoma—e.g., whether the cells are large or small, cleaved or uncleaved. This system was widely used in the United States; the Kiel classiÞcation, which is based on both morphological and immunologic criteria to deÞne disease entities, remained popular in Europe. Gradually, clinicians found that some types of NHLs did not Þt into any of the WF categories. Moreover, improvements in immunologic and cytogenetic techniques led to the discovery of new NHL subtypes (e.g., mantle-cell

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TABLE 1. Clinical Groups of Non-Hodgkin’s Lymphomas As Recognized by the Working Formulation Recognized disease subtypes within the WF Low grade Small lymphocytic, consistent with chronic lymphocytic leukemia (SC) Follicular, predominantly small cleaved cell (FSC) Follicular, mixed small cleaved and large cell (FM) Intermediate grade Follicular, predominantly large cell (FL) Diffuse, small cleaved cell (DSC) Diffuse mixed, small and large cell (DM) Diffuse, large cleaved or noncleaved cell (DL) High grade Immunoblastic, large cell (IBL) Lymphoblastic, convoluted or nonconvoluted cell (LL) Small noncleaved cell, Burkitt’s or non-Burkitt’s (SNC) Recognized disease subtypes not included within the WF Other low-grade subtypes Mantle-cell lymphoma (MCL) Mucosa-associated lymphoid tissue (MALT), monocytoid B-cell lymphoma Other intermediate-grade subtypes Anaplastic large-cell lymphomas Other subtypes Mycosis fungoides Adult T-cell lymphomas/leukemias WF = Working Formulation, developed in 1982.

lymphoma [MCL]). The result was the Revised European-American Lymphoma (REAL) classiÞcation system. Devised by an international group of pathologists, this system divides lymphoid malignancies into three main groups depending on their cellular lineage: B-cell lymphomas, T-cell lymphomas, or Hodgkin’s disease. The REAL system subdivides these three groups into more than 20 subtypes based on cellular morphology, immunophenotype, and the genetic and clinical features of the lymphoma. Because the REAL classiÞcation system bases its main subdivisions on cellular lineage, it can also be used to classify leukemia. Under the auspices of the World Health Organization (WHO), and in conjunction with representatives of the international lymphoma community, the REAL classiÞcation system has undergone an update. The Revised European-American ClassiÞcation of Lymphoid Neoplasms/World Health Organization ClassiÞcation of Lymphoid Malignancies (Table 2) is expected to replace existing lymphoma classiÞcation systems throughout the world (Armitage JO, 1998). This was noted some time ago and is still applicable today. As previously mentioned, the REAL classiÞcation system divides NHLs into subtypes that have a B-cell origin (85% of cases) and subtypes that have a T-cell origin (15%). When considering treatment options, most clinicians divide the subtypes into lymphomas that progress quickly (aggressive lymphomas) and lymphomas that initially progress slowly (indolent lymphomas).

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TABLE 2. Updated REAL/WHO Classification of Non-Hodgkin’s Lymphomas (excluding Hodgkin’s lymphomas) Clinical Groups B-cell neoplasm Ia

II

First Subdivision Precursor B-cell neoplasm

B C D E F G

H I J K L T-cell and putative NK-cell neoplasms Precursor T-cell Ia neoplasm

A

B C D E F G

Clinical Nature of Subdivision

Precursor B-acute lymphoblastic leukemia/lymphoblastic lymphoma (B-ALL, LBL)

A

B-cell chronic lymphocytic leukemia/small lymphocytic lymphoma B-cell prolymphocytic leukemia Lymphoplasmacytic lymphoma/immunocytoma Mantle-cell lymphoma Follicular lymphoma Extranodal marginal zone B-cell lymphoma of MALT type Nodal marginal zone B-cell lymphoma (+/− monocytoid B cells) Splenic marginal zone lymphoma (+/− villous lymphocytes) Hairy-cell leukemia Plasmacytoma/plasma cell myeloma Diffuse large B-cell lymphoma Burkitt’s lymphoma

I

Peripheral B-cell neoplasms

A

II

Second Subdivision

A I A I I I

I I NSb A A

Precursor T-acute lymphoblastic leukemia/lymphoblastic lymphoma (T-ALL, LBL)

A

T-cell chronic lymphocytic leukemia/prolymphocytic leukemia T-cell granular lymphocytic leukemia Mycosis fungoides/Sezary’s syndrome Peripheral T-cell lymphoma, not otherwise characterized Hepatosplenic gamma/delta T-cell lymphoma Subcutaneous panniculitis-like T-cell lymphoma Angioimmunoblastic T-cell lymphoma

A

Peripheral T-cell and NK-cell neoplasms

I I A A A A

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TABLE 2. (continued) Clinical Groups H I J K L M

First Subdivision

Second Subdivision

Clinical Nature of Subdivision

Extranodal T-/NK-cell lymphoma, nasal type Enteropathy-type intestinal T-cell lymphoma Adult T-cell lymphoma/leukemia (HTLV 1+) Anaplastic large-cell lymphoma, primary systemic type Anaplastic large-cell lymphoma, primary cutaneous type Aggressive NK-cell leukemia

A A A A A A

a This subgroup is not additionally subdivided. b Myeloma and plasmacytoma are not clinically categorized as being indolent or aggressive in nature.

A = Aggressive/highly aggressive. HTLV = Human T-Cell Leukemia Virus I = Indolent. MALT = Mucosa-Associated Lymphoid Tissue. NK = Natural killer. NS = Not specified. REAL = Revised European-American Lymphoma. WHO = World Health Organization.

Aggressive NHLs account for approximately 60% of all diagnosed NHLs in the United States and Europe and 90% of NHLs diagnosed in Japan. Although the higher proliferation rate of the lymphoma cells means that aggressive NHLs are lethal within months if untreated, it also means that aggressive NHLs are more responsive to treatment than indolent NHLs. Indeed, about 60% of treated patients are cured. Diffuse large B-cell lymphoma (DLBCL) is the most prevalent subtype, constituting approximately 60% of aggressive NHLs. Indolent NHLs, also known as low-grade lymphomas under the WF, comprise approximately 40% of diagnosed NHLs in the United States and Europe and approximately 10% of NHLs diagnosed in Japan. Advanced-stage indolent NHLs are considered incurable, but because of the low physiological impact of these lymphomas, 70–85% of diagnosed patients survive Þve years after diagnosis and 50–70% are alive after ten years. Most indolent NHLs are initially chemosensitive, and patients often experience long-term remissions after initial chemotherapy. However, almost all patients ultimately relapse. On relapse, fewer patients’ NHL is chemosensitive, and subsequent remissions are shorter. In 20% of patients with indolent NHLs, death is the result of infectious complications that arise after the disease has become chemorefractory; in approximately 80% of patients, death results from causes unrelated to NHLs. On relapse, indolent NHLs may undergo transformation into an aggressive subtype. The proportion of patients whose disease undergoes transformation is unknown. Clinicians estimate that 20–40% of patients with indolent NHLs have

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transformed disease at relapse, but autopsies suggest that up to 90% of indolent patients have pathological characteristics of more aggressive disease. The clinical outlook for transformed NHL patients is poor; although patients may initially respond to treatment, remissions are short-lived. This report focuses on the aggressive lymphomas only, including lymphomas that have transformed from indolent NHLs. The clinical characteristics of aggressive NHL subtypes are described in Table 3. Molecular Stratification. While it is clear that lymphoma classiÞcation has progressed enormously in the last decade, signiÞcant clinical and biological heterogeneity exists within single disease subtypes. DLBCL is a particularly good example: some cases respond well to chemotherapy, but up to 60% are incurable, raising the question of whether this single diagnostic category hides more than one molecular disease (Staudt L, 2003). Scientists have used gene-expression, microarray-proÞling technology to further classify DLBCL into three subtypes based on the expression patterns, or “signatures,” of groups of genes (Rosenwald A, 2002; Shipp MA, 2002; Staudt L, 2003). Two of these subtypes are named according to the kinds of genes that distinguish them: germinal center B-celllike (GCB) and activated B-cell-like (ABC). The third group is known as “type 3” and was added to reßect the heterogeneity of gene expression in DLBCL. Clinical analysis showed that the GCB group is associated with signiÞcantly better patient survival than the ABC group. Type 3 is heterogeneous and poorly deÞned, but patient outcome is similar to outcome for patients in the ABC group. Investigators continue to analyze the gene signatures to elucidate the biological mechanisms behind these malignancies. The use of gene expression microarray to further deÞne DLBCL is a major breakthrough in our molecular and clinical understanding of this disease. However, it is an expensive and technically difÞcult task and is impractical for routine clinical use. A more practical aim is to characterize a group of prognostic genes that will predict therapy outcome and can be tested in a cost-effective assay. Some studies have used immunohistochemistry to analyze the expression of a small subset of genes in order to classify samples as GCB or non-GCB (Barrans SL, 2002; Colomo L, 2003; Linderoth J, 2003; McClintock S, 2003). The survival data resulting from these studies conßicted, and no microarray data were available to correlate with immunohistochemistry. A study published in 2004 was the Þrst to directly evaluate the expression of six prognostically important genes both by immunohistochemistry and by microarray (Hans CP, 2004). Investigators showed that the expression patterns of CD10 , bcl-6 , and MUM1 can be determined by using immunohistochemistry, which would enable classiÞcation of DLBCL samples into GCB and non-GCB subtypes. When compared with microarray data, immunohistochemistry results reproduced gene expression results in 71% of GCB and 88% of non-GCB cases and predicted similar patient survival outcomes. Discordant results were obtained in 22 cases that were classiÞed as GCB by microarray but as non-GCB by immunohistochemistry. These 22 patients had a median survival rate comparable

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TABLE 3. Clinical Characteristics of Aggressive Non-Hodgkin’s Lymphoma Subtypes

Subtypea

Clinical Characteristics

Subgroups from Different Staging Systems in Which Lymphomas May Have Been Classifiedb

Diffuse large B-cell lymphoma (DLBCL)

Most patients present with one or more Diffuse large cell, diffuse mixed rapidly enlarging symptomatic masses cell, immunoblastic, diffuse in nodal or extranodal sites. Considered centroblastic/centrocytic, curable; long-term survival for patients immunoblastic. treated with the current standard of care, CHOP-RC, is not yet known. Mantle-cell Male dominance; poor overall survival Newly recognized subtype. lymphoma with only 20–30% of patients surviving Diffuse small cleaved cell (most (MCL) at five years; considered in previous frequently in the Working staging systems to be indolent. Formulation), follicular small cleaved cell lymphoma, small lymphocytic lymphoma, diffuse large-cell lymphoma, lymphoblastic lymphoma, centrocytic lymphoma (most frequently in Kiel classification), or centrocytoid centroblastic lymphoma. Largest group of T-cell lymphomas in the Diffuse small cleaved cell, diffuse Peripheral REAL classification; low five-year mixed cell, diffuse large cell, T-cell survival of 20–30%; includes a wide immunoblastic. lymphomad variety of subtypes that cannot be otherwise specified. Immunophenotyping improves the accuracy of diagnosis by 45%. Anaplastic Second most common type of T-cell large T/nulllymphoma; often present with cell extranodal disease, especially in the lymphoma skin; median age is low (34), with a male predominance. Good prognosis; long-term overall survival is approximately 80%. Staining for CD30 antigen helps make the final diagnosis. Lymphoblastic Very aggressive subtype; five-year lymphoma survival is 20%; low median age at diagnosis; male predominance. Burkitt-like Difficult to distinguish from Burkitt’s lymphoma lymphoma and diffuse large B-cell lymphoma; therefore, difficult to diagnose accurately. Burkitt’s Most common in children in Africa and lymphoma pockets of the Caribbean; uncommon in adults, particularly in the United States and Europe; highly aggressive but sensitive to treatment; five-year survival approximately 50%. a Revised European-American Lymphoma (REAL) classification subtypes. b Alternative names used in the Working Formulation and the Kiel classification. c CHOP-R = Cyclophosphamide, doxorubicin, vincristine, and prednisone (chemotherapy) with rituximab (immunotherapy). d Excludes angiocentric nasal lymphoma and human T-lymphotropic virus type 1—associated lymphomas, which are more common in non-Western countries.

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to that of other non-GCB patients; investigators, therefore, suggested that they were correctly classiÞed by the immunohistochemistry method. One of the major challenges in microarray studies is to obtain a pure population of tumor cells, uncontaminated by healthy tissue that will obscure the gene expression proÞle of the tumor. PuriÞcation of malignant cells is easier in blood-borne tumors, such as chronic lymphocytic leukemia, than in solid tumors, where consistent results depend on dissecting malignant from healthy tissue. One reason for the discordant results obtained in the study described in the preceding paragraph could be the contamination of DLBCL tissue with healthy lymph node or stromal cells (Hans CP, 2004). In addition, microarray analysis measures the level of mRNA expression, which may not always correspond to protein level, thereby highlighting the importance of downstream protein analysis following microarray data analysis (Hans CP, 2004). Gene expression microarray proÞling has ushered in a new era, advancing our understanding of the mechanisms of cancer processes. It provides a wealth of complex information that requires in-depth statistical analyses. The associated costs and technical difÞculties have restricted this vital technology to the research setting, where it will continue to preempt the search for cancer markers that can be analyzed by assays such as immunohistochemistry. The aforementioned study is an important step in bridging the gap between microarray and routine analysis, but the authors state that this study requires conÞrmation and that further comparative analysis of gene and protein expression is needed (Hans CP, 2004) In addition to uncovering differentially expressed genes with potential diagnostic and prognostic signiÞcance, genes identiÞed by microarray technology may become future targets for biologic therapies. Candidate genes need to be further characterized at the protein level, both in tumor and healthy tissues, to determine their suitability as therapeutic targets. This technology provides a powerful initial step in the hunt for much-needed new targets.

Staging. The Ann Arbor staging classiÞcation system (Table 4) is commonly used to stage patients with NHLs. Originally developed for use in Hodgkin’s disease, this system is based on the anatomic distribution of the lymphoma. It subcategorizes patients according to the presence or absence of B symptoms (unexplained fever, night sweats, or weight loss exceeding 10% of body weight in the previous six months). The International Prognostic Index (IPI) was developed to assess risk in patients with an aggressive form of NHL who are treated with anthracyclinecontaining regimens (International Non-Hodgkin’s Lymphoma Prognostic Factors Project, 1993). Since its publication, some clinicians have applied the IPI to most types of NHL, both indolent and aggressive. The index assigns patients a score of 0 to 5 depending on the presence or absence of the following adverse risk factors: age older than 60 years, Eastern Cooperative Oncology Group (ECOG) performance status of 2 or more, elevated lactate dehydrogenase (LDH), Ann Arbor stage III or IV, and two or more extranodal sites. Depending on their

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TABLE 4. Ann Arbor Staging Classification Stage I II

III IV

Criteria Involvement of a single node region or a single extranodal site or organ. Involvement of two or more lymph node regions on the same side of the diaphragm, or localized involvement of an entranodal site or organ AND one or more lymph node regions on the same side of the diaphragm. Involvement of lymph node regions on both sides of the diaphragm, possibly accompanied by localized involvement of an extranodal organ or site. Diffuse or disseminated involvement of one or more distant extranodal organs with or without associated lymph node involvement.

Note: The following symptoms, known as B symptoms are commonly associated with NHLs: unexplained fever, night sweats, and/or weight loss of >10% body weight. The absence or presence of these symptoms is indicated by the suffix A or B, respectively, to the patient’s stage classification.

TABLE 5. International Prognostic Index Scoring for Patients of All Ages IPI Risk Group

factorsa

Number of risk Frequency (%) Relapse-free 5-year survival (%) Overall 5-year survival (%)

Low

Low Intermediate

High Intermediate

0–1 35 70 73

2 27 50 51

3 22 49 43

High 4.5 16 40 26

a Risk factors are age >60, elevated lactate dehydrogenase (LDH) levels, Eastern Cooperative Oncology Group (ECOG) performance status of 2–4, Ann Arbor stage III or IV, and more than one extranodal site. IPI = International Prognostic Index.

scores, patients are assigned to one of four risk groups, each with a different expected long-term survival (Table 5). For patients younger than age 60, a simpliÞed system based on the Ann Arbor staging system has been developed: LDH level and ECOG performance status. Both the Ann Arbor staging system and the IPI can help determine the appropriate therapeutic approach for individual patients. However, one of the criticisms of the IPI is that many patients fall into the intermediate category, thereby reducing its clinical usefulness for predicting survival. Etiology Risk Factors. In the majority of NHLs cases, the cause is unknown. Although several factors have been implicated in this disease’s development, only a minority of cases can be traced to environmental, bacterial, or viral risk factors. Chemical/Physical Agent Exposures. The strongest known risk factor for NHL is exposure to environmental toxins; clinicians are able to attribute 2–10% of NHL cases worldwide to such exposures. IdentiÞed toxins include creosote,

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lead, paint thinner, oils and greases, dark-toned hair dyes, and most notably, agricultural pesticides, whose contaminants dioxin and phenoxyacetic acid are the most well-documented risk factors. Pockets of increased incidence of NHL have been reported in agricultural communities in the United States and Europe (Scherr PA, 1992; Persson B, 1999). Although it might be assumed that NHLs associated with agricultural chemicals would decline along with the downward trend of farming in industrialized countries, the increased use of pesticides and herbicides in home gardens and on golf courses, playing Þelds, and beaches has increased the risk of exposure in residential and recreational environments. Patients with a history of exposure to such agents exhibit chromosomal abnormalities, including frequent translocations within genes that mediate immune functions. In 1997, researchers identiÞed a strong dose-response relation between lipid-corrected serum polychlorinated biphenyl (PCB) concentrations and risk for NHLs: people who had the highest PCB concentrations had as much as a 12fold higher risk of developing NHLs (Rothman N, 1997). Still, the nature and biological plausibility of the association are not clearly understood. In addition, increasing evidence indicates that exposure to nuclear radiation heightens the risk for developing NHLs, but data on this risk factor are sparse. The development of NHLs has also been reported among patients who are receiving therapy for other neoplasms, such as Hodgkin’s disease. The incidence of NHLs in patients treated for Hodgkin’s disease was found to be 32 times that of the general population (Dorreen MS, 1986). Additionally, data from the Surveillance, Epidemiology, and End Results (SEER) database have shown that in the United States, patients who have Hodgkin’s disease die at an increased rate from NHLs compared with the general population (Diehl L, 1999) Infectious Agents. Both bacterial and viral pathogens have been associated with the development of NHLs: •

•

The presence of the Helicobacter pylori bacterium is strongly associated with gastric lymphomas. Infection with H. pylori results in chronic gastritis, and conversely, primary gastric mucosal-associated lymphoid tissue (MALT) NHL cases frequently show evidence of H. pylori infection. Studies have indicated that patients with primary gastric lymphoma were approximately six times more likely to be positive for H. pylori than case-matched controls. Studies of treatments directed against H. pylori have suggested that in many cases, eradicating the bacteria induces a positive clinical response in the lymphoma, further implying a causal role for the bacteria (Wotherspoon AC, 2002). Human T-cell leukemia virus type 1 (HTLV-1) is a human retrovirus originally isolated from aggressive adult T-cell lymphomas found predominantly in Japan. The virus is transmitted through the exchange of body ßuids. Although HTLV-1 is endemic to several regions of Japan, available data indicate that infection with the virus correlates only weakly with the development of NHLs: only about 50% of all NHL cases in these areas are

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•

•

79

associated with the virus. This Þnding suggests that HTLV-1 may predispose a person to develop a T-cell lymphoma but is not directly able to predict lymphoma development. Epstein-Barr virus (EBV) is a herpes virus that readily transforms B lymphocytes into immortal cell lines in vitro. EBV also causes mononucleosis and is strongly associated with neoplasms, including aggressive NHLs such as Burkitt’s lymphoma. The data suggest that EBV-associated NHLs result from virus-mediated changes to the cell surface, enabling infected cells to escape normal cell regulation. However, exposure to EBV without subsequent development of lymphoma is common, strengthening the argument that EBV predisposes patients to, or enables, development of NHLs rather than directly causing it. In addition, the data suggest that infection with EBV is a potent catalyst for the development of NHLs among people with other risk factors—particularly HIV patients and patients who are receiving immunosuppressive therapy after solid organ transplantation. Before the advent of new antiretroviral therapies, the NHLs (particularly those of the central nervous system [CNS], i.e., primary parenchymal lymphoma and leptomeningeal lymphoma) had been occurring with rising frequency among HIV patients. Indeed, CNS NHL is now considered an AIDS-deÞning illness. HIV patients are believed to have as much as a 60to 100-fold higher risk of developing NHLs than people who do not have HIV. The etiology of HIV-associated NHLs is complicated by the underlying immune system dysfunction and the various infections that accompany AIDS: infected patients have uncontrolled B-cell stimulation and proliferation, increasing the likelihood of genetic errors during lymphocyte proliferation. HIV patients are also at higher risk (as are all immunodeÞcient persons) for lymphomas associated with EBV. Thanks to antiretroviral therapy, however, the incidence of NHLs in AIDS patients has plateaued or is declining in most Western countries that have strong AIDS programs.

Medical Immune Suppression. The high level of immunosuppression induced in solid-organ and bone marrow transplantation can result in a post-transplant lymphoproliferative disorder (PTLD), in which lymphocytes proliferate abnormally. PTLDs can range in severity from a benign mononucleosis-like illness to NHLs; they originate most often in B cells and are associated with EBV (Loren AW, 2003). Because a suppressed immune system is less able to control viral infection, the result is unchecked proliferation of virally infected cells that may progress to NHL. Patients who receive heavy immunosuppressive therapy after organ transplantation have a 25- to 50-fold higher risk of developing a lymphoid malignancy, usually of a highly aggressive type. Prior history of blood transfusion has been associated with increased risk of NHL development (Cerhan JR, 1997). One study suggests that at worst, blood transfusion doubles the risk of NHLs, and at best, the risk does not increase after a transfusion (Vamvakas EC, 2000).

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Non-HIV Autoimmune Disorders. Autoimmune disorders such as Sj¨ogren’s syndrome (a chronic autoimmune disorder affecting the glands that produce tears and saliva), Hashimoto’s thyroiditis, and rheumatoid arthritis are all risk factors for NHLs. In many cases, chronic inßammation promotes the development of MALT and other lymphomas; patients with Sj¨ogren’s syndrome, for example, have a 6% increased risk for NHLs. The relationship between NHLs and autoimmune diseases remains unclear. The inherited immune deÞciency syndrome ataxia-telangiectasia, an autosomal recessive multisystem disorder, is associated with a heightened risk of developing lymphomas. The gene that is mutated in this disease (the ataxia-telangiectasia mutated gene, known as ATM ) is located on chromosome 11, and multiple mutations have been described (Stankovi T, 2002). The ATM protein is a protein kinase implicated in the integration of different cellular responses to DNA damage. Diet and Family History. Studies suggest that high dietary intake of meats and fat from animal sources is associated with an increased risk of NHLs (Chiu BC, 1996; Zhang S, 1999). A greater intake of vegetables, particularly cruciferous vegetables (e.g., broccoli, caulißower, Brussel sprouts), may reduce the risk of NHLs in women (Zhang SM, 2000). Some studies have shown that tobacco use is also associated with NHLs. A history of adult-onset diabetes mellitus (type 2 diabetes) and a history of cancer appear to be risk factors for NHLs in older women (Cerhan JR, 1997). Patients with Wiskott-Aldrich syndrome, an X-linked recessive disorder, have a 100-fold greater risk than the general population of developing NHLs (RemoldO’Donnell E, 1996). Cytogenetic Abnormalities. NHLs are associated with an array of cytogenetic abnormalities, many of them chromosomal translocations (Table 6). Such abnormalities occur when part of one chromosome is translocated onto another chromosome, possibly resulting in activation of an oncogene or disruption of a tumor suppressor gene. MCL, which accounts for approximately 6% of all NHLs in the United States, is characterized by a translocation t(11;14)(q13;q32) that involves chromosomes 11 and 14 (Swerdlow SH, 2002). In this translocation, the cyclin-D1 gene (also known as bcl-1, PRAD1, and ccnd-1 ) is juxtaposed with the enhancer of the immunoglobulin heavy (IgH ) chain gene (Vega F, 2003). The enhancer drives expression of cyclin-D1, and the resulting high levels of cyclin-D1 protein push cells through the cell cycle, accounting in part for the aggressive clinical course of MCL. Diffuse large B-cell lymphoma (DLBCL) is heterogeneous, both in biology and clinical course, and has been associated with a wide variety of molecular abnormalities. Two of the more common abnormalities involve the bcl-2 and bcl6 genes. The t(14;18)(q32;q21) puts bcl-2 under the control of the IgH enhancer (as described for MCL), leading to overexpression of the antiapoptotic bcl-2 protein. High levels of bcl-2 protein promote abnormal cell survival and override the normal propensity for cell death. bcl-6 encodes a transcription factor involved

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TABLE 6. Oncogenes, Immunophenotypes, and Cytogenetic Characteristics of Common Aggressive Non-Hodgkin’s Lymphoma Subtypes

Subtypea

Oncogenes Frequently Involved

Diffuse large B-cell lymphoma

bcl-2, c-myc, bcl-6

Mantle-cell lymphoma

bcl-1, PRAD1

Peripheral T-cell lymphomasb Anaplastic large T/null-cell lymphoma

Not known

Precursor T-cell or B-cell lymphoblastic lymphoma

Variable

Burkitt-like lymphoma Burkitt’s lymphoma

bcl-2, c-myc

ALK

c-myc

Typical Immunophenotype CD5+/−, CD10+/−, CD19+, CD20+, CD22+, CD79a+, CD45+/−, surface immunoglobulin (sIg) +/− CD5+, CD10+/−, CD11c-, CD19+, CD20+, CD22+, CD23-, CD43+, CD79a+, sIg+ CD2+, CD3+, CD4+, CD5+, CD7+, CD8+/− CD3+/−, CD15+/−, CD20-, CD25+/−, CD3-, CD30+, CD43+/−, CD45RO+/−, CD68-, EMA+/− T cell; CD1a+/−, CD2+, CD3+, CD4+/− CD5+, CD7+, CD8+/−, TdT+. B cell; CD10+/−, CD19+, CD20+/−, CD22+, CD79a+, CD34+/−, sIgCD19+, CD20+, CD22+, CD79a+, CD5-, CD10-, sIg+/− CD19+, CD20+, CD22+, CD79a+, CD10+, CD5-, CD23-, sIg+

Characteristic Cytogenetics t(14;18)(q32;q21), t(8;14)(q24;q32), t(3;14)(q27;q32) t(11;14)(q13;q32)

Trisomy 8q t(2;5)(p23;q35)

Variable

t(14;18)(q32;q21) t(8;14)(q24;q32) t(8;14)(q24;q32) t(2;8)(q11;q24) t(8;22)(q24;q11)

a Revised European-American Lymphoma (REAL) classification subtypes. b Excludes angiocentric nasal lymphoma and human T-lymphotropic virus type 1-associated lymphomas,

which are more common in non-Western countries.

with cellular differentiation and has been found translocated to many different gene partners. Further analysis is required to determine the prognostic signiÞcance and impact of the various cytogenetic abnormalities associated with NHLs.

CURRENT THERAPIES Combination chemotherapy is the mainstay of treatment for aggressive nonHodgkin’s lymphomas (NHLs); it can cure 30–40% of patients. A series of regimens developed during the 15 years between 1978 and 1993 aimed to improve on CHOP, the most widely used regimen, which consists of the following agents: •

Cyclophosphamide (Bristol-Myers Squibb’s Cytoxan, Baxter’s Endoxan/ Endoxana, generics).

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AGGRESSIVE NON-HODGKIN’S LYMPHOMAS

• • •

Doxorubicin{hydroxydoxorubicin} (PÞzer’s Adriamycin/Adriblastine, Kyowa’s Adriacin, generics). Vincristine (Eli Lilly/EG Labo’s Oncovin, generics). Prednisone (Merck’s Decortin, generics). In 1993, a large, randomized trial found that three of the regimens developed to compete against CHOP conferred no additional beneÞt, so CHOP remains the standard Þrst-line treatment for aggressive NHLs (Fisher RI, 1993). As a result of this trial and others described in this chapter, the alternative regimens discussed in the following sections are reserved largely for second-line use.

Researchers continue to modify CHOP, either increasing doses, reducing intervals between cycles, or adding or substituting drugs to try to improve response and survival rates and/or reduce toxicities. To date, only the addition of the immunotherapy rituximab (Genentech/Biogen Idec/Chugai’s Rituxan, Roche’s MabThera) to CHOP (forming CHOP-R) has conferred a signiÞcant beneÞt in a large trial. Table 7 lists the leading regimens available to treat aggressive NHLs. Other regimens used by a minority of clinicians include the following: •

•

•

ACVBP, which comprises doxorubicin (PÞzer’s Adriamycin/Adriblastine, Kyowa’s Adriacin, generics); cyclophosphamide (Bristol-Myers Squibb’s Cytoxan, Baxter’s Endoxan/Endoxana, generics); vindesine (EG Labo/Cell Pharm/Crinos/Clonmel’s Eldisine, Ciclum Farma’s Enison, Shionogi’s Fildesin); bleomycin (Bristol-Myers Squibb’s Blenoxane/Nippon Kayaku’s Bleo, generics); and prednisone (Merck’s Decortin, generics). EPOCH, which incorporates etoposide (Bristol-Myers Squibb’s VePesid/ Etopophos, PÞzer’s Lastet, generics); vincristine (Eli Lilly/EG Labo’s Oncovin, generics); doxorubicin (PÞzer’s Adriamycin/Adriblastine, Kyowa’s Adriacin, generics); cyclophosphamide (Bristol-Myers Squibb’s Cytoxan, Baxter’s Endoxan/Endoxana, generics); and prednisone (Merck’s Decortin, generics). ICE, which combines ifosfamide (Bristol-Myers Squibb’s Ifex, Baxter’s Holoxan/Mitoxana, Shionogi’s Ifomide, generics); carboplatin (BristolMyers Squibb’s Paraplatin/Carboplat, generics); and etoposide (BristolMyers Squibb’s VePesid/Etopophos, PÞzer’s Lastet, generics).

Table 8 presents the results of trials that have examined the addition of rituximab to several existing regimens. In general, these trials have been small, and although response rates have been high, larger trials are needed to establish the impact of these combination regimens on survival and their role in the treatment of aggressive NHLs. Cyclophosphamide, Doxorubicin, Vincristine, and Prednisone (CHOP) Overview. CHOP, the mainstay of treatment for aggressive NHLs, incorporates cyclophosphamide (Bristol-Myers Squibb’s Cytoxan, Baxter’s

CURRENT THERAPIES

83

TABLE 7. Current Regimens/Classes Used for Aggressive Non-Hodgkin’s Lymphomas Regimen Components Regimen CHOP

Agent

Availability

Cyclophosphamide US, F, G, I, (Bristol-Myers S, UK, J Squibb’s Cytoxan, Baxter’s Endoxan/Endoxana, generics)

Dose Cyclophosphamide: 750 mg/m2 , day 1, IV

Doxorubicin (Pfizer’s US, F, G, I, Doxorubicin: 50 Adriamycin/ S, UK, J mg/m2 , day 1, IV Adriblastine, Kyowa’s Adriacin, generics) Vincristine (Eli Lilly/EG US, F, G, I, Vincristine: 1.4 mg/m2 , day 1 (maximum 2 Labo’s Oncovin, S, UK, J mg), IV generics) Prednisonea (Merck’s US, F, G, I, S Prednisone: 100 Decortin, generics) mg/m2 /d, days 1–5, PO Repeat every 21 days. CHOP-R Cyclophosphamide US, F, G, I, Cyclophosphamide: (Bristol-Myers S, UK, J 750 mg/m2 ,day 1, IV Squibb’s Cytoxan, Baxter’s Endoxan/Endoxana, generics)

DHAP

Common Toxicities Fever Infection Myelosuppression Lung toxicity Alopecia Cardiac toxicity Neurological toxicity

Fever Infection Myelosuppression Lung toxicity Alopecia Cardiac toxicity Neurological toxicity Respiratory symptoms Chills Hypotension

Doxorubicin (Pfizer’s US, F, G, I, Doxorubicin: 50 Adriamycin/ S, UK, J mg/m2 , day 1, IV Adriblastine, Kyowa’s Adriacin, generics) Vincristine (Eli Lilly/ US, F, G, I, Vincristine: 1.4 mg/m2 , day 1 (maximum 2 EG Labo’s Oncovin, S, UK, J mg), IV generics) Prednisonea (Merck’s US, F, G, I, S Prednisone: 100 mg/d, Decortin, generics) days 1–4, PO Repeat every 21 days for 8 cycles of CHOP. Rituximab US, F, G, I, Rituximab: 375 mg/ (Genentech/Biogen S, UK, J m2 /d, day 1 of each of the 8 cycles of Idec/Chugai’s CHOP, IV Rituxan, Roche’s MabThera) Cisplatin (Bristol-Myers US, F, G, I, Cisplatin: 100 mg/m2 , Myelosuppressionday 1 related infection Squibb’s Platinol AQ, S, UK, J Acute lysis syndrome generics)

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AGGRESSIVE NON-HODGKIN’S LYMPHOMAS

TABLE 7. (continued) Regimen Components Regimen

Agent

Availability

Cytarabine (Pfizer’s US, F, G, I, Cytosar-U/Aracytine, S, UK, J Nippon Shinyaku’s Cylocide, generics) Dexamethasone US, F, G, I, (Merck/Banyu’s S, UK, J Decadron, generics) ESHAP

Methylprednisolone US, F, G, I, (Pfizer’s Medrol/ UK, J Medrone/Medrate, generics) Etoposide US, F, G, I, (Bristol-Myers S, UK, J Squibb’s VePesid/Etopophos, Pfizer’s Lastet, generics) Cytarabine (Pfizer’s US, F, G, I, Cytosar-U/Aracytine, S, UK, J Nippon Shinyaku’s Cylocide, generics) Cisplatin (Bristol-Myers US, F, G, I, Squibb’s Platinol AQ, S, UK, J generics)

MACOP- Methotrexate B (generics)

Leucovorin (generics)

US, F, G, I, S, UK, J

US, F, G, I, S, UK, J

Doxorubicin (Pfizer’s US, F, G, I, Adriamycin/ S, UK, J Adriblastine, Kyowa’s Adriacin, generics) Cyclophosphamide US, F, G, I, (Bristol-Myers S, UK, J Squibb’s Cytoxan, Baxter’s Endoxan/Endoxana, generics) Vincristine (Eli Lilly/EG US, F, G, I, Labo’s Oncovin, S, UK, J generics)

Dose

Common Toxicities 2

Cytarabine: 2 mg/m , day 2

Dexamethasone: 40 mg/d, days 1–4 Repeat every 21–28 days. Methylprednisolone: Myelosuppression 500 mg/d, days 1–4 Nausea and vomiting Non-neutropenic sepsis Etoposide: 40–60

mg/m2 /d, days 1–4 Cytarabine: 2 g/m2 , day 5 Cisplatin : 25 mg/m2 /d, days 1–4 Repeat every 21–28 days. Myelosuppression Methotrexate: 400 mg/m2 once weekly, Infection weeks 2, 6, and 10, Cardiotoxicity IV Neurotoxicity Leucovorin: 15 mg/m2 every 6 hours × 6 (24 hours after methotrexate), PO Doxorubicin: 50 mg/m2 per week, weeks 1, 3, 5, 7, 9, and 11, IV

Cyclophosphamide: 350 mg/m2 per week, weeks 1, 3, 5, 7, 9, and 11, IV

Vincristine: 1.4 mg/m2 per week, weeks 2, 4, 6, 8, 10, and 12, IV

CURRENT THERAPIES

85

TABLE 7. (continued) Regimen Components Regimen

Agent Bleomycin (Bristol-Myers Squibb’s Blenoxane, Nippon Kayaku’s Bleo, generics Prednisonea (Merck’s Decortin, generics)

MINE

BEAM

Availability

Dose

Common Toxicities

US, F, G, I, S, UK, J

Bleomycin: 10 units/m2 per week, weeks 4, 8, and 12, IV US, F, G, I, S Prednisonea : 75 mg/d; dose tapered over the last 15 days, PO Repeat every 12 weeks. Mesna (Bristol-Myers US, F, G, I, S Mesna: 1,330 mg/ Neutropenic fever Squibb’s Mesna/ UK, J m2 /d, days 1–3, Sepsis IV Bleeding associated Mesnex, Baxter/ with thrombocytoShionogi’s Uromitexan) penia Ifosfamide (Bristol-Myers US, F, G, I, Ifosfamide: 1,330 Squibb’s Ifex, Baxter’s S, UK, J mg/m2 /d, days 1–2, IV Holoxan/ Mitoxana, Shionogi’s Ifomide, generics) Mitoxantrone (Serono/ US, F, G, I, Mitoxantrone: 8 mg/ Wyeth/Wyeth-Takeda’s S, UK, J m2 , day 1, IV Novantrone, Baxter’s Onkotrone) Etoposide (Bristol-Myers US, F, G, I, Etoposide: 65 mg/ Squibb’s S, UK, J m2 /d, days 1–3, IV VePesid/Etopophos, Repeat every 21 Pfizer’s Lastet, days. generics) Neutropenic fever Carmustine (Bristol-Myers US, F, G, UK Carmustine: 300 Squibb’s mg/m2 , day 1, IV Sepsis Bleeding associated BiCNU/Carmubris) with thrombocytopenia Etoposide (Bristol-Myers US, F, G, I, Etoposide: 400 Squibb’s S, UK, J mg/m2 /d, days 2–5, IV VePesid/Etopophos, Pfizer’s Lastet, generics) Cytarabine (Pfizer’s US, F, G, I, Aracytine: 400 Cytosar-U/Aracytine, S, UK, J mg/m2 /d, days 2–5, IV Nippon Shinyaku’s Cylocide, generics) Melphalan US, F, G, I, Melphalan: 140 (GlaxoSmithKline’s S, UK, J mg/m2 , day 6, IV Alkeran)

a Prednisolone is substituted for prednisone in countries where prednisone is unavailable.

US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan. IV = Intravenous; PO = By mouth; SC = Subcutaneous.

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AGGRESSIVE NON-HODGKIN’S LYMPHOMAS

TABLE 8. Findings of Clinical Trials Investigating the Combination of Rituximab with Currently Used Regimens in Patients with Aggressive Non-Hodgkin’s Lymphomas

Clinical Setting

Regimen

Number of Patients

Relapsed/refractory DHAP-R for 4 45 aggressive B-cell cycles lymphomaa Relapsed/refractory R-ESHAP plus 11 (DLBCL 10 DLBCL or MCL GM-CSF for 6 patients, MCL (including prior RT or cycles 1 patient) ASCT)b R-ESHAP: 2-4 8 Aggressive B-cell cycles of ESHAP, lymphoma in first 8 infusions of relapse, or partially rituximab sensitive to followed by anthracycline-based HDT-ASCT for treatmentc responders 39 Relapsed, refractory or R-EPOCH for 4-6 cycles transformed aggressive CD20-positive B-cell lymphomad R-ICE 31 Previously treated patients with aggressive NHLse Previously untreated Hyper-CVAD-R for 56 stage IV mantle-cell at least 6 cycles lymphomaf

Previously untreated and previously treated mantle-cell lymphomag

Fludarabine, mitoxantrone, and rituximab

20

Overall Response/ Complete Response (%)

Survival Data

48/28

—

84/67

—

87.5/75

—

69

Median overall survival: 14.5 months Projected 2-year survival: 49% —

81/55

—/89

—/88

Two-year failure-free survival: 72% Two-year overall survival: 90% All patients remained alive after a median follow-up of 16.4 months

a Mey UJM, 2003. b Venugopal P, 2003. c Piliotis EG, 2003. d Jost LM, 2001. e Kewalramani T, 2001. f Romaguera J, 2003. g Mohrbacker A, 2004.

Note: Full source citations appear in ‘‘References’’ ASCT = Autologous stem-cell transplantation. DHAP-R = Cisplatin, cytarabine, dexamethasone, and rituximab. DLBCL = Diffuse large B-cell lymphoma. GM-CSF = Granulocyte-macrophage colony-stimulation factor. Hyper-CVAD-R = Cyclophosphamide, doxorubicin, vincristine, dexamethasone, and rituximab. MCL = Mantle-cell lymphoma. R-EPOCH = Etoposide, vincristine, doxorubicin, cyclophosphamide, and prednisone with rituximab. R-ESHAP = Methylprednisolone, etoposide, cytarabine, cisplatin, and rituximab. R-ICE = Ifosfamide, carboplatin, etoposide, and rituximab. RT = Radiotherapy.

CURRENT THERAPIES

87

Endoxan/Endoxana, generics); doxorubicin (PÞzer’s Adriamycin/Adriblastine, Kyowa’s Adriacin, generics); vincristine (Eli Lilly/EG Labo’s Oncovin, generics); and prednisone (Merck’s Decortin, generics). Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

•

Cyclophosphamide (Figure 1) is an alkylating agent and a cyclic phosphamide ester of mechlorethamine that is activated in the liver. Alkylation of DNA causes both strand breaks and the formation of cross-links between strands of DNA. Both of these events result in aberrant DNA replication and transcription of RNA. Cyclophosphamide, like other alkylating agents, is cell-cycle nonspeciÞc. However, rapidly proliferating cells are more susceptible to the action of alkylating agents because of the reduced time available for DNA enzymes to repair the cytotoxic damage. Doxorubicin (Figure 2) is an anthracycline. Anthracyclines interact with several different cellular targets, most importantly topoisomerase II. By inhibiting this DNA religation enzyme, anthracyclines exert their cytotoxic

FIGURE 1. Structure of cyclophosphamide.

FIGURE 2. Structure of doxorubicin (R = OCH3 , R1 = OH, R2 = H, R3 = H, R4 = OH).

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AGGRESSIVE NON-HODGKIN’S LYMPHOMAS

•

•

effect. Another doxorubicin mechanism that leads to cell death is known as DNA intercalation, in which the anthracycline molecule inserts itself between base pairs. This phenomenon causes single- and double-stranded breaks in DNA that inhibit cell proliferation. The free radicals generated by the reductive metabolism of anthracyclines may also damage cellular structures such as proteins and DNA. Vincristine (Figure 3) is a vinca alkaloid. Vinca alkaloids act by binding with microtubular proteins of the mitotic spindle, thereby leading to mitotic arrest or cell death. Vinca alkaloids exert selective toxicity to tumor cells by acting on proliferating cells. Prednisone (Figure 4) is a glucocorticosteroid. Glucocorticosteroids reduce the inßammatory response to tumor tissue by preventing white blood cells from functioning to produce swelling and pain around the tumor site. Corticosteroids induce lysis of peripheral lymphocytes and slow lymphocyte production. Adding corticosteroids to chemotherapy regimens may augment their efÞcacy.

N

OH CH3

NH H3CO H3CO

N

O

H N R

CH3 OAc O

H HO H3CO

FIGURE 3. Structure of vincristine (R = CHO).

CH2OH

O

H 3C

C

O OH

H3C

O FIGURE 4. Structure of prednisone.

CURRENT THERAPIES

89

Clinical Performance. In 1993, the Southwest Oncology Group (SWOG) and the Eastern Cooperative Oncology Group (ECOG) conducted a prospective, randomized Phase III trial that compared CHOP with three regimens that had seemed to offer improved survival over CHOP in Phase II trials (Fisher RI, 1993). The trial involved 899 treatment-naive patients in advanced stages of intermediate-grade or high-grade NHL. Patients in each arm received a maximum of eight cycles of therapy. The three investigational regimens were as follows: • •

•

m-BACOD: consisting of low-dose methotrexate with leucovorin rescue, bleomycin, doxorubicin, cyclophosphamide, vincristine, and dexamethasone; ProMACE-CytaBOM: consisting of prednisone, doxorubicin, cyclophosphamide, and etoposide, followed by cytarabine, bleomycin, vincristine, and methotrexate with leucovorin rescue; and MACOP-B: consisting of methotrexate with leucovorin rescue, doxorubicin, cyclophosphamide, vincristine, prednisone, and bleomycin.

The median ages of the groups ranged from 54 to 57 years. Approximately 25% of the patients studied were aged 65 or older (Fisher RI, 1993). At three years, 44% of all patients were alive and without disease; investigators observed no signiÞcant differences in disease-free survival among the groups: 41% in the CHOP and MACOP-B groups and 46% in the m-BACOD and ProMACE-CytaBOM groups (p = 0.35). Overall survival at three years was 52%: 50% in the ProMACE-CytaBOM and MACOP-B groups, 52% in the mBACOD group, and 54% in the CHOP group (p = 0.90). Researchers found no subgroup of patients in which survival was superior in a non-CHOP arm. No signiÞcant differences occurred in the response rates of the four treatment groups. The rates of objective antitumor responses were 80% in the CHOP group, 82% in the m-BACOD group, 83% in the MACOP-B group, and 87% in the ProMACE-CytaBOM group. The rates of complete response were 44% for CHOP, 48% for m-BACOD, 56% for ProMACE-CytaBOM, and 51% for MACOP-B. The rates of partial response were 36% for CHOP, 34% for m-BACOD, 31% for ProMACE-CytaBOM, and 32% for MACOP-B (Fisher RI, 1993). In patients with Ann Arbor stage I or localized stage II aggressive NHLs, the addition of radiotherapy to CHOP chemotherapy increased survival compared with CHOP alone. In a Phase III trial, conducted by SWOG, 401 patients with localized aggressive NHLs were randomized to receive eight cycles of CHOP alone or three cycles of CHOP and 4–5.5 Gy involved-Þeld radiotherapy (Miller TP, 1998). Five-year survival was 82% in the combined modality arm and 72% in the CHOP-only arm. Fewer life-threatening toxicities were observed in the radiotherapy/CHOP arm (31% of patients) than in the CHOP-only arm (40%). The CHOP-14 study from the German Non-Hodgkin’s Lymphoma Study Group used moderately increased doses (300 mg/m2 cyclophosphamide, 25 mg/m2 doxorubicin, 0.7 g/m2 vincristine) and reduced the interval between

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AGGRESSIVE NON-HODGKIN’S LYMPHOMAS

courses to 14 days (Pfreundschuh M, 2002). This schedule yielded a moderate increase in survival compared with standard CHOP and is favored by some clinicians. Cyclophosphamide, Doxorubicin, Vincristine, Prednisone, and Rituximab (CHOP-R) Overview. The CHOP-R regimen incorporates cyclophosphamide (BristolMyers Squibb’s Cytoxan, Baxter’s Endoxan/Endoxana, generics); doxorubicin (PÞzer’s Adriamycin/Adriblastine, Kyowa’s Adriacin, generics); vincristine (Eli Lilly/EG Labo’s Oncovin, generics); prednisone (Merck’s Decortin, generics); and rituximab (Genentech/Biogen-Idec/Chugai’s Rituxan, Roche’s MabThera). Rituximab has been approved in the United States for patients with low-grade or follicular, CD20+, B-cell NHLs who have relapsed or are refractory to treatment; and Þrst-line treatment of diffuse large B-cell, CD20-positive, nonHodgkin’s lymphoma in combination with CHOP or other anthracycline-based chemotherapy regimens. In Europe, rituximab is approved for CD20+ diffuse large B-cell lymphoma (DLBCL). Although the Þrst published Phase III trial examining CHOP-R in aggressive NHLs involved just elderly patients, clinicians have extrapolated these Þndings as a basis for treating patients of all ages. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the overall regimen activity: •

•

•

Cyclophosphamide (Figure 1) is an alkylating agent and a cyclic phosphamide ester of mechlorethamine that is activated in the liver. Alkylation of DNA causes both strand breaks and the formation of cross-links between strands of DNA. Both of these events result in aberrant DNA replication and transcription of RNA. Cyclophosphamide, like other alkylating agents, is cell-cycle nonspeciÞc. However, rapidly proliferating cells are more susceptible to the action of alkylating agents because of the reduced time available for DNA enzymes to repair the cytotoxic damage. Doxorubicin (Figure 2) is an anthracycline. Anthracyclines interact with several different cellular targets, most importantly topoisomerase II. By inhibiting this DNA religation enzyme, anthracyclines exert their cytotoxic effect. Another doxorubicin mechanism that leads to cell death is known as DNA intercalation, in which the anthracycline molecule inserts itself between base pairs. This phenomenon causes single- and double-stranded breaks in DNA that inhibit cell proliferation. The free radicals generated by the reductive metabolism of anthracyclines may also damage cellular structures such as proteins and DNA. Vincristine (Figure 3) is a vinca alkaloid. Vinca alkaloids act by binding with microtubular proteins of the mitotic spindle, thereby leading to mitotic arrest or cell death. Vinca alkaloids exert selective toxicity to tumor cells by acting on proliferating cells.

CURRENT THERAPIES

•

•

91

Prednisone (Figure 4) is a glucocorticosteroid. Glucocorticosteroids reduce the inßammatory response to tumor tissue by preventing white blood cells from functioning to produce swelling and pain around the tumor site. Corticosteroids induce lysis of peripheral lymphocytes and slow lymphocyte production. Adding corticosteroids to chemotherapy regimens may augment their efÞcacy. Rituximab is a mouse/human chimeric monoclonal antibody (MAb) directed against the cluster of differentiation (CD) 20 molecule. CD20 is a calcium channel that interacts with the B-cell immunoglobulin receptor complex (Bubien JK, 1993) and is expressed on both normal and malignant B cells, making it an ideal target for MAb therapy in B-cell disorders. Rituximab has been proposed to deplete B cells following binding to CD20 in several ways, including antibody-dependent cellular cytotoxicity (ADCC), complementdependent cytotoxicity (CDC), and alteration of calcium ßux and factors involved in apoptosis (Lin T, 2003). CD20 regulates the activation process for cell-cycle initiation and differentiation. CD20 antigen is present in normal and malignant pre-B and mature B lymphocytes, including B lymphocytes found in more than 90% of DLBCLs. Rituximab causes lysis of the B lymphocytes via activation of the complement cascade; antibodydependent, cell-mediated cytotoxicity; and inducing apoptosis. Because the CD20 antigen is absent in hematopoietic stem cells, rituximab treatment does not deplete these cells.

Clinical Performance. A randomized Phase III clinical trial compared the efÞcacy and safety of CHOP-R and CHOP in 399 treatment-naive elderly patients (aged 60–80; median age of 69) who had DLBCL (CoifÞer B, 2003). In this landmark trial, patients were randomly assigned to receive eight cycles of CHOP every three weeks or eight cycles of CHOP-R, whereby rituximab was administered on day 1 of each cycle of CHOP. The rituximab infusion was interrupted in the event of fever, chills, edema, congestion of the head and neck mucosa, hypotension, or any other serious adverse event and was resumed when such event was no longer occurring. Adding rituximab to CHOP resulted in higher response rates and improved event-free and overall survival (CoifÞer B, 2003). The rate of complete response (including unconÞrmed complete response—greater than 75% reduction in tumor size after therapy but a residual mass and indeterminate bone marrow status) was signiÞcantly higher in the CHOP-R group than in the CHOP group (76% versus 63%). Complete responses were seen in 52% and 37% of patients and unconÞrmed complete responses in 23% and 26% (sic) of patients, respectively. At median, two-year follow-up, event-free and overall survival times were signiÞcantly higher in the CHOP-R group without any clinically signiÞcant increase in toxicity. Two-year survival stood at 70% in the CHOP-R group versus 57% in the CHOP group (p = 0.007). Investigators attributed the longer survival in the CHOP-R group to a lower rate of disease progression during therapy and fewer relapses in patients who had a complete response. A total of 19 patients (9%)

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AGGRESSIVE NON-HODGKIN’S LYMPHOMAS

had progressive disease in the CHOP-R group compared with 43 patients (22%) in the CHOP-alone group. Three-year follow-up data presented at the American Society of Clinical Oncology (ASCO) meeting in 2003 demonstrated that three-year overall survival stood at 62% in the CHOP-R group versus 51% in the CHOP-alone group and that three-year event-free survival was 53% and 35%, respectively. In one of the studies with Þve years of follow-up, the GELA trial, R-CHOP improved overall survival by 47 percent compared to CHOP alone (a hazard ratio of 0.68, which is equivalent to a 32 percent decrease in the risk of death). U.S. Intergroup Trial 4494 is a randomized trial investigating CHOP-R versus CHOP as Þrst-line therapy in elderly patients (60+) with DLBCL, followed by rituximab maintenance therapy or observation. A corresponding press release reported on the respective trials, as follows: Wednesday, Aug 17, 2005 Genentech and Biogen Idec File Supplemental Biologics License Application for FDA Review of Rituxan for Front-Line Treatment of Intermediate Grade or Aggressive CD-20-Positive, B-Cell, Non-Hodgkin’s Lymphoma —Filing Based on Data from Three Phase III Trials in more than 1,800 Patients— South San Francisco, Calif. and Cambridge, Mass.—17 August 2005—Genentech, Inc. (NYSE: DNA), Biogen Idec, Inc. (Nasdaq: BIIB) and Roche (SWX Zurich) today announced that the companies completed the Þling of a supplemental Biologics License Application (sBLA) with the U.S. Food and Drug Administration (FDA) for an additional indication for Rituxan® (Rituximab), in previously untreated (front-line) patients with intermediate grade or aggressive, CD-20-positive, B-cell, non-Hodgkin’s lymphoma (NHL) in combination with CHOP (cyclophosphamide, doxorubicin, vincristine and prednisone) or other anthracycline-based chemotherapy regimens. As part of the Rituxan sBLA Þling, the companies have requested Priority Review designation from the FDA. Rituxan is currently approved for use in relapsed or refractory, low-grade or follicular, CD-20positive, B-cell, non-Hodgkin’s lymphoma. The sBLA Þling is based on efÞcacy and safety data from three randomized, controlled, multicenter studies of Rituxan in combination with CHOP or other anthracycline-based chemotherapy induction regimens in 1,854 previously untreated patients with intermediate grade or aggressive, CD-20-positive, B-cell, non-Hodgkin’s lymphoma. All three trials evaluated the efÞcacy endpoint of overall survival. “These three Phase III studies add to the body of data regarding Rituxan in the treatment of non-Hodgkin’s lymphoma,” said Hal Barron,

CURRENT THERAPIES

93

M.D., Genentech senior vice president, development and chief medical ofÞcer. “We are pleased to be Þling these data with the FDA, and we are committed to working closely with the agency during the review process.” “The addition of Rituxan to CHOP represents the Þrst signiÞcant improvement in intermediate grade or aggressive CD-20-positive, Bcell NHL treatment since the development of the CHOP regimen more than 30 years ago,” said Burt Adelman, M.D., Biogen Idec’s executive vice president, Development. The studies included in this Þling are ECOG 4494 (a National Cancer Institute-sponsored intergroup trial led by the Eastern Cooperative Oncology Group), GELA/LNH 98–5 (the Group d’Etude des Lymphome d’Adulte) and MInT (MabThera International Trial M39045). Study E4494 was designed to evaluate the efÞcacy and safety of Rituxan combined with induction CHOP chemotherapy in 632 patients 60 years of age or older with intermediate-grade or aggressive, CD20-positive, B-cell, NHL. The LNH 98–5 trial conducted by GELA was designed to evaluate the efÞcacy and safety of Rituxan in combination with induction CHOP chemotherapy in 399 patients 60 years of age or older with Diffuse Large B-Cell Lymphoma (DLBCL). Based on the results of this trial, in March 2002, Rituxan, which is known as MabThera® in Europe received approval from the European Union health authority to treat aggressive NHL. The M39045 (MInT) trial was designed to evaluate the efÞcacy and safety of Rituxan in combination with CHOP or other anthracyclinebased induction chemotherapy regimens in 823 patients between the ages of 18–60. Rituximab maintenance consists of four doses weekly every six months for two years. Investigators presented Þndings of a similar study at the American Society of Hematology (ASH) annual meeting in 2003. Here, the addition of rituximab to CHOP did not inßuence the overall response rate in the induction phase (78% for CHOP-R, 77% for CHOP alone). Investigators performed additional “weighted analyses” to remove any effects of subsequent maintenance therapy. These analyses found that patients who received CHOP-R experienced a signiÞcant prolongation of time to treatment failure (a 59% increase in median failure-free survival and a 45% increase in overall survival) compared with patients who received CHOP alone. Although a signiÞcant prolongation in time to treatment failure was observed in patients who received rituximab maintenance therapy, this beneÞt was predominantly conÞned to patients who received CHOP alone during the induction phase. A large international study (the MabThera International Trial [MInT]) has investigated the combination of rituximab and CHOP-like chemotherapy in

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previously untreated younger patients with aggressive NHLs. The study was closed early when a preplanned interim analysis of the study data by an independent data and safety monitoring committee (DSMC) showed a statistically signiÞcant improvement in time-to-treatment failure for patients receiving rituximab (Roche’s MabThera) plus chemotherapy. At a median observation time of two years, overall survival among patients who received rituximab in combination with chemotherapy was 95%, compared with 85% for those who received chemotherapy alone. Among patients treated with the combination, 19% failed treatment versus 42% of CHOP patients. CHOP-R patients had a signiÞcantly higher complete response rate (85% versus 65%) (Pfreundschuh MG, 2004). A retrospective analysis of 294 patients older than age 16 who were treated for DLBCL in Canada during the 18-month period on either side of rituximab’s introduction in 2001 found that outcomes improved dramatically after rituximab’s debut (Sehn LH, 2003). Nine percent of patients received experimental treatment with rituximab during the pre-rituximab period; 85% received treatment in the post-rituximab group. Two-year overall survival (OS) for all patients was 53% and 77%, respectively (p = 0.0001); for patients aged 60 or older, OS was 40% and 67%, respectively; and for patients younger than 60, OS was 69% and 87%, respectively. In a trial presented at ASCO in 2004, 122 patients with newly diagnosed stage III or IV MCL were randomized to receive treatment with either CHOP or CHOP-R (Hiddeman W, 2004). The CHOP-R group experienced a signiÞcantly higher rate of both overall and complete responses (94% versus 75% and 34% versus 7%, respectively). Time-to-treatment failure was signiÞcantly longer in the CHOP-R arm as well. Grade III and IV granulocytopenia was more frequent after CHOP-R (63% versus 53%, p = 0.01), but severe infections were few, thus mitigating the clinical relevance of this difference. Investigators at the Memorial Sloan Kettering Cancer Center in New York performed a retrospective analysis of outcomes of newly diagnosed DLBCL patients receiving rituximab-CHOP-14 (CHOP-R-14) (Halaas JL, 2004). Of the 49 patients included in the analysis, the median age was 52; 71% had advancedstage disease. According to the International Prognostic Index (IPI), 30.6% of the patients had low-risk disease, 22.4% low-intermediate-risk, 28.6% highintermediate-risk, and 18.4% high-risk. The investigators found that administering CHOP-R-14 is feasible in terms of toxicity. Most cycles were delivered on time and at full dose, except for vincristine, whose dose was reduced in one-third of the cycles. Ten patients were hospitalized a total of 21 times; neutropenic fever occurred in 3.6% (9/252) of cycles given. Eighty-two percent of the patients achieved a complete response or unconÞrmed complete response, and one patient had refractory disease. Progression-free survival at 18 months was 89%. Cisplatin, Cytarabine, and Dexamethasone (DHAP) Overview. The DHAP regimen incorporates cisplatin (Bristol-Myers Squibb’s Platinol AQ, generics); cytarabine (PÞzer’s Cytosar-U/Aracytine, Nippon

CURRENT THERAPIES

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Shinyaku’s Cylocide, generics); and dexamethasone (Merck/Banyu’s Decadron, generics). Mechanism of Action •

•

•

Cisplatin (Figure 5) is a platinum agent. Platinum agents generate highly reactive charged platinum complexes that bind covalently to DNA, leading to intrastrand, interstrand, and protein cross-linking of DNA that results in the inhibition of transcription and/or DNA replication mechanisms. Cytarabine, an analogue of deoxycytidine, is a pyrimidine antagonist antimetabolite. Antimetabolites interfere with the production of normal nucleic acids. Cytarabine activates the enzyme cytarabine triphosphatase and causes the substitution of arabinoside for a sugar molecule, thereby inhibiting DNA synthesis. Cytarabine undergoes intracellular metabolism into its active, triphosphate form: cytosine arabinoside triphosphate. This metabolite damages DNA via multiple mechanisms, which include the competitive inhibition of DNA polymerases, resulting in inhibition of DNA repair, and direct mis-incorporation into DNA. The latter mechanism is probably the most important for inducing cell death. Cytotoxicity is highly speciÞc for the S phase of the cell cycle. Dexamethasone (Figure 6) is a glucocorticosteroid. Corticosteroids reduce the inßammatory response to tumor tissue by preventing white blood cells from functioning to produce swelling and pain around the tumor site. Corticosteroids induce lysis of peripheral lymphocytes and slow lymphocyte production. Adding corticosteroids to chemotherapy regimens may augment their efÞcacy.

Clinical Performance. Of the 90 patients with progressive, recurrent, intermediate-grade NHLs who were treated with six to ten cycles of DHAP in

FIGURE 5. Structure of cisplatin.

OR O HO

OH

F O

FIGURE 6. Structure of dexamethasone (R = H).

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a Phase II trial, 34% achieved a complete response and 27% achieved a partial response (Velasquez WS, 1988). At a median follow-up of 11 months, only eight of the complete responders had relapsed. The two-year overall survival was 25%. Investigators observed a signiÞcant difference in response and survival rates for patients with low tumor burden and normal lactate dehydrogenase (LDH) compared with patients with both high tumor burden and elevated serum LDH levels. In the former group, 67% achieved a complete response and 61% survived for two years; in the latter group, no patients achieved a complete response and only 5% survived for one year. Patients with either high tumor burden with normal LDH or low tumor burden with elevated LDH achieved intermediate survival rates, compared with the survival rates of the Þrst two groups. Myelosuppression-related infection was the most frequent serious complication of this regimen (31%) and the cause of death in ten patients. Seven deaths occurred within two weeks of therapy. Details of a multicenter Phase II trial evaluating the safety and efÞcacy of DHAP in combination with rituximab (DHAP-R) for the treatment of patients with relapsed/refractory aggressive B-cell lymphoma are shown in Table 11. Methylprednisolone, Etoposide, Cytarabine, and Cisplatin (ESHAP) Overview. The ESHAP regimen incorporates methylprednisolone Medrol/Medrone/Medrate, generics); etoposide (Bristol-Myers VePesid/Etopophos, PÞzer’s Lastet, generics); cytarabine (PÞzer’s U/Aracytine, Nippon Shinyaku’s Cylocide, generics); and (Bristol-Myers Squibb’s Platinol AQ, generics).

(PÞzer’s Squibb’s Cytosarcisplatin

Mechanism of Action •

Methylprednisolone (Figure 7) is a more soluble salt of prednisolone. Prednisolone is a glucocorticosteroid. Corticosteroids reduce the inßammatory response to tumor tissue by preventing white blood cells from functioning to produce swelling and pain around the tumor site. Corticosteroids induce lysis of peripheral lymphocytes and slow lymphocyte production. Adding corticosteroids to chemotherapy regimens may augment their efÞcacy.

OR O HO

OH

O

FIGURE 7. Structure of methylprednisolone (R = H).

CURRENT THERAPIES

R


O O HO

97

O O OH O O O O CH3O

OCH3 RO

FIGURE 8. Structure of etoposide (R = H, R1 = CH3 ).

•

•

•

Etoposide (Figure 8) is an epipodophyllotoxin. Epipodophyllotoxins cause single-strand breaks in DNA and inhibit DNA topoisomerase II to cause metaphase arrest, thereby inducing cytotoxic activity. Etoposide is speciÞc to the cell-cycle phase; its predominant activity occurs in the late S phase and G2 phase. Cytarabine, an analogue of deoxycytidine, is a pyrimidine antagonist antimetabolite. Antimetabolites interfere with the production of normal nucleic acids. Cytarabine activates the enzyme cytarabine triphosphatase and causes the substitution of arabinoside for a sugar molecule, resulting in the inhibition of DNA synthesis. Cytarabine undergoes intracellular metabolism into its active triphosphate form: cytosine arabinoside triphosphate. This metabolite damages DNA via multiple mechanisms, which include the competitive inhibition of DNA polymerases, resulting in inhibition of DNA repair, and direct mis-incorporation into DNA. The latter mechanism is probably the most important for inducing cell death. Cytotoxicity is highly speciÞc for the S phase of the cell cycle. Cisplatin (Figure 5) is a platinum agent. Platinum agents generate highly reactive, charged platinum complexes that bind covalently to DNA, leading to intrastrand, interstrand, and protein cross-linking of DNA that results in the inhibition of transcription and/or DNA replication mechanisms.

Clinical Performance. Researchers at the M.D. Anderson Cancer Center in Houston, Texas, reviewed the Þndings from 122 patients with relapsed or refractory NHLs who received treatment with one cycle of ESHAP (Velasquez WS, 1994). The Þrst 63 patients were randomized to treatment with or without cisplatin (ESHA or ESHAP); however, because of the marked differences in response rates between the two treatment arms (33% versus 75%), the ESHA arm was discontinued and all subsequent patients received ESHAP. Of 122 patients receiving ESHAP, 37% had a complete remission and 27% had a partial remission, for an overall response rate of 64%. The median duration of complete response was 20 months. At three years, 28% of patients remained

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in a state of complete response. The overall median survival duration was 14 months; the survival rate at three years was 31%. At 40 months, 10% of all patients were alive and disease-free. Response and survival rates were similar in patients with low-grade (34 patients), intermediate-grade (67), transformed (18), and high-grade (3) lymphomas. Investigators observed a signiÞcant difference in survival for patients with normal LDH levels and low or intermediate tumor burden and patients with low tumor burden and elevated LDH levels (55% threeyear survival rate) versus patients with elevated LDH levels and intermediate or high tumor burden (<20% three-year survival rate). The major toxicities included myelosuppression, with a median granulocyte nadir of 500/microL, and median platelet nadir of 70,000/microL. Methotrexate, Leucovorin, Doxorubicin, Cyclophosphamide, Vincristine, Bleomycin, and Prednisone (MACOP-B) Overview. The MACOP-B regimen incorporates methotrexate (generics); leucovorin (generics); doxorubicin (PÞzer’s Adriamycin/Adriblastine, Kyowa’s Adriacin, generics); cyclophosphamide (Bristol-Myers Squibb’s Cytoxan, Baxter’s Endoxan/Endoxana, generics); vincristine (Eli Lilly/EG Labo’s Oncovin, generics); bleomycin (Bristol-Myers Squibb’s Blenoxane/Nippon Kayaku’s Bleo, generics); and prednisone (Merck’s Decortin, generics). Mechanism of Action •

•

Methotrexate (Figure 9), a structural analogue of folate, is an antimetabolite. Antimetabolites block normal DNA synthesis by inhibiting several key enzymes. Methotrexate inhibits the enzyme dihydrofolate reductase. This inhibition interferes with maintenance of the reduced folate pool. Reduced folates are essential for de novo synthesis of thymidylate and purine nucleotides, which is required for DNA synthesis and cell replication. Thus, by inhibiting dihydrofolate reductase, methotrexate stops the replication of rapidly proliferating cells such as tumor cells. Leucovorin (Figure 10) is usually used 24 hours after methotrexate to selectively rescue normal cells from the adverse effects of methotrexate caused by inhibiting the production of reduced folates. This agent is not used

H 2N

N N

N

N

CH3 N H N

NH2 O

COOH COOH

FIGURE 9. Structure of methotrexate.

CURRENT THERAPIES

H2N

N

HN

H N N

O

99

NH CH2

NH

CHO

CH2

C

CH

O

COOH

COOH

CH2

FIGURE 10. Structure of leucovorin.

•

•

•

•

•

simultaneously with methotrexate because it might then nullify methotrexate’s therapeutic effect. Doxorubicin (Figure 2) is an anthracycline. Anthracyclines interact with several different cellular targets, most importantly topoisomerase II. By inhibiting this DNA religation enzyme, anthracyclines exert their cytotoxic effect. Another doxorubicin mechanism that leads to cell death is known as DNA intercalation, in which the anthracycline molecule inserts itself between base pairs. This phenomenon causes single- and double-stranded breaks in DNA that inhibit cell proliferation. The free radicals generated by the reductive metabolism of anthracyclines may also damage cellular structures such as proteins and DNA. Cyclophosphamide (Figure 1) is an alkylating agent and a cyclic phosphamide ester of mechlorethamine that is activated in the liver. Alkylation of DNA causes both strand breaks and the formation of cross-links between strands of DNA. Both of these events result in aberrant DNA replication and transcription of RNA. Cyclophosphamide, like other alkylating agents, is nonspeciÞc to the cell-cycle phase. However, rapidly proliferating cells are more susceptible to the action of alkylating agents because of the reduced time available for DNA enzymes to repair the cytotoxic damage. Vincristine is a vinca alkaloid. Vinca alkaloids bind with microtubular proteins of the mitotic spindle, thereby leading to mitotic arrest or cell death. Vinca alkaloids exert selective toxicity to tumor cells by acting on proliferating cells. Bleomycin is an antitumor antibiotic. It causes single-strand and, less frequently, double-strand DNA breaks through formation of an intermediate iron complex. Bleomycin inhibits DNA synthesis and, to a lesser degree, RNA and protein synthesis. The action of bleomycin is speciÞc to the cellcycle phase. Prednisone is a glucocorticosteroid. Glucocorticosteroids reduce the inßammatory response to tumor tissue by preventing white blood cells from functioning to produce swelling and pain around the tumor site. Corticosteroids induce lysis of peripheral lymphocytes and slow lymphocyte production. Adding corticosteroids to chemotherapy regimens may augment their efÞcacy.

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Clinical Performance. The most relevant clinical trial data for MACOP-B come from a trial that compared CHOP with MACOP-B and two other regimens. In 1993, SWOG and ECOG conducted a prospective, randomized Phase III trial that compared CHOP with three regimens that seemed to offer improved survival over CHOP in Phase II trials (Fisher RI, 1993). The trial involved 899 treatment-naive patients in advanced stages of intermediate-grade or highgrade NHLs. Patients in each arm received a maximum of eight cycles of therapy. The three investigational regimens were as follows: m-BACOD (low-dose methotrexate with leucovorin rescue, bleomycin, doxorubicin, cyclophosphamide, vincristine, and dexamethasone); ProMACE-CytaBOM (prednisone, doxorubicin, cyclophosphamide, and etoposide, followed by cytarabine, bleomycin, vincristine, and methotrexate with leucovorin rescue); and MACOP-B (methotrexate with leucovorin rescue, doxorubicin, cyclophosphamide, vincristine, prednisone, and bleomycin). The median ages of the groups ranged from 54 to 57 years; approximately 25% of the patients were age 65 or older. At three years, 44% of all patients were alive without disease; investigators observed no signiÞcant differences in disease-free survival between the groups: 41% in the CHOP and MACOP-B groups and 46% in the m-BACOD and ProMACE-CytaBOM groups (p = 0.35). Overall survival at three years was 52% (50% in the ProMACE-CytaBOM and MACOP-B groups, 52% in the m-BACOD group, and 54% in the CHOP group; p = 0.90). Researchers found no subgroup of patients in which survival was superior in a non-CHOP arm. No signiÞcant differences occurred in the response rates of the four treatment groups. The rates of objective antitumor responses were 80% in the CHOP group, 82% in the m-BACOD group, 83% in the MACOP-B group, and 87% in the ProMACE-CytaBOM group. The rates of complete response were 44% for CHOP, 48% for m-BACOD, 56% for ProMACE-CytaBOM, and 51% for MACOP-B; the rates of partial response were 36% for CHOP, 34% for mBACOD, 31% for ProMACE-CytaBOM, and 32% for MACOP-B (Fisher RI, 1993). Mesna, Ifosfamide, Mitoxantrone, and Etoposide (MINE) Overview. The MINE regimen incorporates mesna (Bristol-Myers Squibb’s Mesna/Mesnex, Baxter/Shionogi’s Uromitexan); ifosfamide (Bristol-Myers Squibb’s Ifex, Baxter’s Holoxan/Mitoxana, Shionogi’s Ifomide, generics); mitoxantrone (Serono/Wyeth/Wyeth-Takeda’s Novantrone, Baxter’s Onkotrone); and etoposide (Bristol-Myers Squibb’s VePesid/Etopophos, PÞzer’s Lastet, generics). Mechanism of Action •

Mesna reacts with acrolein and other urotoxic metabolites of oxazaphosphorines (cyclophosphamide or ifosfamide) to form stable, non-urotoxic compounds. Mesna does not have any antitumor activity, nor does it appear to interfere with the antitumor activity of antineoplastic drugs. Rather, it protects against the bladder toxicity of ifosfamide and cyclophosphamide.

CURRENT THERAPIES

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•

•

101

Ifosfamide is an alkylating agent pro-drug and a structural analogue of cyclophosphamide. After being metabolized in the liver to its active form, ifosfamide binds DNA and transfers alkyl groups, causing DNA cross-links and strand breaks. As a result, this agent prevents DNA replication and causes tumor cell death. Mitoxantrone is an antitumor antibiotic. It is an anthracinedione and structural analogue of the antibiotic anthracyclines. Mitoxantrone’s exact mechanism of action is unknown, but it includes intercalation with DNA to cause interstrand and intrastrand cross-linking. Mitoxantrone also causes DNA strand breaks through binding with the phosphate backbone of DNA. The agent is nonspeciÞc to the cell-cycle phase. Mitoxantrone also impairs the strand-reunion reaction of topoisomerase II. These effects result in the production of protein-linked, double-strand DNA breaks. Although mitoxantrone is cytotoxic to cells throughout the cell cycle, cells in late S phase are more sensitive. Etoposide is an epipodophyllotoxin. Epipodophyllotoxins cause single-strand breaks in DNA and inhibit DNA topoisomerase II to cause metaphase arrest, thereby inducing cytotoxic activity. Etoposide is speciÞc to the cell-cycle phase; its predominant activity occurs in the late S phase and G2 phase.

Clinical Performance. Forty-eight patients with recurring intermediate- or high-grade lymphomas received a maximum of six cycles of MINE (Rodriguez MA, 1995). Patients had received a median of two prior regimens (range 2–5). Forty-eight percent of patients responded, and 21% achieved a complete response. Median survival was nine months. For patients who achieved a complete response, median time to treatment failure was 12 months, versus 5 months for patients who achieved only partial response. The most signiÞcant toxicity was myelosuppression. Twenty-Þve episodes of neutropenic fever occurred, and two patients died of sepsis. Three patients experienced bleeding associated with thrombocytopenia. Carmustine, Etoposide, Cytarabine, and Melphalan (BEAM) Overview. The BEAM regimen incorporates carmustine (Bristol-Myers Squibb’s BiCNU/Carmubris); etoposide (Bristol-Myers Squibb’s VePesid/Etopophos, PÞzer’s Lastet, generics); cytarabine (PÞzer’s Cytosar-U/Aracytine, Nippon Shinyaku’s Cylocide, generics); and melphalan (GlaxoSmithKline’s Alkeran). Mechanism of Action •

Carmustine (Figure 11) is a nitrosourea-alkylating agent. Metabolites of carmustine cause alkylation and cross-linking of DNA, which prevent DNA replication and cell division. Other biological effects include inhibition of DNA repair. Nitrosoureas generally lack cross-resistance with other alkylating agents.

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FIGURE 11. Structure of carmustine.

Cl

H 2N COOH N

Cl FIGURE 12. Structure of melphalan.

•

•

•

Etoposide (Figure 8) is an epipodophyllotoxin. Epipodophyllotoxins cause single-strand breaks in DNA and inhibit DNA topoisomerase II to cause metaphase arrest, thereby inducing cytotoxic activity. Etoposide is speciÞc to the cell-cycle phase; its predominant activity occurs in the late S phase and G2 phase. Cytarabine, an analogue of deoxycytidine, is a pyrimidine antagonist antimetabolite. Antimetabolites interfere with the production of normal nucleic acids. Cytarabine activates the enzyme cytarabine triphosphatase and causes the substitution of arabinoside for a sugar molecule, resulting in inhibition of DNA synthesis. Cytarabine undergoes intracellular metabolism into its active triphosphate form: cytosine arabinoside triphosphate. This metabolite damages DNA by multiple mechanisms, which include the competitive inhibition of DNA polymerases, resulting in the inhibition of DNA repair, and direct mis-incorporation into DNA. The latter mechanism is probably the most important for inducing cell death. Cytotoxicity is highly speciÞc for the S phase of the cell cycle. Melphalan (Figure 12) is an alkylating agent and a phenylalanine derivative of mechlorethamine. Alkylation of DNA causes both strand breaks and the formation of cross-links between strands of DNA. Both of these lesions result in aberrant DNA replication and transcription of RNA. Melphalan, like other alkylating agents, is nonspeciÞc to the cell-cycle phase. However, rapidly proliferating cells are more susceptible to the action of alkylating agents because of the reduced time available for DNA enzymes to repair the cytotoxic lesion.

Clinical Performance. Clinicians use BEAM to destroy malignant cells prior to autologous stem-cell transplantation (ASCT). BEAM is the most commonly used form of high-dose therapy (HDT) in the seven major pharmaceutical markets

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under study. The clinical performance of HDT followed by autologous bone marrow transplantation or ASCT is described in the following section. Nonpharmacological Approaches Radiotherapy. In patients with Ann Arbor stage I or localized stage II aggressive NHLs, adding radiotherapy to CHOP chemotherapy increases survival compared with CHOP alone. In a Phase III trial conducted by SWOG, 401 patients with localized aggressive NHLs were randomized to receive eight cycles of CHOP alone or three cycles of CHOP and 4–5.5 Gy involved-Þeld radiotherapy (Miller TP, 1998). Five-year survival was 82% in the combined modality arm and 72% in the CHOP-only arm, and fewer life-threatening toxicities were observed in the radiotherapy/CHOP arm (31% of patients) than in the CHOP-only arm (40%). Local radiotherapy is not generally used in the treatment of advanced aggressive NHLs. Bone Marrow Transplantation Following the results of a randomized trial, autologous bone marrow transplantation (ABMT) became the treatment of choice for relapsed aggressive NHLs in chemotherapy-sensitive patients who are young and Þt enough to undergo this arduous treatment (Philip T, 1995). Hematopoietic stem cells were traditionally harvested from bone marrow but are now more commonly obtained from peripheral blood. Accordingly, the procedure is usually described as autologous stem-cell transplantation (ASCT) rather than ABMT. The purpose of stem-cell transplantation is to reestablish bone marrow function after HDT and radiation therapy have been administered to destroy malignant cells. However, with available second-line regimens, only 50% of patients respond well enough to undergo transplantation. The randomized trial involved 109 patients who responded to reinduction chemotherapy and were randomized to receive either consolidation chemotherapy and radiotherapy, or radiotherapy and HDT, followed by an ASCT (Philip T, 1995). Five-year event-free survival was 46% in the ASCT group and 12% in the chemotherapy group; overall Þve-year survival was 53% and 32%, respectively. The incidence of fatal toxic events was higher in the ASCT group than in the chemotherapy group. This high treatment-related mortality restricts the use of ASCT to young (less than 60 years old), Þt patients. Patients with aggressive NHLs who have a high risk of relapsing (high IPI score) may also beneÞt from Þrst-line HDT and ASCT. In a randomized Phase III trial, 451 patients in their Þrst complete remission were randomized to receive additional treatment to consolidate their response—either sequential chemotherapy or ASCT. The two groups did not have signiÞcantly different disease-free survivals. However, a retrospective subset analysis found that patients with two or three IPI risk factors who were treated with ASCT enjoyed signiÞcantly improved survival and disease-free survival rates compared with the chemotherapy-treated

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patients (Haioun C, 2000). Eight-year survival was 64% for the ASCT arm and 59% for the chemotherapy arm; eight-year disease-free survival was 55% and 39%, respectively. Again, because of the toxicities associated with ASCT, this treatment is considered only for Þt patients younger than age 60.

EMERGING THERAPIES A wide range of promising agents are in late stages of development for nonHodgkin’s lymphomas (NHLs). They include monoclonal antibodies (MAbs), molecularly targeted agents, and modiÞed cytotoxics. MAbs dominate the Þeld, both in their unconjugated form as radioimmunotherapy and conjugated with toxins. Herein, we focus on agents in development for aggressive NHLs. We cover therapies for diffuse large B-cell lymphoma (DLBCL), mantle-cell lymphoma (MCL), Burkitt’s lymphoma, peripheral T-cell lymphoma, and transformed disease (patients initially diagnosed with indolent disease whose disease has transformed clinically and histologically to an aggressive subtype, usually DLBCL). Development efforts are focusing primarily on DLBCL, the largest histological subtype of NHL, and MCL, a subtype with a particularly poor prognosis, the worst prognosis being: disease-free and overall survival rates of 11% and 27%, respectively. Barriers to entry are low for noncytotoxics with side effects that do not overlap with those of CHOP (cyclophosphamide, doxorubicin, vincristine, and prednisone) chemotherapy, e.g., myelosuppression, cardiotoxicity, neurotoxicity. For example, uptake of rituximab (Genentech/Biogen Idec/Chugai’s Rituxan, Roche’s MabThera) has been extremely rapid in most geographic markets. This reference does not cover some agents in development for aggressive NHLs because few data are available. Among these agents are: temsirolimus (Wyeth Research’s CCI 779), and interleukin-12, each for MCL. Table 9 summarizes the drug therapies in development for aggressive NHLs. Table 10 outlines the achievements of currently marketed agents. We include this table as a benchmark for comparison of experimental agents with marketed agents. Radioimmunotherapies Overview. Radioimmunotherapy (RIT) was conceived to exploit the radiosensitivity of NHLs while reducing the toxicity associated with total body irradiation. The partial success of unlabeled immunotherapy in the treatment of aggressive NHLs provides a rationale for the use of radiolabeled MAbs. Investigations are under way to establish the optimal timing of treatment and the best way to combine RIT with standard and new therapies. Two anti-CD20 RITs are in development for aggressive NHLs: yttrium-90labeled ibritumomab tiuxetan (Biogen Idec/Schering AG’s Zevalin) and iodine131-labeled tositumomab (Corixa/GlaxoSmithKline/GE Healthcare’s Bexxar). A

EMERGING THERAPIES

105

TABLE 9. Emerging Therapies in Development for Aggressive Non-Hodgkin’s Lymphomas Compound

Development Phase

Marketing Company

II II I

Biogen Idec Schering AG Biogen Idec

Iodine-131-labeled tositumomab (Bexxar) United States Europe Japan

II II —

GlaxoSmithKline GE Healthcare —

Yttrium-90-labeled epratuzumab (LymphoCide/LymphoCide Y-90) United States Europe Japan

III II —

Immunomedics Immunomedics —

Unlabeled monoclonal antibodies Epratuzumab (LymphoCide; Immu-103) United States Europe Japan

II II —

Immunomedics Immunomedics —

Antisense oligonucleotides Oblimersen (Genasense) United States Europe Japan

II — —

Genta — —

Proteasome inhibitors Bortezomib (Velcade, PS-341) United States Europe Japan

II II —

Millennium Pharmaceuticals Millennium Pharmaceuticals —

Cytotoxic agents Sphingosomal vincristine (Onco-TCS) United States Europe Japan

PR — —

Enzon/Inex Pharmaceuticals — —

III III —

Novuspharma — —

II — —

Fujisawa/ National Cancer Institute — —

Radioimmunotherapies Yttrium-90-labeled ibritumomab tiuxetan (Zevalin) United States Europe Japan

Pixantrone (formerly BBR-2778) United States Europe Japan Histone deacetylase inhibitors Depsipeptide (FR-901228) United States Europe Japan

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TABLE 9. (continued) Compound Immunotoxins Denileukin diftitox (Ontak) United States Europe Japan

Development Phase

Marketing Company

II — —

Seragen — —

Vaccines Recombinant idiotype-KLH vaccine (MyVax) United States II Europe — Japan —

Genitope — —

PR = Preregistered.

third anti-CD22 RIT in development for aggressive NHLs is yttrium-90-labeled epratuzumab (Immunomedics’ LymphoCide/LymphoCide Y-90). No randomized data are available comparing the two agents in late-stage development for aggressive NHLs (ibritumomab [Biogen Idec/Schering AG’s Zevalin] and tositumomab [Corixa/GlaxoSmithKline/GE Healthcare’s Bexxar]) (ibritumomab and tositumomab). Corixa and GlaxoSmithKline are planning an open-label Phase III trial to compare the safety and efÞcacy of ibritumomab and tositumomab in patients with indolent NHLs. Uptake of these agents will be hampered by logistical and patient ownership issues (oncologists must refer patients to nuclear medicine specialists), cost, and concern about toxicities. However, if radiolabeled antibodies are used in place of high-dose therapy (HDT), a procedure that carries a great risk of secondary malignancy, the risk of secondary malignancy is likely to decline. Mechanism of Action. Radioimmunotherapy uses MAbs directed against antigens on the tumor surface to target radiation to the tumor. Radiolabeled antibodies have three advantages over unlabeled antibodies. First, the radiolabeled antibodies do not heavily rely on an often-deÞcient host immune system; second, the radioisotopes emit beta particles that induce lethal DNA damage both to the cell targeted by the antibody and to nearby cells that are inaccessible to the antibody or express insufÞcient antigen (known as a bystander or “cross Þre” effect); third, the tumor-killing effects of the beta particles are powerful, yielding higher overall response rates and longer remission durations than were achieved with unconjugated antibodies. The potential beneÞts of high-dose treatment with targeted, radiolabeled antibodies rather than total body irradiation include the potential to deliver higher doses of radiation to the tumor and lower doses to normal organs, with a consequent reduction in the incidence of secondary malignancy resulting from radiation exposure (most notably, myelodysplastic syndrome [MDS] and acute myeloid leukemias). In previous studies of total body irradiation, observed incidence rates

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TABLE 10. Achievements of Current Treatment Approaches in Aggressive Non-Hodgkin’s Lymphomas: Benchmarks for Evaluating Emerging Therapies Three-Year Overall

Two-Year

Disease-Free Event-Free

Treatment

Response

Survival/

Modality

Complete

Overall

Overall

Principal

or Regimen

Response (%)

Survival

Survival

Toxicities (%)

80/44

41/54

—

Fatal toxicity (1) Life-threatening (grade 4) toxicity (31)

CHOP-R Elderly patients with diffuse large B-cell lymphoma (DLBCL)b

82/75c

—

57/70

Grade 3 or 4 cardiac toxicity (8) Grade 3 or 4 infection (12)

Elderly patients with DLBCLb

CHOP

69/63c

—

38/57

Grade 3 or 4 cardiac toxicity (8) Grade 3 or 4 infection (20)

Second-line treatment of patients with aggressive NHLs (progressive recurrent disease)d

DHAP

61/34

—

—/25

Infection (31) Fatal infection (9)

Second-line treatment of patients with aggressive NHLs (recurrent or refractory disease), median age 63 yearse

DHAP-R

48/28

—

—

Hematologic Renal Nausea/vomiting

Mantle-cell lymphoma (MCL), first-line treatmentf

HyperCVAD-R

—/89

—

72/90

Setting First-line treatment of intermediate- or high-grade non-Hodgkin’s lymphomas (NHLs), bulky stage II, stage III, or stage IV disease, all agesa

CHOP

Survival/

Grade 4 hematologic toxicity (60) Grade 3 infections (14)

a Fisher R, 1993. b Coiffier B, 2002. c Includes unconfirmed complete response. d Velasquez WS, 1988. e Mey UJM, 2003. f Romaguera J, 2003. Note: Full source citations appear in ‘‘References.’’ CHOP = Cyclophosphamide, doxorubicin, vincristine, prednisone. CHOP-R = Cyclophosphamide, adriamycin, vincristine, prednisone. DHAP = Cisplatin, cytarabine, dexamethasone. DHAP-R = Cisplatin, cytarabine, dexamethasone, rituximab. Hyper-CVAD-R = Cyclophosphamide, doxorubicin, vincristine, dexamethasone, rituximab.

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of secondary malignancies ranged from 4% to 8%, with incidence projections of 12–20% at 20 years post-transplantation (Press OW, 2000). The incidence of secondary malignancy associated with radiolabeled antibodies has not yet been established because most trials have treated patients who had previously undergone chemotherapy and radiation. Yttrium-90-Labeled Ibritumomab Tiuxetan. Yttrium-90-labeled ibritumomab tiuxetan (Biogen Idec/Schering AG’s Zevalin) is a murine CD20 MAb attached to yttrium-90 as a radiation source. 90 Y-ibritumomab tiuxetan has been launched in the United States for relapsed or refractory, low-grade, follicular or transformed B-cell NHLs. It is preregistered in Europe for the same indication; a positive opinion from the Committee of Proprietary Medicinal Products (CPMP) was issued in September 2003. Several Phase II trials for aggressive lymphomas are under way. 90 Y-ibritumomab tiuxetan uses MAbs directed against CD20 antigens on the tumor surface to target radiation to the tumor. Yttrium-90 has the following advantages over iodine-131: •

• •

•

It emits a particle that has higher energy and a longer path length than iodine-131, thereby enabling deeper tissue penetration, which is potentially valuable in the case of bulky tumors. It is a more stable conjugate than iodine-131, with no elution of the radioactive conjugate from the antibody. Because all radioactivity is targeted to the tumor, dosimetry (calculation of patient-speciÞc radiation dose from internally administered radionucleotides) is not required. As a pure beta emitter, yttrium-90-based treatment can be administered on an outpatient basis.

The role of 90 Y-ibritumomab tiuxetan in aggressive NHLs has not yet been deÞned. Small trials have investigated the use of ibritumomab in three distinct settings: as single-agent salvage therapy for patients with B-cell lymphomas; in patients with MCL; and in combination with HDT and autologous stem-cell transplantation (ASCT). Additionally, retrospective analyses have investigated the activity of 90 Y-ibritumomab tiuxetan in patients with transformed disease, and the most appropriate timing of 90 Y-ibritumomab tiuxetan treatment relative to chemotherapy and unconjugated antibodies. In the salvage setting, in patients with advanced B-cell NHLs, a Phase I/II dose-Þnding study found that 90 Y-ibritumomab tiuxetan has good single-agent activity (Gordon LI, 2004). Two patients with relapsed DLBCL experienced complete responses lasting more than Þve years. Fifty-one patients (median age 58) with relapsed or refractory B-cell lymphoma (including three with diffuse mixed histology and nine with diffuse large-cell histology) received 5 mCi 90 Yibritumomab tiuxetan on day 1 for imaging and 0.3–0.4 mCi/kg 90 Y-ibritumomab tiuxetan on day 8 (after preinfusion with rituximab to clear peripheral B cells).

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The median number of prior regimens was two; all patients had received CHOP or CHOP-like regimens. Investigators noted an overall response rate of 58%, with 33% of patients achieving a complete response. This result compares favorably with results achieved in the salvage setting by DHAP (cisplatin, cytarabine, and dexamethasone) plus rituximab: 48% overall response rate and 28% complete response (Mey UJM, 2003). The median duration of response had not been reached at 35.5 months of follow-up. Median time to next anticancer treatment for all patients was 9.9 months. Toxicity was primarily hematologic: grade 4 neutropenia and grade 4 thrombocytopenia occurred in 17% and 8% of patients, respectively. A retrospective analysis of data from 15 patients with transformed (previously indolent) B-cell NHLs, presented at the American Society of Clinical Oncology (ASCO) meeting in 2002, demonstrated 90 Y-ibritumomab tiuxetan’s activity in this subpopulation (Bartlett NL, 2002). In one of four trials, the patients had been treated with 90 Y-ibritumomab tiuxetan using the regimen described in the preceding paragraph. Inclusion criteria for the four trials were as follows: less than 25% bone marrow involvement, absolute neutrophil count greater than 1,500/mm3 , and no prior myeloablative therapy. Median patient age was 60 and the median number of prior therapies was two. Toxicity was primarily hematologic: grade 4 neutropenia (40% of patients) and thrombocytopenia (20%). Investigators noted an overall response rate of 53%; 13% of all patients achieved a complete response. The median time to progression was 8.5 months, with two patients remaining in remission at 31 and 37 months, respectively. With a view to improving survival rates, three trials have investigated the role of 90 Y-ibritumomab tiuxetan in combination with HDT and ASCT in heavily pretreated patients. The Þrst, a Phase I/II trial, combined high-dose 90 Y-ibritumomab tiuxetan (40–100 mCi) with etoposide (40–60 mg/kg) and cyclophosphamide (100 mg/kg) (Nademanee A, 2002). The RIT protocol involved pretreatment with rituximab (to deplete B cells from the peripheral circulation and optimize levels of 90 Y-ibritumomab tiuxetan at the tumor site) and Indium (In)-111 ibritumomab (for dosimetry calculations). Twenty-six patients entered the trial; 18 advanced to HDT. The remaining patients achieved inadequate uptake at tumor sites, had progressive disease before treatment, or had an allergic reaction to rituximab (one patient). Nine patients had follicular lymphoma, seven had DLBCL, and two had MCL. All but two patients had previously received rituximab treatment. No treatment-related deaths occurred, and all patients engrafted; the median time to reach the target neutrophil and platelet counts was 10 days and 18 days, respectively. All seven patients with active disease at transplant achieved complete remission. Seventeen patients were alive in remission at a median follow-up of eight months. Both one-year estimated survival and disease-free survival are 92%. Investigators concluded that adding high-dose 90 Y-ibritumomab tiuxetan to etoposide and cyclophosphamide does not increase toxicity or delay engraftment, and that the combination is effective in heavily pretreated patients with refractory B-cell NHLs.

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The second HDT trial of 90 Y-ibritumomab tiuxetan is ongoing. In this trial, investigators are combining 90 Y-ibritumomab tiuxetan with high-dose BEAM (carmustine, etoposide, cytarabine, and melphalan) and autologous peripheral blood progenitor-cell transplant (Winter JN, 2002). In 12 relapsed or refractory patients with CD20+ aggressive NHLs who received treatment, engraftment was rapid. At a median follow-up of 15 months, four deaths had occurred, resulting in one-year survival of 75%. Two-year survival was also 75%; two-year progressionfree survival was 54%. The third trial, a pilot study, is investigating the combination of standard-dose 90 Y-ibritumomab tiuxetan with high-dose BEAM for older patients in order to achieve targeted intensiÞcation of the conditioning regimen (Fung HC, 2003). Twelve patients (median age 66 years) with high-risk, aggressive CD20+ B-cell lymphomas underwent treatment. Two patients had received prior dose-limiting radiation. Seven patients had DLBCL; Þve had MCL. After imaging with In-111ibritumomab, patients received 90 Y-ibritumomab tiuxetan at a Þxed dose of 0.4 mCi/kg (without dosimetric guidance) with their conditioning regimen of highdose BEAM. Prior to transplantation, Þve patients had failed induction therapy, three were in second complete response, and two were in third complete response. All patients tolerated treatment well, with only two grade 3 or 4 gastrointestinal (GI) toxicities. One patient developed steroid-responsive pneumonitis 17 days after transplantation. All patients engrafted promptly; the median time to reaching both target neutrophil and platelet counts was 11 days. The median total dose of 90 Y-ibritumomab tiuxetan was 32 mCi (range: 20.7–40). At a median follow-up of nine months, one patient with MCL had died of progressive disease, but the remaining 11 patients were well, without evidence of lymphoma in computer-aided tomography (CT) and positron-emission tomography (PET) scans. Investigators concluded that this approach is well tolerated and that results look promising, given the age and refractory disease status of the trial participants. A retrospective analysis examined the positioning of RIT relative to chemotherapy and unconjugated antibodies (Ansell SM, 2002). Fifty-eight patients (most had indolent NHLs, but 14 had transformed, follicular NHLs) received single-agent 90 Y-ibritumomab tiuxetan followed (after relapse) by a median of two additional therapies. The study found that ibritumomab does not compromise subsequent chemotherapy or ASCT. Toxicity with subsequent therapies is similar to toxicity in patients not treated with 90 Y-ibritumomab tiuxetan. A Phase II trial has assessed the activity of 90 Y-ibritumomab tiuxetan in heavily pretreated patients with relapsed MCL. Twelve patients received a single dose of 90 Y-ibritumomab tiuxetan. Patients with platelet counts higher than 500,000/mm3 received 0.4 mCi/kg (maximum dose 32 mCi); patients with lower platelet counts received 0.3 mCi/kg. The median patient age was 64, the median number of prior regimens was three, and 50% of patients had previously been treated with Hyper-CVAD (cyclophosphamide, doxorubicin, vincristine, and dexamethasone). Three patients had complete remissions, and one had a partial remission, for an overall response rate of 33%; these four patients’ responses lasted for six, nine, Þve, and four months, respectively. Two other patients had

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stable disease for 11 and 6 months, respectively. Responses were observed in patients receiving the higher dose and in patients receiving the lower dose. Investigators described the treatment as well tolerated; the most common toxicity was hematologic. Three patients required between one and three platelet transfusions, and one patient was hospitalized with neutropenic fever. 90 Y-ibritumomab tiuxetan’s limitations include its toxicity, high price, and relatively cumbersome administration protocol (predosing with unlabeled antibody, followed a week later by a second infusion of unlabeled antibody and a single injection of labeled antibody, followed by weekly monitoring of platelet counts until hematopoietic recovery). The toxicity of 90 Y-ibritumomab tiuxetan is primarily hematologic. A distinctive feature of 90 Y-ibritumomab tiuxetan’s hematologic toxicity is that it occurs seven to nine weeks after treatment (compared with a matter of days for chemotherapy). Toxicity correlates with the number of prior treatments and the extent of bone marrow involvement; patients with more than 25% bone marrow involvement are ineligible for treatment. Infections occur in 29% of patients; severe infections occur in 5% of patients, with 7% requiring hospitalization (Witzig TE, 2003). The risk of serious infections is mitigated by the fact that ibritumomab does not cause severe mucositis (unlike most cytotoxics), a condition that allows infectious pathogens to enter the body. Myelodysplasia or acute myelogenous leukemia occurred in 1% of patients 1–34 months after treatment. As mentioned previously, the most appropriate timing for RIT in treating aggressive NHLs has not yet been established. However, on the basis of available trial data, 90 Y-ibritumomab tiuxetan could be approved in 2005 for second-line treatment of MCL, for second-line treatment of patients with relapsed or refractory disease who are ineligible for HDT and ASCT, and for third-line treatment of other patients. Iodine-131-Labeled Tositumomab. Iodine-131-labeled tositumomab (GlaxoSmithKline/GE Healthcare’s Bexxar) is a cell-speciÞc, anti-CD20 murine MAb labeled with iodine-131. In July 2003, it launched in the United States for the treatment of patients with CD20+, follicular NHLs, with or without transformation, whose disease is refractory to rituximab and has relapsed following chemotherapy. 131 I-tositumomab was granted orphan drug status in Europe for follicular lymphoma in February 2003. 131 I-tositumomab uses MAbs directed against CD20 antigens on the tumor surface to target radiation to the tumor. Iodine-131 has the following advantages over yttrium-90: • • •

The radio-iodination methods available for antibody labeling are relatively simple. Iodine-131’s cost is low, compared with the cost of yttrium-90. It emits gamma emissions (in addition to the toxic beta emissions), thus enabling imaging of the patient and localization of the radioimmunoconjugate to assess tumor biodistribution.

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The third advantage is offset by the need for hospitalization to control the risks from increased radiation exposure and the potential hazard posed to health care workers. Unlike yttrium-90-labeled antibodies, the metabolism of iodine-131labeled antibodies results in the release of free iodine-131 and iodine-131 tyrosine into the blood. The RIT protocol for 131 I-tositumomab involves a dosimetric dose with an unlabeled antibody given prior to a trace dose of iodine-131-labeled antibody. For individualized dose calculation, three whole-body gamma camera images are taken during the course of the week following the dosimetric dose to calculate patient-speciÞc elimination of radioactivity. The therapeutic dose is administered seven days after the dosimetric dose. The therapeutic dose is calculated to deliver a maximum dose of 75 cGy (attenuated to 65 cGy for patients with low platelet counts). Again, a predose of unlabeled antibody is given to deplete B cells from the peripheral circulation and to optimize 131 I-tositumomab levels at the tumor site; the predose is followed by the labeled antibody. In patients with aggressive disease, 131 I-tositumomab has been studied in several settings: at myeloablative doses in combination with HDT and ASCT; at nonmyeloablative doses following chemotherapy for response consolidation; as a single agent for patients with recurrent disease; and as Þrst-line treatment for patients with MCL. A subgroup analysis has examined the safety of 131 I-tositumomab for elderly patients. Before we describe trials involving patients with de novo aggressive disease (i.e., patients whose initial disease is an aggressive form of NHLs), we will examine data from trials that focused primarily on patients with indolent—as opposed to aggressive—disease but also included patients with transformed disease. (Transformed disease would have initially diagnosed as indolent but transformed clinically and histologically to an aggressive subtype.) We discuss these trials because relatively few patients with de novo aggressive disease have received treatment with 131 I-tositumomab and because patients with transformed disease usually have DLBCL. Clinicians say that because data in DLBCL are limited, they extrapolate from data in transformed disease to afÞrm activity in DLBCL. In the clinic, transformed disease is often used as a surrogate marker for DLBCL. The pivotal trial that won approval of this drug for use in indolent patients was a multicenter, nonrandomized study that evaluated 131 I-tositumomab in 60 patients with follicular, low-grade, or transformed NHLs who had a particularly poor prognosis (Kaminski MS, 2001). Patients had previously received at least two courses of chemotherapy and had become chemotherapy-resistant. A subgroup of 23 patients had transformed disease. Patients received a pretreatment dose of 450 mg unlabeled tositumomab and a dosimetry dose of 131 I-tositumomab (5 mCi [35 mg]). One to two weeks later, patients received 450 mg of unlabeled tositumomab followed by a calculated therapeutic dose of 131 I-tositumomab (35 mg tositumomab). In the subgroup of patients with transformed disease, 39% achieved a response and 13% achieved a complete response. The median duration of response in

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113

responding patients with transformed disease was long, more than 16 months; in three patients, the response lasted for more than 35 months. The normal course of disease for patients with transformed disease is rapidly progressive; median survival is only 6–12 months (Bastion Y, 1997). Radiolabeled 131 i-tositumomab’s most signiÞcant side effects are transient and reversible hematologic toxicities. Eighteen percent of patients developed grade 4 neutropenia, and 22% experienced grade 4 thrombocytopenia. The median duration of grade 4 toxicity was 15 days. Four patients were diagnosed with myelodysplasia at a median of 35.5 months after treatment and a median of 5.1 years after Þrst exposure to alkylating agents. Eight percent of patients become seropositive for human anti-mouse antibody (HAMA). Nonhematologic toxicities included fatigue, fever, and nausea; they were transient grade 1 or 2 toxicities. In a multicenter, expanded-access study involving 273 patients (median age 58), of whom 57 had transformed NHLs, patients received a single dose of singleagent tositumomab as a salvage therapy (Gockerman J, 2001; Schenkein DP, 2001). Ninety percent of all 273 patients had stage III or IV disease at entry, 44% had bulky disease, 43% had bone marrow involvement, and 60% had elevated lactate dehydrogenase (LDH). The median number of prior chemotherapies was two; 45% of patients had failed rituximab, and 21% had prior radiotherapy. Of the patients with transformed disease, 40% achieved a response, and 16% achieved a complete response. The most common toxicity in all patients was reversible myelosuppression; the median absolute neutrophil count nadir was 1,300 cells/mm3 , the platelet nadir was 68,000 cells/mm3 , and the hemoglobin nadir was 11.2 gm/dL. Fourteen percent of all patients developed grade 4 neutropenia, and 2% developed grade 4 thrombocytopenia. The most common nonhematologic adverse events were grades 1 or 2; they included asthenia (23%), nausea (17%), and fever (11%). Four percent of patients developed a serious infection that required hospitalization, and 8% developed HAMA. Seven patients with transformation to DLBCL received treatment as part of a trial of 131 I-tositumomab in patients experiencing Þrst or second recurrence (Davies AJ, 2003). Five patients proceeded to treatment with individualized therapeutic doses after dosimetry; the response rate in these patients was 80%. In all patients treated in the trial (including an additional 33 patients with indolent lymphomas), the median duration of response was 10.5 months; in complete responders, median duration was 15.2 months. Median duration of response in patients with transformed disease is not available. Toxicity was primarily hematologic: 16% and 13% of patients developed grade 3 or 4 thrombocytopenia or neutropenia, respectively. Median platelet and neutrophil nadirs occurred at six weeks; all patients recovered completely by nine weeks. Encouraging preliminary results are available for 131 I-tositumomab in the highdose setting. 131 I-tositumomab in combination with BEAM and ASCT has shown promise in patients with chemotherapy-resistant aggressive NHLs (Vose JM, 2003). Few other treatment options are available once patients become resistant to chemotherapy, and standard HDT and ASCT are not recommended in

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this setting (Shipp MA, 2002). Twenty-three patients underwent treatment with one of four doses of tositumomab (30–75 cGy), followed by BEAM, followed by ASCT. Fifty-nine percent of patients achieved a complete response and 9% of patients achieved a partial response, for an overall response rate of 68%. Threeyear, event-free survival was 39%; overall survival was 55%. Toxicities were similar to historical controls who received BEAM alone. No treatment-related deaths occurred. Two patients developed myelodysplastic syndrome more than two years post-transplantation. In a single-arm, dose-Þnding study, 52 patients with relapsed B-cell lymphomas underwent HDT with 131 I-tositumomab followed by chemotherapy and ASCT (Press OW, 2000). Patients received a myeloablative dose of tositumomab (1.7 mg/kg) labeled with an amount of I-131 calculated to deliver a target dose of radiation of 20–27 Gy to critical normal organs, followed by 60 mg/kg etoposide and 100 mg/kg cyclophosphamide, followed by ASCT. The majority of patients had follicular lymphomas; 14 had aggressive disease (6 patients with MCL, 6 with transformed DLBCL, and 2 with de novo DLBCL). Nineteen patients had high-intermediate-risk disease (based on the International Prognostic Index [IPI]), and 31 had low-intermediate-risk disease. Only two patients had low-risk disease. The maximum tolerated dose of 131 I-tositumomab that could be safely combined with the chemotherapeutic regimen delivered 25 Gy to critical normal organs. A retrospective comparison of this high-dose, multimodality treatment group with a control group of patients treated with conventional HDT and total body irradiation found that overall survival and progression-free survival at two years were superior in the multimodality treatment group: 83% versus 53%, and 68% versus 36%, respectively. Toxicities appeared similar in the trial and control groups, although more treatment-related deaths occurred in the control group (17% versus 7.6% in the 131 I-tositumomab group). Of the 131 I-tositumomab-treated patients, eight patients (15%) had grade 3 or 4 toxicities, including three patients with respiratory distress, two of whom died; three patients with reversible grade 3 mucositis or GI toxicity; one with reversible, grade 3 veno-occlusive disease; and four with a fatal infection (several patients experienced more than one toxicity). Grade 1 and 2 toxicities included mucositis (92%), nausea (63%), and infections (63%). Analysis by histology is not available, but both the indolent-disease and aggressivedisease groups had superior overall survival and progression-free survival with the 131 I-tositumomab regimen. At two-year follow-up, one patient had developed myelodysplastic syndrome, but no patients developed acute myeloid leukemia (AML) or other secondary malignant diseases. In a trial presented at ASCO in 2001, three patients with DLBCL received treatment with CHOP followed by 131 I-tositumomab within six to eight weeks of the last cycle of chemotherapy (Leonard J, 2001). All patients achieved a response, and investigators concluded that the treatment was safe, noting that dosimetry calculations will be important to minimize toxicity in a combined modality setting.

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In a Phase II dosimetry validation study involving 45 patients with indolent or transformed NHLs, patients received a 75 cGy total body dose of 131 Itositumomab (Vose JM, 2000). The overall response rate was 60% in patients with transformed disease; the complete response rate (including clinical complete responses) was 50%. The median duration of complete response in all patients was 19.9 months. Treatment was well tolerated; the principal toxicities were hematologic. Other toxicities were mild to moderate and included fatigue (32%), nausea (30%), fever (26%), vomiting (15%), infection (13%), and pruritis (13%). One patient developed HAMA. 131 I-tositumomab has been investigated as Þrst-line treatment of patients with MCL (Zelenetz A, 2003). Thirteen patients received 131 I-tositumomab for cytoreduction, followed by chemotherapy. Patients with measurable disease and less than 25% bone marrow involvement received 75 cGy tositumomab, followed 13–16 weeks later by six cycles of CHOP chemotherapy (1,000 mg/m2 cyclophosphamide). Twelve patients were evaluable for response to RIT and ten for response to RIT followed by chemotherapy. The overall response rate to induction with 131 I-tositumomab was 83%: 50% of patients achieved a complete response, and 33% achieved a partial response. Nine of ten patients completing all therapy achieved a complete response. At median follow-up of 11 months, three relapses had occurred, at 6, 9, and 12 months after completion of therapy. Five episodes of febrile neutropenia occurred during the CHOP chemotherapy (consistent with the incidence expected in an elderly population, according to investigators). Nonhematologic toxicities were mostly grades 1 and 2. One patient developed pericarditis a week after RIT; one developed an intussusception at week 13. Two patients developed HAMAs. Longer follow-up will establish whether this combined modality treatment is an improvement over existing therapies. For example, Hyper-CVAD-R (multiagent chemotherapy combined with rituximab) has achieved a complete response in 89% of MCL patients (Romaguera J, 2003). 131 I-tositumomab can be safely administered to elderly patients (Gregory SA, 2003). A subgroup analysis of more than 1,500 patients who received 131 Itositumomab in the context of a clinical trial or the expanded access program compared adverse events in three age categories: younger than 60 years, 60–70 years old, and older than 70 years. The study found that 131 I-tositumomab can be safely administered to patients older than 60 years, even to patients older than 70 years, with statistical equivalence in all end points. A trend was observed (p = .02) toward more serious infections in older patients: 3.8% (younger than 60), 8.4% (aged 60–70), and 5.7% (older than 70). In patients with indolent disease, 131 I-tositumomab has shown promising activity when used for a second time in previous responders (Kaminski MS, 2003). This strategy has not yet been tried in patients with aggressive disease. After receiving treatment, patients must remain in the hospital for several days owing to the possibility of exposing other people to radiation. The risk to others is small and lasts for up to a week, until the isotope has been excreted from

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the patient’s body. During this time, certain restrictions are necessary: patients must receive treatment in isolation from patients who are not receiving similar radiotherapy treatments; lead screens must be placed on either side of the patient’s bed to absorb any radiation emitted while receiving hospital care; and visiting privileges are restricted to adults, excluding pregnant women. Patients who leave the hospital after a few days must take precautions; while the isotope is still in their bodies, they are at risk of contaminating the public. Patients must maintain a distance of six feet from other people; they must not take long trips sitting near other people in a car or plane; and they must sleep alone in a separate room. Data from a study evaluating the amount of radiation received by family members exposed to patients treated with 131 I-tositumomab showed that doses absorbed by family members were below the 5-mSv (500 mrem) limit set by the U.S. Nuclear Regulatory Commission. However, in Japan, according to the Medical Service Law Enforcement Regulations, patients receiving gamma radiation must be isolated in a hospital environment with lead-lined facilities. European regulations vary among countries—for example, some countries require extended hospitalization of patients, and others require lead-lined hospital drainage. Yttrium-90-Labeled Epratuzumab. Yttrium-90-labeled epratuzumab (Immunomedics’ LymphoCide) is a humanized anti-CD22 antibody directed against the CD22 determinant RFB4, present on 75% of B lymphocytes. At the time of composing this reference, its Phase II trials in the United States and Europe have now been completed. This antibody can be given repeatedly with minimum risk of eliciting neutralizing antibodies, thus making fractionated administration feasible. Investigators hope that fractionated dosing may reduce heterogeneity in absorbed dose and widen the therapeutic window. In a Phase I/II trial, 23 patients with various types of B-cell lymphomas who had failed previous chemotherapy received repeated weekly dosing of yttrium90-labeled epratuzumab: 5 mCi/m2 or 2.5 mg/m2 infusion over one hour (Lind´en O, 2003). The patient group included Þve cases of transformed disease, six cases of DLBCL, and one case of MCL. Patients who had previously undergone HDT with ASCT received the lower dose. Patients were treated in cohorts and received either two, three, or four weekly infusions. Treatment could be repeated once if toxicity allowed, and disease was not progressing. The Þrst infusion included 4 mCi of indium-111 for scintigraphic veriÞcation of tumor targeting. With each infusion, 1.5 mg epratuzumab was administered. Of the 23 treated patients, response data are available for 22 patients: 16 treated at the higher dose and 6 at the lower dose. In the higher-dose group, 62% achieved a response; 25% achieved a complete response. Toxicity was primarily hematologic. In the lower-dose group, no toxicities of grade 3 or higher were seen. In the higher-dose group, grade 3 toxicity was observed in 0/3, 3/7, and 2/4 patients at the two-, three-, and four-times-weekly infusion levels, respectively. Grade 4 toxicity was observed only in two patients who had recently received high-dose

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chemotherapy and received four infusions of yttrium-90-labeled epratuzumab at the higher dose. SigniÞcant interpatient and intrapatient variability in tumor dose occurred. In patients treated at the higher dose, median absorbed dose to the tumor was 11.5 mGy/MBq (range 2.2–30.3 Gy); to the kidneys, 1.6 Gy; to the liver, 1.3 Gy; to the lungs, 2.1 Gy; and to the bone marrow, 0.4 Gy. In two patients whose absorbed dose could be estimated at re-treatment, the dose was doubled compared with the Þrst treatment. Unlabeled Monoclonal Antibodies Mechanism of Action. The introduction of the MAb rituximab has revolutionized treatment of NHLs. MAbs target cell-surface proteins and can interfere with receptor/ligand interactions, thus affecting downstream signaling and subsequent cell growth and proliferation. They can also alert the immune system to target a cell for death via cell antibody–dependent cellular cytotoxicity. Each of the identical cells that make up each lymphoma carries the same immunoglobulin on its surface. The unique portion of the immunoglobulin, the idiotype, is an ideal target for therapy. Early studies targeted the idiotype with MAbs designed for individual patient lymphomas, but MAbs have recently become more generic—targeting molecules found on many B-cell lymphoma cells. The ideal characteristics of an antigen target for a MAb include the following: • • •

Presence on all NHL cells, or at least on the clonogenic cells. Persistence of expression despite negative selection by the antibody. Sustained presence on the cell surface after antibody binding, which allows the exposed constant portion to activate host immune mechanisms—i.e., the antigen should not be shed, internalized, or modulated.

In NHLs, the Þrst MAb to reach the clinic—rituximab—targeted CD20. CD20 is present on 95% of all B-cell lymphomas; it is not present on stem cells, and its expression does not vary at different stages of cell division. Furthermore, CD20 does not respond to antibody binding by internalizing or shedding from the cell surface. The only other MAb in an advanced stage of development is epratuzumab (discussed next), which targets CD22. Galiximab (Biogen Idec’s IDEC-114), which targets CD80, is also in development for NHLs Epratuzumab. Epratuzumab (Immunomedics’ LymphoCide; Immu-103) is a humanized IgG1 , anti-CD22 MAb in development for combination with rituximab in the treatment of indolent and aggressive NHLs. In 2003, Amgen suspended Phase III monotherapy trials in indolent NHLs because of lack of activity as a monotherapy, and in November 2003, Amgen decided not to begin a second Phase III trial for epratuzumab. Immunomedics is also developing a radioactive form of epratuzumab (discussed previously) linked to yttrium-90.

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Epratuzumab is directed against the CD22 determinant RFB4, which is present on 75% of B lymphocytes. One study has shown that—at least in vitro and in mice—unconjugated epratuzumab’s primary mechanism of action takes place via antibody-dependent, cell-mediated cytotoxicity rather than complement-mediated cell lysis or direct growth inhibition (Gada P, 2002). Unconjugated epratuzumab is administered over a period of 30–60 minutes, a shorter infusion time than rituximab, which requires four to six hours’ infusion. Epratuzumab has been investigated in several NHL settings, including the following: single-agent salvage therapy; in combination with CHOP-R; in combination with rituximab; and as a second course following relapse in patients already treated with epratuzumab. Fifty-two patients received escalating doses, from 120 mg to 1,000 mg, by weekly intravenous infusion for a total of four treatments. Patients had received a median of Þve prior regimens. Twelve patients (23%) achieved stable disease; Þve (10%) achieved a response (three complete responses, two partial). The median duration of response was 38+ weeks, and median time to progression was 35+ weeks. Responders had lower tumor burden, fewer prior treatments, and normal lactate dehydrogenase compared with nonresponders. All responses were observed in DLBCL patients and at the 240–600 mg/m2 /week levels. Epratuzumab was well tolerated, with no dose-limiting toxicity. Another single-agent study evaluated re-treatment of NHL patients who had responded to epratuzumab (Leonard JP, 2003[a]). Two patients with DLBCL received a second four-week course of epratuzumab, with no intervening therapy. Both patients achieved a second response, with a time-to-disease progression of 27 weeks and 62 weeks, respectively. Re-treatment was well tolerated; toxicities occurred only during the Þrst infusion and were grade 1. If this level of toxicity is conÞrmed in larger studies, epratuzumab would represent an advantage over rituximab, with which grade 3 infusion-related toxicity has been observed in 12% of patients and grade 4 toxicity in 3% of patients who were given a standard dose once weekly for four weeks. Several studies have investigated the combination of epratuzumab with rituximab, alone or with CHOP. At the ASCO meeting in 2004, investigators presented preliminary data involving 15 patients treated with epratuzumab, rituximab, and CHOP in the Þrst-line setting (Micallef IN, 2004). These patients received epratuzumab (360 mg/m2 ) followed by rituximab (375 mg/m2 ) and standard-dose CHOP, all given on day 1, every 21 days, for six to eight cycles. The incidence of grade 4 neutropenia was 31%. Nine patients required dose reductions, eight of which were secondary to grade 4 neutropenia, despite the use of growth factor support. Fourteen patients were assessable for response: 50% achieved a complete response, and 36% a partial response. At a median follow-up of 8.1 months, 14 of 15 patients remained alive without disease progression. In this small trial, epratuzumab did not appear to confer any beneÞt over rituximab and CHOP. Twenty-one previously treated patients (median age 63) with indolent or aggressive NHLs were treated with four weekly doses of 360 mg/m2 epratuzumab

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in combination with 375 mg/m2 rituximab and monitored for toxicity and clinical response to therapy (Leonard JP, 2002[b]). All patients were rituximab-naive. The combination of these two agents was well tolerated, with grades 1 and 2 infusion-related toxicities, and was comparable to antibody monotherapy. Of the 19 evaluable patients, 53% achieved an objective response; of the responders, 80% achieved a complete response. After a follow-up of more than 13 months, nine out of ten patients had ongoing responses. A second Phase II trial investigated the combination of rituximab and epratuzumab in patients with relapsed/refractory B-cell NHLs (Strauss SJ, 2004). The study involved 65 patients, 13 of whom had DLBCL. Median patient age was 61 years; 35% had more than two prior treatments; and 77% of the patients were rituximab-naive. Patients received 360 mg/m2 of epratuzumab intravenous over 60 minutes, followed by infusion of 375 mg/m2 of rituximab weekly for four weeks. The most frequent toxicities were rigors (22%), fever (15%), hypotension (14%), and nausea (11%). One infusion reaction (hypotension) necessitated termination of therapy. Forty-six percent of DLBCL patients achieved a response. Half of responding patients had a complete response (including unconÞrmed complete responses), and half had a partial response. The disadvantage with epratuzumab is its limited application. Only 75% of lymphomas express the target antigen CD22, and in single-agent trials involving heavily pretreated patients, only 10% of patients achieved a response. By comparison, in the Þrst-line setting, CHOP-R achieves 75% complete response in DLBCL patients. Antisense Oligonucleotides Overview. No antisense oligonucleotides are on the market for cancer. This drug class has been plagued with disappointing trial results, such as those demonstrated in trials of Isis Pharmaceuticals/Eli Lilly’s ISIS-3521/AfÞnitak in non-small-cell lung cancer. Despite these compounds’ high speciÞcity and low toxicity, clinicians lack faith in their in vivo mechanism of action. Additionally, administration is difÞcult because these drugs require long periods of intravenous drip administration. Recent data showing positive results in melanoma trials may help reverse some of these setbacks. Mechanism of Action. Antisense oligonucleotides are short sequences of single-stranded DNA that bind to a speciÞc region of target messenger RNA (mRNA). This binding, or hybridization, triggers enzymatic degradation of the mRNA, thereby blocking the translation of the mRNA and the generation of the corresponding protein. Oblimersen. Note: At the time of composing this reference, the drug has been discontinued in the US for leukemia. Still in Phase III in Europe for CLL with chemotherapy.

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Oblimersen (Genta/Aventis’s Genasense [formerly G-3139]) is an antisense oligonucleotide designed to block the production of the bcl-2 protein from bcl2 , a proto-oncogene. Oblimersen is a fully phosphorothioated, 18-mer antisense oligonucleotide directed at the Þrst six exons of the bcl-2 open reading frame. It is being investigated broadly as a drug that can amplify the effectiveness of anticancer treatments. Phase II trials of oblimersen in combination with rituximab are ongoing in patients with MCL and recurrent NHLs. In April 2004, oblimersen received a negative recommendation for malignant melanoma from the FDA Oncologic Advisory Committee. The committee stated that the data presented did not provide substantial evidence for effectiveness, as measured by response rate and progression-free survival, to outweigh the increased toxicity endured by patients receiving this drug. Genta subsequently announced it was withdrawing the NDA for oblimersen therapy in malignant melanoma. The company plans to meet with the FDA to discuss key issues and the next steps in developing oblimersen for malignant melanoma. Bcl-2 is an apoptosis regulator that, when overexpressed, inhibits the process of natural cell death that should occur when cells are damaged—for example, by chemotherapy. Bcl-2 is located in the mitochondrial membrane and prevents the release of cytochrome c, thereby protecting the cell from entering the intrinsic apoptotic pathway and promoting cell survival. Inhibition of bcl-2 allows cells to progress through the cell death pathway. Bcl-2 is overexpressed in most cases of MCL and in approximately 25–50% cases of DLBCL (Hill ME, 1996; Kramer MH, 1996; Vose JM, 2002). It is expressed more frequently in B-cell lymphomas (83%) than in T-cell lymphomas (45%) (Lai R, 1998). Bcl-2 overexpression is associated with poor clinical outcome. Overexpression correlates with an increased risk of relapse after chemotherapy and reduced survival, disease-free survival, and relapse-free survival (Gascoyne RD, 1997). Upregulation of bcl-2 is associated with resistance to rituximab. At ASCO in 2004, investigators presented preliminary results from a Phase II trial of oblimersen and rituximab in patients with recurrent B-cell NHLs (indolent or aggressive) (Pro B, 2004). The trial is evaluating the effects of this combination on the level of Bcl-2 expression and correlating the results with clinical and molecular response. Oblimersen (3 mg/kg per day) is administered as continuous intravenous infusion for seven days in weeks 1, 3, and 5; rituximab (375 mg/m2 ) is given weekly for six doses. One of three MCL patients achieved a partial response. Common grade 3–4 toxicities included reversible neutropenia in 36% of patients, thrombocytopenia in 10%, and non-neutropenic fever in 21%. A Phase II trial tested the ability of oblimersen monotherapy to stabilize treatment-naive and pretreated MCL (Leonard JP, 2003[b]; Genta, press release, June 2, 2003). Patients were divided into two cohorts: 9 patients with chemotherapy-n¨aive disease and 16 patients with relapsed/refractory disease. Both cohorts received oblimersen 3 mg/kg per day for 7 days every 21 days for up to six cycles. Relapsed/refractory patients were allowed to receive six additional cycles of therapy. Chemotherapy-naive patients who progressed or

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failed to achieve a complete response went on to receive up to six cycles of oblimersen plus R-CHOP. Four of nine newly diagnosed patients (44%) and 6 of 16 previously treated patients (38%) remained stable without progression during all six treatment cycles when oblimersen was used alone. At time of reporting, seven newly diagnosed patients had crossed over to oblimersen plus R-CHOP. Of these patients, two achieved a complete response (29%) and two (29%) a partial response; other patients had not been in treatment long enough for evaluation. Of previously treated patients who crossed over to combination therapy, 38% achieved stable disease and 6% complete response. Higher doses of oblimersen will be tested in the relapsed/refractory cohort. No serious adverse events related to oblimersen occurred. Researchers have investigated the impact of oblimersen on the effectiveness of the antibodies rituximab and alemtuzumab.The rationale for combining the two classes of drug is the fact that oblimersen can reduce the level of bcl-2 protein in target tumor cells and sensitize these cells to other apoptotic stimuli. A preclinical study found that the addition of oblimersen to alemtuzumab greatly enhances the effectiveness of the latter, with similar cell kill at 20% of the standard dose (Cotter FE, 2003). Another preclinical trial found that downregulation of bcl-2 by oblimersen followed by rituximab resulted in longer survival in lymphoma-bearing, severe combined immunodeÞciency (SCID) mice in the case of a rituximab dose of 1 mg/kg but not in the case of a 2.5 mg/kg dose (Tucker C, 2003). Oblimersen is being tested in rituximab-resistant cells to determine whether bcl-2 downregulation will lessen acquired resistance (Ramanarayanan J, 2003). A Phase II trial examined the combination of rituximab (given weekly for six standard doses) and oblimersen (continuous infusion for seven days in weeks 1, 3, and 5) in patients with refractory/recurrent B-cell NHLs, with or without prior rituximab exposure (Pro B, 2003). One of two patients with stage IV mucosalassociated lymphatic tissue (MALT) lymphoma achieved a complete response, and one of two patients with MCL achieved a minor response. Grade 3 or 4 reversible neutropenia occurred in three patients (20%); grade 3 non-neutropenic fever occurred in four patients (26%). Other adverse events included anemia, hypotension, fatigue, and chills. Other preclinical trials have shown that downregulation of bcl-2 in NHL cell lines by oblimersen enhances the antitumor effects of the proteasome inhibitor bortezomib (Millennium Pharmaceuticals’ Velcade) without any excess toxicity, compared with single-agent treatment (O’Connor O, 2003). At 24 days, animals receiving either single agent had a 20–25% reduction in tumor volume versus controls, while animals receiving the combination experienced a 50% reduction. The principal limitations of oblimersen are its limited efÞcacy and the requirement for seven days continuous infusion. Because oblimersen’s toxicity proÞle is favorable, ambulatory administration may be possible in the future via a pump system. The favorable toxicity proÞle also facilitates combination with other agents.

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Proteasome Inhibitors Overview. The proteasome is a novel target for cancer drugs, and clinical studies in myeloma have generated intense interest in this Þeld. Because the proteasome interacts with many proteins (described further on), research into the downstream consequences of drugs that inhibit this target is ongoing. Mechanism of Action. Proteasomes are enzyme complexes involved in the disposal of damaged cellular proteins and the degradation of short-lived regulatory proteins that regulate cell proliferation, apoptosis, adhesion, angiogenesis, and signal transduction. Accordingly, inhibition of proteasomes can stimulate apoptosis and suppress tumor growth and spread. Regulatory proteins degraded by proteasomes include p53, p21, p27, NF-kB, I-kB {nuclear factor kappa beta (NF-κβ), I-κβ} and bcl-2. Downregulation of the NF-κβ survival pathway, activated by anthracyclines, can reverse drug resistance. In preclinical studies, cancer cells appear more susceptible to the effects of proteasome inhibition than normal cells. Bortezomib. Bortezomib (Millennium Pharmaceuticals’ Velcade; formerly PS341, MLN-341, and LDP-341 is a small-molecule proteasome inhibitor in Phase II trials for aggressive NHLs. The agent has been approved in the United States and Europe for the treatment of multiple myeloma and is under investigation for indolent NHLs. Bortezomib has high selectivity for the proteasome over other proteases (e.g., thrombin) and has demonstrated in vitro cytotoxicity against a wide range of tumor-cell lines. Bortezomib inhibits the 26S proteasome that works through multiple pathways, including pathways that inßuence apoptosis and angiogenesis. Because NF-κβ transcriptionally activates bcl-2, its inhibition may induce tumor-cell apoptosis or reduce bcl-2-associated drug resistance. Phase II trials are investigating bortezomib in several settings: as salvage therapy in patients with MCL or DLBCL; as single-agent therapy early in the course of disease history (0–2 prior regimens) in MCL patients; in combination with dose-adjusted EPOCH (etoposide, prednisone, vincristine, cyclophosphamide, and doxorubicin) chemotherapy for DLBCL patients; and in combination with pegylated (polyethylene glycol–conjugated) doxorubicin in patients with hematologic malignancies, including NHLs. A Phase II trial presented at the ASCO meeting in 2004 demonstrated bortezomib’s antitumor activity in the salvage setting (Goy A, 2004). At the time of presentation, data were available for 25 patients with MCL and 10 patients with DLBCL. The accrual goal is 152 patients. Patients’ median age was 63; the median number of prior therapies was 3.8. Patients received 1.5 mg/m2 intravenous push on days 1, 4, 8, and 11 every 21 days for up to six cycles (median 2.1). Restaging was done every two cycles. Two infectious episodes occurred, one fatal. In the MCL group, whose median number of prior therapies was three, four patients achieved a complete response, and seven achieved a partial response; the median duration of response was 5.7 months at time of reporting (range 1–15

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months). One of the DLBCL patients had a partial response. Five patients had grade 3 or 4 GI toxicities; four experienced hypotension, fatigue, or dehydration. One patient who had been treated previously with vincristine and taxanes developed grade 3 neuropathy. Four patients developed transient rash. Dose reduction was required in three patients. One patient had grade 4 neutropenia; six had grade 3 neutropenia. Ten patients developed thrombocytopenia; eight patients had a platelet nadir of less than 20,000/microL, and one had a nadir of less than 10,000/microL. A Phase II trial investigated single-agent bortezomib in 14 patients with measurable MCL who had received 0–2 prior regimens (Assouline S, 2003). Patients received bortezomib 1.3 mg /m2 twice weekly for two of every three weeks until progression or unacceptable toxicity. Patients achieving stable disease received four courses of treatment. Median patient age was 69 years; all patients had stage IV disease. Six patients had received no prior course of chemotherapy or only one; two other patients had received two prior regimens. Of 12 evaluable patients, 4 achieved a partial response (33%), and 5 others achieved stable disease (42%). Five patients experienced serious adverse events associated with ßuid retention; all had baseline dyspnea, ßuid retention, or both. Two patients died while receiving bortezomib; one death was related to grade 4 vascular leak syndrome, the second was caused by progressive MCL with severe edema. A second Phase II trial presented at ASCO in 2004 found that 6 of 11 previously treated patients with MCL achieved a response to treatment with bortezomib. Patients received bortezomib 1.5 mg/m2 twice weekly for two consecutive weeks for two of every three weeks. The only grade 3 or 4 toxicity was a grade 3 sensory and motor neuropathy in one patient. Responses lasted between 1 and 19 months. The patient who attained a partial response lasting 19 months was re-treated with four cycles of bortezomib and achieved a second partial response (O’Connor OA, 2004). A small sequential treatment trial investigated the efÞcacy and toxicity of bortezomib treatment followed by dose-adjusted EPOCH chemotherapy (Dunleavy KM, 2003). At the time data were presented at the American Society of Hematology (ASH) meeting in 2003, 13 patients had been enrolled. Median patient age was 43, the median number of prior regimens was two, and 92% of patients had stage 3 or 4 disease. In the Þrst part of the trial, patients received bortezomib 1.3 mg/m2 intravenous on days 1, 4, 8, and 11, every 21 days for six cycles. Patients who did not achieve a complete response crossed over to a second trial part; patients who had unstable disease at accrual proceeded directly to the second part. In the second part, patients received an escalating dose of bortezomib (0.5, 1, 1.5, then 1.7 mg/m2 ) on days 1 and 4 every 21 days and doseadjusted EPOCH with granulocyte colony-stimulating factor (G-CSF). (EPOCH comprised etoposide 50 mg/m2 /day on days 1–4, prednisone 60 mg/m2 twice daily orally on days 1–5, vincristine 0.4 mg/m2 /day on days 1–4, cyclophosphamide 750 mg/m2 on day 5, and doxorubicin 10 mg/m2 /day intravenous on days 1–4.) Etoposide, cyclophosphamide, and doxorubicin were dose-escalated 20% per cycle to achieve a nadir absolute neutrophil count of less than 500/mL.

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Of Þve patients who had been enrolled in the Þrst part of the trial, two achieved stable disease. Four cases of nonhematologic grade 2 toxicities occurred: one each of pain, diarrhea, vomiting, and taste disturbance. One grade 3 and one grade 4 thrombocytopenia occurred. The second part of the trial achieved one partial response and two cases of stable disease. Toxicities recorded for dose-adjusted EPOCH with bortezomib were similar to those seen in a study of Þxed-dose EPOCH (Gutierrez M, 2000): febrile neutropenia occurred in 21% and 18% of patients, respectively; grade 4 neutropenia in 68% and 48% of patients; and grade 3/4 thrombocytopenia in 24% and 27% of patients. As expected, a higher percentage of patients suffered gastrointestinal toxicity grade 2 in the dose-adjusted EPOCH-bortezomib trial (47% versus 14%), though grade 2 neurotoxicity was not increased (0% versus 22%). A small dose-Þnding trial combining bortezomib with pegylated liposomal doxorubicin achieved partial responses in two of six patients with NHLs; one patient had cutaneous T-cell lymphoma, and one had peripheral T-cell lymphoma (Orlowski RZ, 2003). The maximum tolerated dose was 1.5 mg/m2 bortezomib and 30 mg/m2 pegylated liposomal doxorubicin. Bortezomib’s drawbacks include its limited efÞcacy in DLBCL and its significant toxicities—most notably vascular leak syndrome. Cytotoxic Agents Overview. Cytotoxic anticancer drugs are collectively the largest and most established chemotherapy group. In this section, we discuss the most promising novel cytotoxics. These agents include modiÞcations of cytotoxics that are well established in the treatment of NHLs as well as members of new classes. Compounds that are marketed for other cancer investigations that have undergone limited investigations in aggressive NHLs include paclitaxel (Bristol-Myers Squibb’s Taxol), oxaliplatin (SanoÞ-Synth´elabo’s Eloxatin), gemcitabine (Eli Lilly/Spaly Bioquimica’s Gemzar), bendamustine (Salmedix’s Ribomustine) and liposomal doxorubicin (Alza/Schering-Plough’s Doxil/Caelyx). Paclitaxel achieved promising preliminary results in a small trial as a low-dose weekly treatment (Rizzieri D, 2004), and gemcitabine and oxaliplatin achieved promising preliminary results in combination with rituximab (El Gnaoui T, 2004). Mechanism of Action. The fundamental aim of all cytotoxic agents is to exert selective toxicity toward cancer cells. They do so via a variety of pathways. Sphingosomal Vincristine. Sphingosomal vincristine (Enzon, Inex Pharmaceuticals’ Onco-TCS, vincristine sulphate liposomal injection) is a therapeutic that contains the cytotoxic vincristine encapsulated in sphingomyelin liposomes. Inex describes its drug delivery system as a “transmembrane carrier system,” or TCS. Onco-TCS is preregistered in the United States. The FDA accepted the new drug application (NDA) in May 2004 and gave it a Standard Review

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designation. The agency’s target date for completion of review is January 15, 2005. The NDA is seeking marketing approval for Onco-TCS as a single-agent treatment for patients with relapsed aggressive NHLs previously treated with at least two combination chemotherapy regimens. The agent is also in Phase II development for Þrst-line treatment of aggressive NHLs and in clinical trials for small-cell lung cancer and several hematologic malignancies. Encapsulation of cytotoxics in liposomes is intended to extend the active agent’s circulation in the bloodstream and promote the accumulation of the drug in tumors. Free-form (unencapsulated) drugs circulate throughout the body, diluting their effectiveness and damaging healthy tissues. Therefore, optimal doses of vincristine are often not administered to elderly patients or to patients who require repeated treatment because of its neurotoxicity. Inex released the Þnal results of the Phase II/III trial that forms the basis of its NDA at the ASH meeting in 2003 (Winter JN, 2003). One hundred and nineteen patients with relapsed or refractory, diffuse, aggressive NHLs received 2 mg/m2 sphingosomal vincristine by one-hour intravenous infusion every two weeks for a median of 4.6 cycles. Twenty-Þve percent of the patients responded to treatment; 6% achieved a complete response. Another 26% of the patients achieved stable disease. Median time to progression was three months, four months for responders. In patients with sensitive disease (patients responding for more than six months to their previous therapy), the response rate was 48%; in patients with resistant disease, the response rate was 16%. The investigators estimated median survival to be 206 days. Despite prior vincristine exposure in 98% of patients, and despite the increased dose intensity compared with standard regimens, treatment was well tolerated. Grade 3 neuropathy occurred in 32% of patients and caused 13% to stop treatment. Grade 3 or 4 hematologic toxicities included neutropenia (21%), anemia (12%), and thrombocytopenia (9%). Patients involved in the study had been heavily pretreated (median of three prior treatment regimens) and had multiple poor prognosis characteristics (predominantly stage III/IV disease, predominantly IPI score of 2 or greater; 46% had refractory disease). Investigators have reported preliminary results of a Phase II trial of sphingosomal vincristine combined with rituximab in patients with multiple relapsed DLBCLs (Ashcroft AJ, 2003). Eighteen evaluable patients had received rituximab (375 mg/m2 ) weekly for four weeks, plus 2 mg/m2 sphingosomal vincristine every two weeks for up to 12 cycles (median 5 cycles). The overall response rate was 56% (23% complete response); median duration of response was 156 days, and median overall survival was 167 days. The regimen was well tolerated, with paresthesia in 77% of patients (grade 3 in Þve patients). Hematologic toxicity was not dose-limiting for any patient. Patients involved in the study were elderly (median age 70) and heavily pretreated (median four chemotherapeutic regimens), and 61% had stage IV disease. Investigators concluded that this regimen may be appropriate for patients who are not suitable for other regimens because of previous toxicities or comorbidities. A trial of unmodiÞed vincristine provides comparison: in a large randomized trial comparing CHOP with CHOP-R

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in treatment-naive patients with DLBCL, unmodiÞed vincristine caused neuropathy in 51–54% of patients and grade 3 or 4 neurotoxicity in 5–9% of patients (CoifÞer B, 2002). Pixantrone. Pixantrone (Novuspharma’s BBR-2778) is a new aza-anthracenedione in Phase II development for aggressive NHLs and in Phase III development for indolent NHLs. Pending discussions with the FDA, Novuspharma plans to initiate a Phase III trial in 2004 in patients who need third-line treatment of aggressive NHLs. In the aggressive disease setting, Novuspharma is developing pixantrone for combination use with other cytotoxic agents in younger patients and for single-agent use in elderly, heavily pretreated patients. Pixantrone’s cytotoxic mechanism of action has not been fully elucidated. The compound intercalates DNA and inhibits topoisomerase II, but DNA damage is not correlated with cytotoxicity. In hematologic malignancies, pixantrone exerts a more powerful antitumor effect than either doxorubicin or pixantrone’s parent drug, mitoxantrone; it is designed to be less myelodepressive and cardiotoxic than either agent. A Phase II trial plans to accrue 75 patients for treatment with pixantrone in combination with cytarabine, methylprednisolone, and cisplatin (the BSHAP regimen). The trial will investigate the efÞcacy of BSHAP as a salvage regimen in patients experiencing their Þrst relapse and will assess its role as an induction regimen before bone marrow transplant. Patients receive two cycles of BSHAP before assessment of their transplantation eligibility; patients who are unsuitable may receive up to six cycles of BSHAP. Patients who achieve a response and are considered appropriate for transplantation proceed to stem-cell mobilization with one or more cycles of BSHAP plus rituximab, followed by high-dose chemotherapy and reinfusion of stem cells. In a Phase I trial, pixantrone was administered in place of doxorubicin as part of the CHOP regimen (CPOP) to 22 patients with relapsed aggressive NHLs (Borchmann P, 2003). Patients had previously received their maximum permitted dose of standard anthracyclines (median 428.8 mg/m2 ). The mean age of patients was 66.1 years. The majority suffered from relapsed DLBCL (12 patients) and had advanced disease (eight patients stage IV, Þve patients stage III). The median number of prior chemotherapy regimens was two. Eleven patients had previously been treated with radiation. At the time of publication, six patients had achieved a complete response, seven a partial response, and three had progressive disease. Final evaluations presented at ASH and published in a Novuspharma press release showed that 13 patients (59%) achieved a complete response and another 4 patients achieved a partial response, for a major response rate of 77%. At the highest dose of pixantrone, all seven patients responded, with six patients (85%) achieving a complete response. Dose-limiting neutropenia occurred in three of six patients receiving the 150 mg/m2 dose, but only one case of febrile neutropenia occurred. Cardiac toxicity was observed in a single patient who had a reduced left ventricular ejection fraction (LVEF; 44%) at study entry; this patient’s cardiac

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function fell to 27% after six courses of treatment. The trial established 120 mg/m2 as the most appropriate dose for further trials. A Phase II, single-agent, salvage treatment trial involving 31 elderly patients with DLBCL (24 patients) or MCL (7 patients) investigated the efÞcacy and toxicity of a lower dose of pixantrone. Patients received 85 mg/m2 three times weekly (Borchmann P, 2003). Patients’ median age was 66 years, and they had been pretreated with a median of two prior regimens. Responses included Þve complete and four partial responses, with a median duration of more than 17 months. The principal toxicity was neutropenia. Another Phase I/II trial investigated pixantrone in place of etoposide in the ESHAP regimen in 21 patients with relapsed or refractory disease after a median of one prior treatment (Levine AM, 2003). Patients received pixantrone, 80 mg/m2 intravenous, over one hour on day 1; methylprednisolone, 500 mg intravenous, on days 1–5; cisplatin, 25 mg/m2 intravenous, on days 1–4; and cytosine arabinoside, 2,000 mg/m2 intravenous, on day 5. Treatment was given on an outpatient basis when possible. Chemotherapy cycles were repeated every 21 days. Patients had a median age of 50 years, and most had advanced disease (85% stage III or IV). Most patients had DLBCL or variants (86%), two had follicular large-cell lymphomas, and one had transformed lymphoma. All patients had received a prior anthracycline-containing regimen. Seven patients had relapsed disease; the remainder were refractory to their prior regimens. Of 18 patients evaluable for response, 7 patients achieved a complete response (39%), and 4 achieved a partial response (22%). Of the responding patients, four had primary refractory disease. The median duration of complete or partial response was 195+ days (range 24+ to 290+). The most common grade 4 toxicities were hematologic, including neutropenia in 13 patients (62%), 5 of whom developed febrile neutropenia (26%); thrombocytopenia in 8 patients (38%); and anemia in 4 patients (19%). No clinically signiÞcant cardiac events or reductions in LVEF of 20% or more were noted. Nonhematologic side effects, primarily grade 1 or 2 in severity, included nausea (79%), blue urine (chromaturia; 63%), and fatigue (58%). Pixantrone’s main limitation is its hematologic toxicity. We note that the relatively high rate of grade 4 neutropenia described in an earlier paragraph (62%, with 24% febrile neutropenia) occurred in patients who had received a median of one prior regimen. Pixantrone’s contribution to the toxicity associated with the CPOP regimen is difÞcult to disentangle from that of the other agents. Histone Deacetylase Inhibitors Overview. A wide range of histone deacetylase (HDAC) inhibitors are in development for NHLs. In this section, we discuss depsipeptide, the most promising HDAC inhibitor. Other HDAC inhibitors in early stages of development include SAHA (Memorial Sloan-Kettering Cancer Center/Aton Pharma), pyroxamide (National Cancer Institute/Southern Research Institute), the benzamides MS-275

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(Schering AG) and CI-994 (PÞzer), and the butyrate derivative 4-PBA (Johns Hopkins University). Mechanism of Action. Depsipeptide’s mechanism of action is not completely understood, but in vitro studies have shown that the compound reduces mRNA expression of the c-myc oncogene and causes p53-independent cell-cycle arrest at G0/G1 by inhibiting signal transduction through MAP kinase. The agent activates SV40 promoter transcription and inhibits HDAC. In vitro studies have shown that alone and in combination with hypomethylating agents, depsipeptide induces a number of cellular proteins involved in susceptibility to immunologic manipulation, apoptosis, and proliferation. It may also have antiangiogenic activity. Depsipeptide is a substrate for P-glycoprotein but is not cross-resistant with several cytotoxic drugs. Depsipeptide. Depsipeptide (Fujisawa/National Cancer Institute’s FR-901228, formerly FK-228 and NSC 630176) is a bicyclic peptide in Phase II development in the United States for cutaneous or peripheral T-cell lymphoma. Phase II data demonstrate single-agent depsipeptide activity in T-cell lymphoma (Peikarz R, 2003[a]). Of 20 patients with T-cell lymphoma, six achieved partial responses and two achieved complete responses. Patients received 14 mg/m2 on days 1, 8, and 15 of a 28-day cycle. Ten patients had been previously treated with doxorubicin. Some cardiac abnormalities were observed in more than half of the electrocardiograms taken post-treatment (Grade 1 T-wave ßattening or grade 2 segment depression), but investigators found no signiÞcant change in cardiac function or evidence of myocardial damage. Electrolyte replacement was incorporated into patient regimens to mitigate the potential side effects of QTc interval prolongation. In December 2003, more data from the same trial were presented at the 45th annual ASH meeting. Thirty-seven patients had been enrolled in four arms based on histology and prior therapy. Response data were presented only for patients in arm 1 (cutaneous T-cell lymphoma): three patients achieved a complete response and Þve patients a partial response, for an objective response rate of 57%. Five patients who had received depsipeptide for 12–30 months showed no evidence of cumulative toxicity (Piekarz RL, 2003[b]). In May 2002, at the 38th annual ASCO meeting, investigators presented Phase I/II data from 13 patients (Piekarz RL, 2002). Depsipeptide was administered at 14 mg/m2 in a four-hour infusion on days 1, 8, and 15 of a 28-day cycle. One patient with peripheral T-cell lymphoma exhibited a complete response, one showed a partial response, and two showed disease progression. In evaluable patients with cutaneous T-cell lymphoma, nine partially responded to treatment. The major toxicities were granulocytopenia, thrombocytopenia, hypocalcemia, fatigue, nausea, and vomiting. Depsipeptide has demonstrated antitumor activity in clinical trials, the sine qua non for successful development.

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Immunotoxins Mechanism of Action. Immunotoxins use MAb technology conjugated to natural toxins. Immunotoxins consist of peptides, usually an antibody or growth factor, that are linked to toxins such as diphtheria toxin, Pseudomonas exotoxin, or ricin. This “magic bullet” mechanism is designed to target the toxic moiety to a speciÞc cell by binding the growth factor/antibody portion of the immunotoxin to the cell-surface receptor. When internalized, the toxin is cleaved into an active form and causes cell death. Denileukin Diftitox. Denileukin diftitox (Seragen’s Ontak) is in Phase II development in the United States for the treatment of B-cell and T-cell NHLs. This agent has been launched in the United States and preregistered in Western Europe for the treatment of patients with cutaneous T-cell lymphoma whose malignant cells express the p55 (CD25) component of the interleukin (IL)-2 receptor. It is also in development for chronic lymphocytic leukemia and psoriasis. Denileukin diftitox is an IL-2 diphtheria toxin fusion protein designed to direct the cytocidal action of diphtheria toxin to cells that express the IL-2 receptor. The human IL-2 receptor exists in three forms: low (CD25), intermediate (CD122/CD132), and high (CD25/CD122/CD132) afÞnity. Denileukin diftitox targets NHLs expressing intermediate- and high-afÞnity IL-2 receptors. A Phase II trial investigated the activity of denileukin diftitox in heavily pretreated patients with relapsed/refractory, low-grade or intermediate-grade, B-cell or T-cell NHLs of any histology, barring cutaneous T-cell lymphoma (Dang NH, 2003). Thirty-nine evaluable patients received 18 ug/kg/day by daily intravenous infusion for 5 days, every 21 days, for up to eight cycles (mean 3.2 cycles). Patients were premedicated with a corticosteroid. CD25 expression was measured by immunohistochemistry or ßow cytometry. Patients’ median age was 64 years, and most had undergone multiple prior treatments, including autologous stem-cell transplantation. Four patients (10%) achieved a complete response, including one patient with DLBCL, one with MCL, and one with peripheral Tcell lymphoma. Seven patients (18%) achieved a partial response, and 10 (26%) achieved stable disease. Patients with CD25-positive and patients with CD25negative NHLs responded to treatment. Of the patients with B-cell disease, 24% achieved a response, and another 21% achieved stable disease; for patients with T-cell disease, the Þgures were 40% and 40%, respectively. The mean number of treatment cycles for patients with responsive or stable disease was 4.6. Treatment was well tolerated. Most toxicities were mild and transient and included elevation of transaminases, fatigue, and hypoalbuminemia. No hematologic toxicities occurred. Denileukin diftitox has an inconvenient administration protocol (infusion over Þve days) and signiÞcant toxicity. Data published to date do indeed show relatively low response rates, although it should be noted that the patients who received denileukin diftitox had undergone multiple prior treatments. Also, the agent has proven activity in a very difÞcult-to-treat setting, and signiÞcant unmet need exists for patients with T-cell lymphomas.

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Vaccines Overview. Therapeutic vaccines are designed to induce the patient’s own immune system to make antibodies to combat disease. Vaccine development is most advanced for indolent lymphomas. Two features of indolent NHLs have spurred investigators to invest more than two decades in developing a vaccine for this group of diseases: (1) the surface of each lymphoma cell in a patient with B-cell lymphoma carries a unique identifying protein (idiotype), and (2) some patients have long periods of stable disease, possibly because the immune system is holding the disease in check, and a few patients achieve spontaneous remissions. Aggressive lymphomas are less attractive targets for vaccines because only the Þrst of these two features holds true for these diseases. Because aggressive lymphomas progress rapidly, little time is available to stimulate an immune response. Active immunotherapy may generate a lasting response. Investigators hope that recognizing the idiotype protein will become part of the “memory” of the immune system so that the immune system will continue to Þght the protein for several years. Because each patient’s lymphoma is unique, each idiotype vaccine is made from an individual patient’s tumor cells, an expensive and laborious process. In ongoing Phase III trials in indolent lymphomas, vaccine therapies are being administered after chemotherapy. This schedule is designed to reduce the burden of disease so that the immune reaction has fewer malignant cells to Þght. Ongoing Phase II trials are investigating vaccines combined with passive immunotherapy (administration of antibodies). In the early days of vaccine development for indolent NHLs, investigators used short courses of unconjugated idiotypic protein. However, trying to get the host to “break tolerance” and react against the idiotypic protein has been problematic. Current Phase III trials administer the vaccine over a longer period (three to six months), link the idiotypic protein to keyhole limpet hemocyanin (KLH) to heighten its immunogenicity, and coadminister granulocyte-macrophage colony-stimulating factor (GM-CSF). The role GM-CSF plays is to increase the likelihood of an immune reaction by recruiting dendritic cells to the injection site to “cross-talk” between the vaccine and the immune cells. Vaccines in development are well tolerated; soreness at the injection site is the most common reaction. Some patients develop ßulike symptoms, including muscle aches and mild fever. Although two companies have vaccines in Phase III trials for indolent lymphomas (Genitope and the National Cancer Institute [NCI]), we discuss only Genitope’s vaccine because it is the only one in trials for aggressive NHLs as well—notably, MCL. The NCI vaccine’s method of manufacture differs from that of Genitope’s vaccine; it is made by means of rescue fusion rather than gene cloning. Large-Scale Biology Corporation and Cellgenix also have lymphoma vaccines in early stages of clinical development. Mechanism of Action. Vaccines in development for NHLs are designed to stimulate the immune system to launch a response against the tumor-speciÞc,

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variable regions of the clonal immunoglobulin (idiotype [Id]) expressed by the malignant B-cell population. Recombinant Idiotype-KLH Vaccine. Recombinant idiotype-KLH vaccine (Genitope’s MyVax, formerly GTOP-99) is a recombinant Id protein conjugated to KLH. The vaccine is in Phase III development for indolent NHLs and in Phase II trials for aggressive NHLs, including MCL. To be eligible for vaccine treatment, patients with aggressive NHLs must be in complete or partial response following CHOP chemotherapy and have an adverse prognosis (IPI 2–4). The vaccine is coadministered with GM-CSF. Genitope manufactures the vaccine by means of gene cloning. The company hopes the vaccine will improve time to disease progression following chemotherapy. In a Phase II study, Þve patients with MCL were treated with recombinant idiotype-KLH vaccine according to an initial schedule of Þve immunizations starting six months after chemotherapy (Leonard JP, 2003[c]). Because three of these patients were unable to complete treatment because of rapidly progressing disease, investigators changed the treatment schedule: vaccination began only 13 weeks after completion of chemotherapy and continued every two weeks for seven doses; an eighth dose was administered at week 18 of vaccination. At a median follow-up of 522 days after the completion of chemotherapy, median time to progression had not been reached. Investigators did not provide details of adverse events but described them as mainly grade 1 and 2, transient and manageable. Vaccines are less well suited to most of the aggressive lymphomas. Owing to the rapid course associated with aggressive disease, the window of opportunity in which to stimulate an immune response is narrower. In the context of aggressive lymphomas, the most notable limitations of idiotype vaccines are their cumbersome, time-consuming manufacturing process and the duration required to stimulate an immune response. Furthermore, the idiotype is a poor immunogen, and hence requires the attachment of KLH and coadministration of GM-CSF. Even so, such vaccines have the potential to be approved as Þrst-line treatment of MCL, in which existing therapies achieve very low survival rates, and vaccines offer a greatly improved toxicity proÞle.

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Bladder Cancer

ETIOLOGY AND PATHOPHYSIOLOGY Background Bladder cancer (BC), the seventh most common malignancy in the world, is primarily a disease of men older than 65 and is rarely diagnosed before the age of 40 (Ferlay J, 2001; Herr HW, 2001). The link between BC and smoking, as well as occupational exposure to certain carcinogens, is well established. Cigarette smoking is the most important cause of BC, accounting for up to 50% of BC in men and 33% in women (IARC, 1987). The development of BC is associated with the excretion of carcinogenic metabolites in the urine (Van der Meijden APM, 1998). The bladder urothelium may have direct contact with these carcinogens, which damage the urothelial cells that line the inside of the bladder and increase the chances of developing cancer (Silverman DT, 1992). High consumption of ßuids, however, may reduce the exposure by diluting the urine and reduce the contact time through increased frequency of urination (Braver DJ, 1987). The ultimate recurrence and progression of tumors may require multiple insults to the uroepithelium. In addition, further molecular alterations, such as genetic mutations, may be necessary for tumor progression (Lee R, 2000). Predicting future tumor behavior is a major task when treating BC (Zlotta AR, 2000); researchers are just beginning to understand the genetic events that occur in BC. Determining the prognostic signiÞcance of each genetic alteration and designing new therapies that directly target these alterations is a signiÞcant Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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challenge for researchers who are working to prevent BC and improve treatment outcomes. Pathophysiology Anatomy of the Bladder. The normal human urinary bladder (Figure 1) is a hollow, balloon-shaped organ that has a muscular wall that allows it to expand and contract. It is located in the lower abdomen and collects urine from the kidneys via thin tubes called ureters; the urine is stored until it is excreted through another tube called the urethra. The urinary bladder has three layers: the epithelium and the subepithelial connective tissue, the muscularis, and the perivesical fat (Fleming ID, 1997). A layer of urothelial cells (transitional cells) lines the inside of the kidney, ureter, bladder, and urethra. This layer is known as the urothelium or transitional epithelium. Under the urothelium is a thin region of connective tissue called the lamina propria. The next deeper layer is a wider region of muscle tissue called the muscularis propria. Beyond this muscle tissue, another region of fatty connective tissue separates the bladder from other, nearby organs (American Cancer Society [ACS], 2004). Figure 2 shows the extent of primary BC with tumor staging according to the tumor, node, metastasis (TNM) system. Pathogenesis and Natural History. BC is a disease in which cancer cells form in the tissues of the bladder. It appears that alteration in cell-cycle regulation is a key event in determining the biological behavior of BC (Cote RJ, 2003). Normal cellular proliferation takes place in an orderly fashion through the cell cycle. Cell-cycle-associated protein complexes composed of cyclins and cyclindependent kinases (Cdks) regulate normal cellular proliferation. Several tumor suppressor genes (TSGs), together with their protein products (p53, pRb, p16, and p14), act at the G0/G1 checkpoint to prevent the loss of cell-cycle control and

FIGURE 1. Structure of the normal human urinary system.

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FIGURE 2. Extent of primary bladder cancer with tumor staging according to the TNM system.

tumor progression (Quek ML, 2003; Williams SG, 2001). Mutations in a single TSG are unlikely to be the only factor resulting in carcinogenesis; alterations of one of the several pathways within the cell cycle are likely to be responsible for the progression of BC (Williams SG, 2001). The natural history of BC correlates with several prognostic factors, including histological grade, the depth of penetration into the bladder wall (stage), the appearance of vascular/lymphatic invasion, and the presence of carcinoma in situ (CIS) (Lee R, 2000). The biological foundation underlying the different types of BC within each of these categories is not fully understood. Some tumors, for example, appear to have favorable histology, but they progress and express aggressive activity despite theoretically curative treatment, while other tumors appear to have unfavorable histology but are less aggressive (Lee R, 2000). Two risks have been attached to all superÞcial tumors: a risk of recurrence with no worsening of the stage or grade, and a risk of progression leading to muscle-invasive and metastatic disease (Chopin DK, 2002). Low-grade, lowstage tumors pose less risk of progression than do high-grade, high-stage tumors (Chopin DK, 2002). CIS is by deÞnition a high-grade tumor; it is found in more

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than half of bladders with multiple papillary tumors (Herr HW, 2001). Primary CIS constitutes 1–2% of newly detected cases of BC (Herr HW, 2001). CIS is believed to be a predominant precursor of invasive tumors (Herr HW, 2001). A previous study described three types of CIS. Type 1 CIS is an early stage of BC; tumors are not very aggressive and are unifocal. Type 2 CIS causes bladder irritative symptoms, and tumors are multifocal. Type 3 CIS increases the risk of recurrence, progression, and cancer-related death, and tumors are associated with one or more other tumors (Lamm DL, 1992). Multiple pathways of human bladder carcinogenesis have been proposed. We brießy describe one of these pathways below. When BC arises, the initial step is intraepithelial neoplastic transformation (Lee R, 2000). The transformed urothelial cells acquire growth advantages that allow them to expand and achieve dominance over normal urothelial cells. Assuming that tumorigenesis is a multiple-step process, papillary forms of BC arise via the ampliÞcation of cyclin D1, upregulation of vascular endothelial growth factor (VEGF), and loss of heterozygosity of chromosome 9. In contrast, p53 or Rb inactivation and platelet-derived endothelial cell growth factor (PDECGF) expression are implicated in the development of invasive forms of urothelial carcinoma, also known as transitional-cell carcinoma, or TCC (Lee R, 2000). Metastatic BC is a fatal disease; the most common sites of metastases are regional lymph nodes, lung, liver, bone, and skin. Less common sites are the brain, meninges, and the organs within the peritoneal cavity (Al-Sukhun S, 2003). The distribution of metastasis is particularly important when considering treatment because the sites of involvement appear to correlate with prognosis (Al-Sukhun S, 2003). Classification. The three major types of cancers that affect the bladder are urothelial carcinoma (TCC), squamous-cell carcinoma, and adenocarcinoma (Cote RJ, 2003). More than 90% of BCs are TCCs derived from the uroepithelium (Herr HW, 2001). About 3% of BCs are pure squamous-cell carcinomas, and 2% are adenocarcinomas (Herr HW, 2001). The remaining 5% are a mix of transitionalcell and squamous-cell carcinomas or adenocarcinomas. Nearly all squamous-cell carcinomas and adenocarcinomas of the bladder are invasive. Signs and Symptoms. Hematuria, the appearance of blood in the urine, is the key symptom of BC (Van der Meijden APM, 1998). Hematuria is either visible or microscopic. Hematuria occurs in 80–90% of cases, but urinary frequency, bladder irritability, and reduction in overall bladder capacity are also common (Herr HW, 2001). Rarely, metastatic bone lesion pain or local progression of disease is the presenting sign (Herr HW, 2001). Screening asymptomatic patients for hematuria does not seem to have any effect on overall survival; it does, however, increase the probability of diagnosing the disease at an earlier stage (Herr HW, 2001). Prospective studies are ongoing to assess the role of screening in high-risk populations.

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Staging. Physicians in the seven major markets generally stage BC according to the internationally recognized TNM (primary tumor/regional lymph nodes/ distant metastasis) classiÞcation system developed by the American Joint Committee on Cancer (AJCC) (see Table 1). BC staging is determined by the depth to which the tumor invades the bladder wall. For speciÞc deÞnitions of the BC TNM staging system and one- to Þve-year survival rates, see Table 2. SuperÞcial BC can be classiÞed as low-risk, intermediate-risk, or high-risk according to the likelihood of progression. Table 3 lists the risk of progression and cancer-related death according to tumor stage and grade. Clinically, BCs are classiÞed as superÞcial, muscle-invasive, or metastatic disease (Herr HW, 2001). SuperÞcial tumors conÞned to the bladder mucosa, submucosa, or lamina propria account for 70–80% of newly diagnosed BCs. About 20% of BCs invade the muscle layer (invasive BC) and up to 5% have de novo metastases (metastatic BC) (Cote, RJ 2003; Al-Sukhun S, 2003; Herr HW, 2001). SuperÞcial BCs consist of papillary tumors that can be low-grade or highgrade (Ta, grade 1–3) and CIS (Crawford ED, 2003). In patients who have CIS TABLE 1. AJCC TNM Classification System for Bladder Cancer Primary Tumor (T) TX T0 Ta Tis T1 T2 pT2a pT2b T3 pT3a pT3b T4 T4a T4b

Criteria Primary tumor cannot be assessed. No evidence of primary tumor Noninvasive papillary carcinoma Carcinoma in situ: ‘‘flat tumor’’ Tumor invades subepithelial connective tissue Tumor invades muscle Tumor invades superficial muscle (inner half) Tumor invades deep muscle (outer half) Tumor invades perivesical tissue Microscopically Macroscopically (extravesical mass) Tumor invades any of the following: prostate, uterus, vagina, pelvic wall, or abdominal wall Tumor invades the prostate, uterus, vagina Tumor invades the pelvic wall or abdominal wall

Regional node status (N) NX Regional lymph nodes cannot be assessed N0 No regional lymph node metastasis N1 Metastasis in a single lymph node, 2 cm or less in greatest dimension N2 Metastasis in a single lymph node, more than 2 cm but not more than 5 cm in greatest dimension; or multiple lymph nodes, none more than 5 cm in greatest dimension N3 Metastasis in a lymph node, more than 5 cm in greatest dimension Distant metastasis (M) MX Distant metastasis cannot be assessed M0 No distant metastasis M1 Distant metastasis AJCC = American Joint Committee on Cancer. TNM = Tumor, node, metastasis.

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TABLE 2. Definitions for the TNM Staging Classification of Bladder Cancer

AJCC Stage

Bladder Cancer Survival by One-Year Two-Year Three-Year Four-Year Five-Year Stage (%) (%) (%) (%) (%)

Definition

0a

Ta, N0, M0

0is

Tis, N0, M0

0

94.00

88.09

82.25

76.63

I II

70.35

T1, N0, M0

I

91.33

82.50

75.24

68.59

62.44

T2a, N0, M0 T2b, N0, M0

II

75.07

58.03

49.01

42.48

37.88

III

T3a, N0, M0 T3b, N0, M0 T4a, N0, M0

III

64.84

45.02

35.74

30.44

26.04

IV

T4b, N0, M0 Any T, N1, M0 Any T, N2, M0 Any T, N3, M0 Any T, any N, M1

IV

41.65

23.37

16.85

13.35

11.46

Overall

84.96

74.82

67.85

61.96

56.39

AJCC = American Joint Committee on Cancer. TNM = Primary tumor (T), lymph node spread (N), distant metastasis (M). Note: Survival data from National Cancer Data Base (NCDB). Data accessed August 15, 2004.

TABLE 3. Risk of Progression and Cancer-Related Death According to Tumor Stage and Grade

Risk Group Low

Intermediate

High

Types of Tumor Single pTa G1 pTa G1–G2 (not recurring in less than three months) Multifocal pTa G2 pTa with multiple recurrences pTa G3, single pT1 G2 pT1 G3 Diffuse pTis Multifocal pT1 pT1 recurring in less than six months

Risk of Disease Progression at Five Years (%)

Risk of Cancer-Related Mortality at Ten Years (%)

7.1

4.3

17.4

12.8

41.6

36.1

Adapted from Rischmann P, 1998. Full source citations appear in ‘‘References.’’

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associated with papillary tumors, the potential for developing muscle-invasive disease is 42–83%, compared with 20–34% in patients who have isolated CIS (Hudson MA, 1995). Overall, the risk of bladder muscle invasion for noninvasive disease (pTa, 9%) is half that for invasive disease (pT1, 18%) (Kurth K, 1992). CIS (pTis) makes up less than 5% of all bladder tumors (Chopin DK, 2002). Left untreated, CIS will progress to invasive disease in Þve years in more than 50% of patients (Herr HW, 2001). The risk of CIS progression increases with widespread disease. Grading. Bladder tumors are classiÞed as low-grade (G1) or high-grade (G2, G3) according to the World Health Organization (WHO) classiÞcation (Epstein JI, 1998). Grading is of major importance for noninvasive tumors (superÞcial tumors) because almost all invasive neoplasms (T1 or greater) are high-grade (Herr HW, 2001). Grade 1 tumors are much less likely (6%) to progress than grade 3 tumors (30%) (Kurth K, 1992). Researchers have found that over 20 years of follow-up, the risk of progression increases to 14% for pTaG1 tumors and 45% for pT1G3 tumors; the risk of recurrence, however, is the same (approximately 50%) regardless of the aggressive nature of the tumor (Herr HW, 1997). Also, tumors of similar grade may behave very differently; additional markers are therefore crucial for identifying patients at risk for disease progression or recurrence (Zlotta AR, 2000). For invasive (muscle-invasive/metastatic) tumors, however, stage is the most important independent prognostic variable for progression and overall survival (Herr HW, 2001). The healthy epithelium has normal nuclear polarity, no pleomorphism, and a thickness of less than Þve to seven layers (Herr HW, 2001). Low-grade papillary carcinomas are relatively benign tumors that closely resemble the normal urothelium: normal nuclear polarity in more than 95% of the tumor; no, or only slight, pleomorphism; and more than seven layers of urothelium. In addition, low-grade papillary carcinomas rarely progress to a higher stage. High-grade papillary tumors show loss of polarization of the nuclei or moderate or prominent pleomorphism (Herr HW, 2001); progression to higher-stage lesions is therefore frequent. Prognostic Factors. For BC, the Þve-year survival rates range from approximately 11% for patients with advanced disease to approximately 70% for patients diagnosed with localized disease (National Cancer Data Base, 2004). The biggest challenge for researchers is to identify the group of patients whose disease has the potential to evolve into muscle-invasive disease and, subsequently, metastatic disease. Studies have found improved median survival in patients with metastasis limited to the lymph nodes, skin, and possibly the lungs; substantially worse prognoses have been documented in patients with liver, bone, and visceral metastasis (Geller NL, 1991; Loehrer PJ, 1992). The pattern of growth is important; superÞcial tumors are associated with better prognosis. Additionally, histology (TCC versus squamous-cell carcinoma and adenocarcinoma) is an important factor in treatment outcome. Evidence shows that patients with metastatic adenocarcinoma or squamous-cell carcinoma do not

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generally respond to chemotherapy and have a worse median survival compared with TCC patients (Saxman SB, 1997). Etiology Risk Factors. Most known cancer risk factors can be categorized as processes that increase a person’s exposure to mutagens; interfere with or prevent normal DNA repair; and/or increase the proliferation rate of cells (e.g., as a result of repetitive injury and tissue repair). Cigarette smoking is the principal known risk factor for BC, though occupational exposure to arylamine N -acetyltransferases (NATs), polycyclic aromatic hydrocarbons (PAHs), and aromatic amines is also associated with its development (Hung RJ, 2004; Yu MC, 1998). Workers who have an increased risk of developing BC include painters, truck drivers, and drill press operators (Silverman DT, 1989). Other industries that pose an increased risk to workers include the dye, rubber, and leather industries. Cigarette Smoking. Cigarette smoking is by far the most important risk factor for BC: 25–65% of all cases are attributable to this habit (Lee R, 2000). Smokers are two to four times more likely to develop BC than the general population (Lee R, 2000). Researchers have found that the duration and intensity of cigarette smoking independently increases the risk of developing BC (Castelao JE, 2001). The latency period from initial exposure to the development of the urothelial tumor is a median of 18 years (Herr HW, 2001). However, study results of the effect of cigarette smoking on BC have been inconsistent. One study found a positive correlation between smoking history and histological grade, stage, number, and size of bladder tumors (Thompson IM, 1987). In another study, tenyear survival data showed that 40% of smokers died of their disease compared with 27% of nonsmokers, even though no correlation between smoking history and histological grade or stage was observed (Raitamen MP, 1995). A subsequent study found no difference in BC risk between smokers of Þltered or low-tar cigarettes and smokers of nonÞltered or high-tar cigarettes, implying that lowtar cigarettes are no less mutagenic to human bladder urothelium than high-tar cigarettes (Castelao JE, 2001). The risk of TCC is related to the type of tobacco smoked; air-cured “black” tobacco is associated with higher risk because of its greater concentration of carcinogenic byproducts than is found in ßue-cured “blond” tobacco (Lee R, 2000; Silverman DT, 1996). A marker of arylamine exposure found in cigarette smoke (3- and 4-aminobiphenyl [ABP]-hemoglobin adduct) may play a role in smoking-induced bladder carcinogenesis (Yu MC, 1998). The 4-aminobiphenyl (ABP)-hemoglobin adducts, and the adducts of several other aromatic amines, are higher for smokers of black tobacco than for smokers of blond tobacco. Further, the urine of people who smoke black tobacco is more mutagenic than the urine of those who smoke blond tobacco (Silverman DT, 1996). The exact roles that pipe tobacco, cigars, and smokeless tobacco play in BC are unclear; however, most of the evidence suggests that pipe smokers are at increased risk for developing BC (Silverman DT, 1996).

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Smoking cessation has been found to reduce the risk of developing BC by 30–60% (Silverman DT, 1996). A reduction in smoking-related incidence of BC to a rate equal to that of the nonsmoking population comes ten years after cessation of smoking (Herr HW, 2001). The effect of reduced smoking is proportional to the length of time since quitting (Castelao JE, 2001). Other Risk Factors. Compared with smoking, other risk factors play a minor role in BC development. Nevertheless, they can potentiate a smoker’s risk of developing BC. •

•

•

Genetic susceptibility and carcinogen exposure. Glutathione S-transferases, including GSTM1, GSTP1, and GSTT1, play a role in detoxifying metabolites of carcinogens in tobacco smoke. GSTs are also involved in detoxifying the reactive metabolites of polycyclic aromatic hydrocarbons (PAHs) (Hung RJ, 2004). The data available for GSTP1 polymorphism and BC risk are inconclusive (Toruner GA, 2001; Harries LW, 1997; Katoh T, 1999). GSTM1 and GSTT1 polymorphisms may modulate individual susceptibility to BC (Hung RJ, 2004). Aromatic amines. The aromatic amines are urothelial carcinogens metabolized by arylamine N -acetyltransferases (NATs). Two types of NATs—NAT1 and NAT2 —are involved in the activation and detoxiÞcation of aromatic amines (Hein DW, 2000). The lack of two functional NAT2 alleles leads to a slow acetylation phenotype, while NAT1 *10 and *11 are associated with faster enzyme activity. A meta-analysis of NAT2 showed an increased risk of BC among smokers with slow acetylators (Marcus PM, 2000[a,b]). Data available for NAT1 in fast acetylators and BC risk are less conclusive (Hung RJ, 2004). Sulfotransferase 1A1 (SULT1A1 ) and cytochrome P450 1B1 (CYP1B1 ) are also involved in metabolizing PAHs and aromatic amines. Recent Þndings show that the risk of BC may be modulated by GSTM1, GSTT1, NAT2, and SULT1A1 polymorphisms, possibly in combination with exposure to aromatic amines (Hung RJ, 2004). Additionally, both GSTM1 and GSTT1 deletions appear to play a more signiÞcant role in bladder carcinogenesis among younger subjects. The GSTM1 null genotype also has a strong effect in heavy smokers, suggesting that GSTM1 null genotype is not an independent risk factor but acts synergistically with smoking on BC risk (Hung RJ, 2004). The Þndings show no association between NAT1 polymorphisms and BC risk. GSTM1, GSTT1, NAT2, and SULT1A1 polymorphisms may therefore modulate individual susceptibility to BC (Hung RJ, 2004). Gender. Approximately 75% of BC patients are men (van der Meijden APM, 1998). However, the risk of BC may eventually become higher in women. A population-based, case-control study involving 1,514 BC patients and 1,514 matched controls found that the risk of developing BC is higher in women than in men who smoked comparable numbers of cigarettes (Castelao JE, 2001). Researchers found that women who smoke have more elevated

ETIOLOGY AND PATHOPHYSIOLOGY

•

•

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levels of 3- and 4-ABP-hemoglobin adducts than do men who smoke comparable amounts. Arylamines (including ABPs) found in cigarette smoke play a major role in smoking-induced bladder carcinogenesis; consequently, women may have a higher risk of developing BC (Castelao JE, 2001). Diet. A prospective health professional follow-up study in 47,909 men over a ten-year period found that the intake of cruciferous vegetables (e.g., broccoli, cabbage, caulißower, brussels sprouts) is inversely associated with BC risk (Michaud DS, 1999[a]). The data have reached statistical signiÞcance was speciÞcally reached in the cases of broccoli and cabbage. This inverse association was strongest among nonsmokers. The data suggest that a diet in cruciferous vegetables may substantially reduce BC risk (Michaud DS, 1999[a]). Researchers proposed that an increase in cruciferous vegetable intake protects the bladder epithelium from carcinogens by enhancing detoxiÞcation of xenobiotics prior to excretion (Michaud DS, 1999[a]). The prospective evaluation of ßuid intake in the health professional follow-up study found a clear link between increased ßuid intake and a decreased risk of BC. The Þndings from this U.S. study suggest that high intake of water may reduce the risk of developing BC by approximately 50% (Michaud DS, 1999[b]). A recent population-based, case-control study involving 1,592 BC patients and 1,592 controls (matched by sex, date of birth [within Þve years], and race) found a strong inverse association between BC risk and intake of dark green vegetables, after adjusting for nondietary risk factors, including cigarette smoking (Castelao JE, 2004). The study also found a strong inverse association between BC risk and intake of citrus fruits/juices—or, in terms of nutrients, total carotenoids and vitamin C (Castelao JE, 2004). Consumption of fried meat and fat in high quantities is associated with increased risk of BC (Herr HW, 2001; Silverman DT, 1996). Arsenic. Ingestion of arsenic in drinking water is a strong risk factor for several forms of cancer, including BC. Long-term arsenic exposure is associated with a greater risk of developing BC. A recent study found that higher levels of arsenic exposure are associated with higher levels of chromosomal instability in patients with bladder tumors (Moore LE, 2002). The study also found that deletions in part or all of chromosome 9q are increased in tumors from patients exposed to high levels of arsenic. Most of the chromosomal alterations associated with arsenic exposure were also associated with tumor stage and grade, possibly suggesting that bladder tumors from arsenic-exposed patients may behave more aggressively and result in a higher mortality rate than tumors from unexposed patients (Moore LE, 2002). The molecular mechanisms by which arsenic causes cancer are uncertain, but researchers hypothesize that arsenic probably causes increased genetic instability in BC, possibly by deregulating the cell-cycle control pathways via epigenetic mechanisms or by reducing the cell’s ability to respond properly to or to repair DNA damage (Moore LE, 2002).

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Familial predisposition. Persons with family members who have BC are at increased risk of developing the disease (Silverman DT, 1996). The risk is high in patients with environmental exposure such as heavy cigarette smoking, implying that both genetic and environmental interactions play a role in BC. Race. White men are twice as likely as black men to be diagnosed with BC, and the diagnosis rate is higher in urban than in rural areas (Herr HW, 2001). The higher incidence in whites than in blacks is limited to localized cases; in more advanced tumors, the risk for whites and blacks is similar (Silverman DT, 1996). Asians and Hispanics have the lowest rates of BC, and BC risk is very low in American Indians (Silverman DT, 1996). Age. The incidence and mortality of BC increases with age; approximately two-thirds of all cases occur in people aged 65 or older (Silverman DT, 1996). The median age at diagnosis is between 68 and 69 years (ACS, 2004). BC is rarely diagnosed in people younger than 40 (Herr HW, 2001). Schistosoma haematobium. Exposure to S. haematobium, a parasite found in many developing countries, is associated with increased risk of both squamous-cell carcinoma and TTC of the bladder (Herr HW, 2001). Cyclophosphamide. High doses of cyclophosphamide, an alkylating agent used in the treatment of both malignant and nonmalignant diseases, increases the risk of BC. Cyclophosphamide was found to produce bladder tumors in both rats and mice (Silverman DT, 1996). Patients who are taking higher doses or taking cyclophosphamide for long periods of time are at increased risk for developing BC. Radiation. Ionizing radiation causes BC, but this exposure contributes very little to the incidence of BC in the general population (Silverman DT, 1996).

Genetic Mutations. Studies of different stages and grades of BC have shown that mutations in proto-oncogenes and TSGs may be critical to the development and progression of bladder tumors. In general, loss of TSGs—the p53 on chromosome 17 and the retinoblastoma [Rb, on chromosome 13]) are common in patients with invasive disease, while aberrations of 9q occur mainly in superÞcial tumors (Herr HW, 2001). Deletions of several chromosomal arms—including 9p, 9q, 11p, 13q, and 17p—have been noted in BC (Cote RJ, 2003). Deletions of chromosome 9 are the most common abnormalities associated with BC (Quek ML, 2003). In fact, chromosome 9 alterations may represent an early event in TCC, because deletions of this chromosome are found in both low- and highgrade BC (Quek ML, 2003). The majority of chromosome 9 deletions involve the 9p21 locus, which encodes three distinct proteins: p16, p14, and p15. All three proteins act as negative cell-cycle regulators and are therefore considered potential TSGs (Quek ML, 2003; Williams SG, 2001). Additionally, deletions on the short arm of chromosomes 3 (3p) and 8 (8p) have been found in high-grade, muscle-invasive BC, though the relevant genes have yet to be identiÞed (Quek ML, 2003; Williams SG, 2001).

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Abnormalities in Rb and p53 genes are pervasive in many types of cancer, including BC, and are often important in the cascade of events that leads to the malignant phenotype. When either p53 or Rb gene product is missing or altered, negative checks on cell growth are deactivated; the “switch” is turned on, and uncontrolled cell proliferation ensues. The pervasiveness of these genetic abnormalities has led researchers to suspect that these genes act as “master switches” that control tumor formation. Table 4 brießy describes the oncogenes and TSGs implicated in BC and their clinical signiÞcance. Evidence suggests that p53 and pRb proteins have a cooperative negative effect on both progression and survival in primary BC. A marked increase in progression and decreased overall survival have been observed in BC patients with both p53 and pRb alteration (Cordon-Cardo C, 1997). Studies have shown that patients with p53 wild-type or pRb wild-type tumors are at signiÞcantly decreased risk of developing BC recurrence and mortality, compared with patients in whom both proteins are altered (Shariat SF, 2004). Altered expression of p53 and pRb/p16 and p21 is associated with an increased risk of BC progression and death (Shariat SF, 2004). Recurrent Disease. Generally, superÞcial BC is fairly benign. Approximately 10–30% of patients with superÞcial tumors will progress to muscle-invasive or metastatic disease, despite frequent recurrences; recurrence rates after initial successful treatment range from 30% to 85%, with grade progression occurring in 10–30% of patients and stage progression occurring in 4–30% of patients (Crawford ED, 2003). Despite complete eradication of the primary tumor, approximately two-thirds of patients will develop tumor recurrences within the Þrst Þve years of follow-up (Crawford ED, 2003). In contrast, invasive tumors are highly malignant and have a less favorable prognosis (Al-Sukhun S, 2003); for example, approximately 50% of patients treated locally for invasive tumors relapse with metastases within two years of treatment (Williams SG, 2001). Therefore, the prevention of invasive disease and of noninvasive superÞcial disease continues to be the ultimate goal for researchers interested in preventing BC and improving treatment outcomes. CURRENT THERAPIES Unless noted otherwise, our deÞnition of bladder cancer (BC) is mainly interchangeable with that of transitional cell cancer (TCC) of the urothelium. BC and TCC are not pathologically the same. However, we have grouped all forms of BC together when evaluating clinical data because TCC encompasses more than 90% of all BCs (Herr HW, 2001), and because other pathological types of BC are treated similarly (staging and grading being more important determinants of treatment and patient stratiÞcations in clinical trials). Treatment options for BC patients include surgery, radiotherapy, intravesical pharmacotherapy, and systemic chemotherapy. Treatment decisions are guided primarily by the stage of disease, although the pathological grade of the disease

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TABLE 4. Select Genetic Mutations in Bladder Cancer Gene/Protein

Function

Tumor suppressor genes p53 The p53 protein normally prevents the propagation of DNA damage by causing cell arrest at the G1/S checkpoint through the induction of p21 and inducing apoptosis. Mutations in the p53 gene remove this regulatory control, allowing mutation-carrying cells to progress through the cell cycle. Mutated p53 results in the nuclear accumulation of the abnormal protein with a prolonged half-life (Al-Sukhun S, 2003). Generally, an intact p53 regulatory mechanism is thought to be necessary for the activity of DNA-damaging cytotoxic agents.

p21

pRb/p16

p21 is an important target of the p53 protein. The p21 protein is both a p53-inducible and p53-independent cyclin-dependent kinase inhibitor that can arrest the cell by inhibiting DNA replication (Shariat SF, 2004; Zlotta AR, 2000). The retinoblastoma tumor suppressor gene (Rb) encodes a protein (pRb) that acts downstream of p53, also causing cell arrest at the G1/S checkpoint. The p16 gene regulates pRb function by inhibiting CDK4 enzyme activity. Inactivation of this gene therefore has the downstream effect of disrupting pRb-mediated cell-cycle control.

Comments A recent study found altered expression of p53 in 56% of patients undergoing radical cystectomy (45 out of 80) (Shariat SF, 2004). Altered expression of p53 was independently associated with disease progression. p53 was the strongest predictor of BC outcome, followed by p21, suggesting a fundamental role of the p53/p21 pathway in BC progression (Shariat SF, 2004). Tumors negative for both p53 and p21 proteins, even in high-risk superficial BC, seem to have favorable prognosis in terms of recurrence (Zlotta AR, 2000). A previous study found that accumulation of p53 protein in the tumor cell nuclei is associated with an increased risk of recurrence and decreased overall survival in patients with transitional-cell carcinoma (TCC) confined to the bladder (Esrig D, 1994). Altered expression of p21 was found in 49% of patients undergoing radical cystectomy (39 out of 80) (Shariat SF, 2004). The p21 protein seems to be associated with the early stages of BC progression. pRb or p16 proteins were altered in 54% of patients undergoing radical cystectomy (43 out of 80) (Shariat SF, 2004). Altered expression of p53, pRb/p16 and p21 is associated with an increased risk of BC progression and death (Shariat SF, 2004). While p21 seems to be associated with the early stages of BC progression, p53 and pRb/p16 seem to be associated with the later stages. Several other studies have shown significant association between loss of the Rb gene, tumor stage, and tumor grade; the data suggest that this loss is more commonly associated with invasive BC (Al-Sukhun S, 2003).

TABLE 4. (continued) Gene/Protein Oncogenes E2F3

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Function

Comments

E2F3, a candidate BC oncogene located on chromosome 6p22, is the latest oncogene to be implicated in BC. E2F3 protein is involved in controlling cell division. E2F3 appears to provide growth advantage to tumor cells by activating cell proliferation in a subset of bladder tumors (Oeggerli M, 2004).

Overexpression of E2F3 protein was found to be associated with cell proliferation and development of BC (Feber A, 2004). A high level of E2F3 protein was associated with tumor stage and grade (Feber A, 2004). Researchers have also found that a high level of E2F3 protein expression is associated with high grade, advanced stage, and E2F3 gene amplification (Oeggerli M, 2004). E2F3 was found to be frequently amplified and overexpressed in invasive BC (stage pT1-T4) (Oeggerli M, 2004). Mutations of the H-ras gene have been implicated in the development and progression of BC. Alterations in codon 12 and 61 of the H-ras gene were found in up to 20% of BC (Quek ML, 2003). cH-ras overexpression is correlated with early recurrence in patients with superficial BC; however, subsequent studies have not found this prognostic effect (Quek ML, 2003). Myc genes are overexpressed in high-grade bladder tumors, but correlation with progression or survival has not been found (Al-Sukhun S, 2003). Further studies are needed to determine the true prognostic role of c-myc gene expression in BC. Some studies found a positive association between overexpression of c-erbB-1 and high-grade, high-stage BC, thereby identifying this receptor as an independent prognostic factor in patients with advanced BC (Al-Sukhun S, 2003). Several studies have found that overexpression of HER-2/neu in patients with BC is correlated with higher-stage tumors, increased tumor progression, greater incidence of metastases, and reduced overall survival (Quek ML, 2003; Williams SG, 2001). Other studies found that HER-2/neu provide no additional prognostic value compared with previously established markers of grade and stage for TCC.

cH-as

The ras family of genes encodes for the signal transduction proteins involved in transmitting growth-promoting signals.

c-myc

The myc family of genes are nuclear transcription factors that activate genes that drive cell growth.

Epidermal growth factor receptor (c-erbB-1)

Receptor tyrosine kinase is involved with epithelial cell growth and division. Overexpression of the growth receptor on the surface of cells enhances their metastatic potential.

Epidermal growth factor receptor (c-erbB-2, HER-2/neu)

Receptor tyrosine kinase is involved with epithelial cell growth and division. Overexpression of the growth receptor on the surface of cells enhances their metastatic potential.

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TABLE 4. (continued) Gene/Protein

Function

bcl-2

The bcl-2 (B-cell lymphoma/leukemia-2) gene encodes a protein that blocks apoptosis. Overexpression of bcl-2 allows cells to escape apoptosis.

Survivin

An apoptosis inhibitor that is expressed during fetal development but is normally undetectable in adulthood.

Cyclin D1

Cyclin D1, a key cell-cycle regulator during the G1/S phase, is a putative proto-oncogene overexpressed in several cancers, including TCC of the bladder.

MDM2

MDM2 gene inhibits the cell-growth arrest mediated by wild-type p53 protein.

Other relevant events in BC Ki-67 The Ki-67 biomarker is a proliferation antigen detected by immunohistochemical staining. Tumors with positive Ki-67 represent actively growing tumor cells.

Comments Bcl-2 expression in bladder tumors is associated with a less aggressive phenotype (Shiina H, 1996). However, others have found bcl-2 overexpression to be significantly associated with disease progression and upstaging of the tumor during radiotherapy in patients with invasive BC (Pollack A, 1997; Rodel C, 2000). Previous studies found survivin in 78% of patients with BC but not in normal urothelium, and its expression correlated with increased rate of recurrence (Swana HS, 1999). Urine detection of survivin appears to be a marker for diagnosing patients with new or recurrent BC (Smith SD, 2001). In contrast, recent findings suggest that survivin expression does not correlate with early relapse; rather, its expression seems to be associated with the histological grade of tumors (Gazzaniga P, 2003). The cyclin D1 gene (CCND1) variant A allele may be associated with an increased risk of TCC of the bladder, especially in men without a history of smoking (Wang L, 2002). Recent findings suggest that while CCND1 polymorphism cannot serve as a prognostic marker for BC, the CCND1 variant A allele may increase the risk of CIS incidence in patients with superficial BC (Ito M, 2004). The role of MDM2 in regulating p53 protein levels in transitional carcinoma of the bladder is unclear. MDM2 gene amplification is not common in BC (Williams SG, 2001). Researchers have found that the proliferation markers, including Ki-67 and proliferating cell nuclear antigen (PCNA), correlate with grade, stage, and risk of recurrence (Gontero P, 2000; Bush C, 1991; Cheng G, 1997). The Ki-67 nuclear antigen also appears to be associated with invasive cancers and worse prognosis. It appears to be the most promising single marker in predicting BC recurrence and progression (Kausch I, 2002).

TABLE 4. (continued) Gene/Protein E-cadherin Basic fibroblast growth factor (bFGF) Vascular endothelial growth factor (VEGF) Matrix metalloproteinases (MMPs)

Function The presence of E-cadherin is essential for full adhesive function of the cell (Pirinen RT, 2001). bFGF promotes vascular repair and angiogenesis both in vivo and in vitro.

VEGF is essential in the regulation of endothelial cell proliferation and angiogenesis, both of them key contributors to the growth of cancer and vascular diseases. MMPs are involved in tumor-associated degradation of the extracellular matrix.

CD44

CD44, a widely expressed adhesion molecule, is involved in cell-cell and cell-matrix interactions as well as signal transduction through ras in response to hyaluronic acid (Quek ML, 2003).

Thymidine kinase (TK)

TK is an enzyme that plays a key role in the complementary, or alternative, salvage pathway of pyrimidine synthesis.

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Full source citations appear in ‘‘References.’’

Comments The loss of E-cadherin is associated with increased invasion and metastasis in BC (Cresswell J, 2002). Higher levels of bFGF have been found in the urine of BC patients than in the urine of controls. Urinary bFGF levels were also correlated with pathological stage in patients with muscle-invasive BC (Quek ML, 2003). Higher levels of VEGF have been found in the urine of BC patients than in the urine of controls. VEGF levels were also correlated with tumor recurrence in patients with Ta and T1 disease (Quek ML, 2003; Williams SG, 2001). Elevated levels of MMP-2 and MMP-9 have been found in the serum and urine of patients with muscle-invasive TTC and correlated with a decreased disease-free survival (Quek ML, 2003). Additionally, elevated levels of MMP-9 have been found in TTC compared with normal urothelium and correlated with increasing tumor stage (Quek ML, 2003; Williams SG, 2001). Increased CD44 expression has been found in superficial TTC, with a fall in expression at the time of muscle invasion. Data suggest that CD44 status is a prognostic marker in urothelial cancer and may be a potential target when screening BC patients (Quek ML, 2003). A recent study found that TK activity is correlated with both stage and grade of BC and that increased TK activity predicted early recurrence in Ta and T1 BC (Mizutani Y, 2003). TK may therefore be used as a prognostic marker in patients with BC.

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is also important. Physicians across the major markets now stage BC according to the tumor, node, metastasis (TNM) staging system. However, only three clinically relevant stages of cancer can be correlated with the TNM staging system: superÞcial BC (SBC), muscle-invasive BC (MIBC), and metastatic BC (MBC). Advanced BC (ABC) usually comprises cases in the MIBC and MBC categories. SBC can be further subdivided into low risk, intermediate risk, and high risk of progression or recurrence, which further takes into consideration pathological grading of disease, size, and number of lesions (G1 being a low grade and G3 being a high grade). The subdivisions of SBC guide decisions on the use of intravesical therapy in conjunction with surgery. The overall aim of SBC treatment is to prevent recurrence and progression to incurable disease; the main decision here is whether, in addition to transurethral resection (TUR), intravesical pharmacotherapy is warranted. The aim of MIBC treatment is to determine which tumors can be treated curatively with surgery and which require an integrated systemic approach to achieve cure. The aim of MBC treatment is to prolong survival and palliate disease symptoms because, at this stage, the disease is too advanced for the majority of patients to be cured. Combination chemotherapy is the standard approach for MBC. Table 5 lists the most common chemotherapy regimens used to treat BC. We discuss them in the following sections. Gemcitabine/Cisplatin Overview. The combination regimen of gemcitabine (Eli Lilly/Spaly Bioquimica’s Gemzar) (Figure 3) and cisplatin (Bristol-Myers Squibb’s Platinol-AQ, Nippon Kayaku’s Randa, generics) (Figure 4) is the new gold-standard therapy for the treatment of MBC. Gemcitabine/cisplatin (GC) is indicated for the treatment of ABC (muscle-invasive stage IV tumors with or without metastases). GC is also one of the most widely used regimens for the treatment of non-small-cell lung cancer. Gemcitabine and cisplatin have complementary mechanisms of action; in preclinical studies, gemcitabine was found to be synergistic with cisplatin (Bergman AM, 1996). GC is increasingly replacing methotrexate/vinblastine/doxorubicin/cis platin (MVAC) and cisplatin/methotrexate/vinblastine (CMV) and has been investigated in several settings, including as Þrst-line therapy in metastatic disease and in neoadjuvant and adjuvant settings. In some cases of BC, the patient is too ill to tolerate the side effects of cisplatin. In such cases, researchers have substituted carboplatin for cisplatin. Carboplatin is a platinum agent, like cisplatin, but is less toxic. We discuss the results of studies examining this substitution in this section as well. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the overall regimen activity: •

Gemcitabine is an antimetabolite. Antimetabolites block normal DNA synthesis, thus stopping cell replication. Gemcitabine exhibits cell-phase speciÞcity, primarily killing cells that are undergoing DNA synthesis

TABLE 5. Current Regimens/Classes Used for Bladder Cancer Regimen Components Regimen

Agent

Availability

Dose

Gemcitabine (Eli Lilly/Spaly Bioquimica’s Gemzar)

US, F, G, I, S, UK, J

Gemcitabine: 1,000 mg/m2 IV days 1, 8, 15.

US, F, G, I, S, UK, J

Cisplatin: 70 mg/m2 IV day 2. Cycle repeated every 28 days

Gemcitabine, single agent

Cisplatin (Bristol-Myers Squibb’s Platinol-AQ, Nippon Kayaku’s Randa, generics) Gemcitabine (Eli Lilly/Spaly Bioquimica’s Gemzar)

US, F, G, I, S, UK, J

Gemcitabine: 1,200 mg/m2 IV days 1, 8, 15. Cycle repeated every 28 days.

MVAC

Methotrexate (generics)

US, F, G, I, S, UK, J

Methotrexate: 30 mg/m2 /d IV days 1, 15, 22.

Vinblastine (Eli Lilly’s Velbe, Eli Lilly/Shionogi’s Exal, generics) Doxorubicin (Pfizer’s Adriamycin/Adriblastine, Kyowa’s Adriacin, generics) Cisplatin (Bristol-Myers Squibb’s Platinol-AQ, Nippon Kayaku’s Randa, generics)

US, F, G, I, S, UK, J

Vinblastine: 3 mg/m2 /d IV days 2, 15, 22. Doxorubicin: 30 mg/m2 IV day 2.

Gemcitabine/cisplatin

US, F, G, I, S, UK, J

US, F, G, I, S, UK, J

Cisplatin: 70 mg/m2 IV day 2. Cycle repeated every 28 days.

Grade III/IV Toxicities Neutropenia Anemia Thrombocytopenia Nausea/vomiting Alopecia Neutropenia Thrombocytopenia Nausea/vomiting Edema Respiratory Leukopenia Neutropenia Neutropenic fever Thrombocytopenia Mucositis Nausea/vomiting Alopecia

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TABLE 5. (continued) Regimen Components Regimen CMV

Bacillus ´ Calmette-Guerin (BCG)

Mitomycin, single agent

Agent

Availability

Dose

Cisplatin (Bristol-Myers Squibb’s Platinol-AQ, Nippon Kayaku’s Randa, generics)

US, F, G, I, S, UK, J

Cisplatin: 70 mg/m2 IV day 2.

Methotrexate (generics) Vinblastine (Eli Lilly’s Velbe, Eli Lilly/Shionogi’s Exal, generics) Organon’s Tice BCG/OncoTICE, Sanofi-Aventis’s Theracys/ImmuCyst, Nippon Kayaku’s ImmuCyst, Japan BCG’s Immunobladder, generics

US, F, G, I, S, UK, J US, F, G, I, S, UK, J

Methotrexate: 30 mg/m2 IV days 1, 8. Vinblastine: 4 mg/m2 IV days 1, 8. Cycle repeated every 21 days. BCG: 1 instillation per week for six weeks (induction therapy). May be repeated once if tumor remission has not been achieved and if the clinical circumstances warrant. Thereafter, intravesical BCG vaccine administration continues at monthly intervals for at least 6–12 months.

Mitomycin (generics)

US, F, G, I, S, UK, J

US, F, G, I, S, UK, J

Mitomycin: 20 mg (prophylactic protocol) and 40 mg (ablative protocol) for six to eight weeks (induction therapy).

Grade III/IV Toxicities Leukopenia Thrombocytopenia Neutropenic fever

Urinary frequency Hematuria Cystitis Dysuria Urgency Urinary tract infection Flu-like syndrome Fever Shaking/chills Malaise/fatigue Cardiac Cramps/pain Dysuria Cystitis Urinary frequency Urgency Hematuria Cramps/pain Chills Fever Malaise/fatigue Rash/itching

TABLE 5. (continued) Regimen Components Regimen Doxorubicin, single agent

Agent Pfizer’s Adriamycin/Adriblastine, Kyowa’s Adriacin, generics

Epirubicin, single agent Pfizer’s Ellence/Farmorubicin, Pfizer/Kyowa’s Farmorubicin

Thiotepa, single agent

Amgen/Wyeth Lederle’s Thioplex, Sumitomo’s Tespamin, generics

Availability

Dose

Grade III/IV Toxicities

US, F, G, I, S, UK, J Doxorubicin: 50–80 mg administered Dysuria intravesically, weekly for 4–8 weeks. Frequency May be repeated at monthly intervals. Hematuria Urgency Cystitis Nausea/vomiting Bladder cramps/pain US, F, G, I, S, UK, J Epirubicin: 100 mg administered Chemical cystitis intravesically immediately after Fever transurethral resection (single dose). Dysuria Or 50 mg weekly for 4–8 weeks. US, F, G, I, S, UK, J Thiotepa: 60 mg administered Dysuria intravesically, weekly for four weeks. Urinary retention May be repeated if necessary, but Chemical cystitis second and third courses must be (rare) given with caution because Hemorrhagic cystitis bone-marrow depression may be (rare) increased. Myelosuppression

US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan. IV = Intravenous.

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NH2 N HO

O O

OH

N F F

FIGURE 3. Structure of gemcitabine.

FIGURE 4. Structure of cisplatin.

•

(S phase) and blocking the progression of cells through the G1/S-phase boundary. The agent is metabolized intracellularly by nucleoside kinases to its active form. Cisplatin is a platinum agent. Platinum agents generate highly reactive charged platinum complexes that bind covalently to DNA, leading to intrastrand, interstrand, and protein cross-linking of DNA that result in inhibition of transcription and/or DNA replication mechanisms.

Clinical Performance. A randomized, multinational Phase III trial compared GC with MVAC in 405 chemotherapy-naive patients with locally advanced or metastatic TCC of the urothelium (von der Maase H, 2000). The primary objective was comparison of overall survival; secondary objectives were comparisons of objective tumor responses, duration of responses, times to disease progression, times to treatment failure, toxicity, changes in performance status and weight, and evaluations of quality of life and medical resource utilization. Patients were randomized to receive either a maximum of six cycles of GC (203 patients) or a maximum of six cycles of MVAC (202 patients) between November 1996 and September 1998. Investigators observed no signiÞcant difference in median overall survival (GC, 13.8 months; MVAC, 14.8 months); time to disease progression, (7.4 months for both arms); or time to treatment failure (GC, 5.8 months; MVAC, 4.6 months). Investigators assessed overall response rates in 164 patients in the GC arm and 151 in the MVAC arm and found similar response rates: GC, 49.4% (81 patients) with 12.2% complete response (CR) and 37.2% partial response (PR); MVAC, 45.7% (69 patients) with 11.9% CR and 33.8% PR. One-third

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of the patients in both arms achieved stable disease. The median durations of response were 9.6 months for the GC arm and 11 months for the MVAC arm. GC caused more grade 3 or 4 anemia (GC, 27%; MVAC, 18%) and thrombocytopenia (GC, 57%; MVAC, 21%) but did not result in a greater number of transfusions. MVAC caused more grade III or IV neutropenia (MVAC, 82%; GC, 71%) and neutropenic fever (MVAC, 14%; GC, 2%) and signiÞcantly more neutropenic sepsis (MVAC, 12%; GC, 1%). Grade III or IV mucositis (MVAC, 22%; GC, 1%) and alopecia (MVAC, 55%; GC, 11%) were also signiÞcantly more common in the MVAC arm. The drug toxicity–related mortality rate was lower for the GC arm (1%) than for the MVAC arm (3%), but the difference was not statistically signiÞcant. Quality of life was maintained during treatment in both arms, but more patients on GC did better regarding weight, performance status, and fatigue. Based on this study, GC provided a survival advantage similar to that of MVAC, but with a better safety proÞle and tolerability. Investigators concluded that GC should be considered the new standard of care for patients with locally advanced or metastatic TCC (von der Maase H, 2000). Results of a neoadjuvant Phase III Egyptian trial presented at the American Society of Clinical Oncology (ASCO) in 2003 showed that the GC regimen is effective and well tolerated when used as neoadjuvant therapy in MIBC (stages T2–T4, N0–N2) (Khaled HM, 2003). A total of 58 patients were randomized to receive three cycles of GC chemotherapy preoperatively, and 56 patients had only radical cystectomy between November 2000 and June 2002. Patients with a CR received three additional chemotherapy cycles followed by radical radiotherapy. The overall response rate was 56% (28 patients) in the GC arm: 30% CR and 26% PR. Eight patients were not evaluable for response. Bladder preservation was feasible in 11 of the patients achieving CR. In the radical cystectomy–only arm, 52 patients underwent radical cystectomy and 4 were found unresectable on exploration. Researchers observed a trend toward increased overall one-year survival for neoadjuvant GC (69%) compared with cystectomy alone (54%). The overall survival for all patients in the study was 63%. Occasional grade III or IV toxicity was observed. The investigators found neoadjuvant GC to be effective and tolerable in MIBC and helped achieve organ preservation in a signiÞcant subset of patients. There is also insufÞcient evidence to support the routine use of adjuvant chemotherapy for the treatment of patients with locally advanced BC because the trials have employed small sample sizes and confusing methodologies (Sylvester R, 2000). However, adjuvant chemotherapy is generally used more often than neoadjuvant chemotherapy. In a retrospective study presented at ASCO 2004, 25 patients with locally advanced BC were treated with adjuvant chemotherapy between 1997 and 2002 (El-Khoueiry AB, 2004). The researchers determined that adjuvant GC chemotherapy was both feasible and effective. Over a median follow-up of 25 months, 44% had grade III or IV hematologic toxicity and 8% had grade III nonhematologic toxicity. The median time to recurrence was 49.7 months, and median overall survival exceeded 49.7 months. Eight

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patients died. Other Phase III trials are ongoing in the United States to investigate the effect of adjuvant chemotherapy. Carboplatin-based regimens are widely used as an alternative to cisplatin-based regimens in unÞt patients. “UnÞt” refers to those with poor performance status and poor renal function. Thirteen trials involving 327 patients with advanced urothelial cancer found that single-agent carboplatin elicited objective responses in 14% of patients: 3% CR and 11% PR (Mottet-Auselo N, 1993). A Phase II dose-Þnding study evaluated the combination of gemcitabine and carboplatin in unÞt patients with advanced BC (Bellmunt J, 2001). Researchers reported an overall response rate of 44%: 6% CR and 38% PR in 16 patients ineligible for the cisplatin-based regimens. At evaluation, six patients had stable disease and three had progressive disease. The gemcitabine/carboplatin regimen appears to be active and well-tolerated and shows acceptable toxicity in this patient population. This encouraging result prompted the European Organization for the Research and Treatment of Cancer (EORTC) to conduct a randomized Phase II/III trial, which is ongoing, to compare gemcitabine/carboplatin with methotrexate/carboplatin/vinblastine in patients ineligible for cisplatin-based chemotherapy. The trials are still open at the time of composing this reference. Gemcitabine, Single Agent (under open trials at the time of composing this reference) Overview. Single-agent gemcitabine (Eli Lilly/Spaly Bioquimica’s Gemzar) has also demonstrated promising activity against advanced or metastatic urothelial cancer. The agent is generally used for patients with poor renal function who cannot tolerate cisplatin-based therapy. Mechanism of Action. •

Gemcitabine is an antimetabolite. Antimetabolites block normal DNA synthesis, thus stopping cell replication. Gemcitabine exhibits cell-phase speciÞcity, primarily killing cells that are undergoing DNA synthesis (S phase) and blocking the progression of cells through the G1/S-phase boundary. The agent is metabolized intracellularly by nucleoside kinases to its active form.

Clinical Performance. Overall response rates for gemcitabine obtained from two Phase II trials in previously treated patients were 22.5% and 29%, respectively (Lorusso V, 1998; Gebbia V, 1999). Two studies evaluating gemcitabine in previously untreated patients conÞrmed its relatively high single-agent activity (Stadler WM, 1997; Moore MJ, 1997). These studies achieved overall response rates of 28% (Stadler WM, 1997) and 24.3% (Moore MJ, 1997). A Phase II study assessed the ablative efÞcacy of intravesical gemcitabine at a concentration of 40 mg/mL (2,000 mg in 50 mL saline solution) in 39 patients with intermediate-risk SBC (Ta, T1, G1, G2) (Gontero P, 2004). Gemcitabine was

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administered weekly for six weeks on a single marker tumor left in the bladder after a complete TUR of all other lesions. A complete response was achieved in 22 (56%) of the 39 patients. Investigators observed no progression among the 17 nonresponders. Side effects generally did not exceed grade I toxicity. Researchers concluded that the ablative effect of gemcitabine produced a higher number of responses than the minimum required by the protocol to indicate a signiÞcant probability of drug efÞcacy. The durability of response should be tested in Phase III trials. MVAC Overview. The regimen known as MVAC is a combination of methotrexate (generics) (Figure 5), vinblastine (Eli Lilly’s Velbe, Eli Lilly/Shionogi’s Exal, generics) (Figure 6), doxorubicin (PÞzer’s Adriamycin/Adriblastine, Kyowa’s Adriacin, generics) (Figure 7), and cisplatin (Bristol-Myers Squibb’s PlatinolAQ, Nippon Kayaku’s Randa, generics). For more than 15 years it has been the most widely used regimen for the treatment of patients with locally advanced or metastatic BC (Culine S, 2002). Randomized trials show MVAC to be superior to cisplatin alone, as well as to cisplatin in combination with cyclophosphamide and doxorubicin. However, the regimen is associated with signiÞcant toxicities, which are especially problematic for older patients, who often present with comorbidities and who constitute a signiÞcant proportion of the population with TCC (von der Maase H, 2000). Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

Methotrexate, a structural analogue of folate, is an antimetabolite. Antimetabolites block normal DNA synthesis by inhibiting several key enzymes. Methotrexate inhibits the enzyme dihydrofolate reductase. This inhibition interferes with the maintenance of the reduced folate pool. Reduced folates are essential for de novo synthesis of thymidylate and purine nucleotides, a requirement for DNA synthesis and cell replication. Thus, by inhibiting dihydrofolate reductase, methotrexate stops the replication of rapidly proliferating cells such as tumor cells.

H2N

N

N CH3

N

N N H N

NH2 O

COOH COOH

FIGURE 5. Structure of methotrexate.

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CH3

N

HN

OH

H3CO O

H

N H

H3CO

CH3

OH N R1

R3 H

R2

FIGURE 6. Structure of vinblastine (R1 = CH3 , R2 = CO(OCH3 ), R3 = OCO(CH3 ).

FIGURE 7. Structure of doxorubicin (R = OCH3 , R1 = OH, R2 = H, R3 = H, R4 = OH).

•

•

Vinblastine is a vinca alkaloid. Vinca alkaloids interact with tubulin and disrupt microtubular function in the mitotic spindle. This action leads to metaphase arrest, resulting in mitotic arrest and cell death. Doxorubicin is an anthracycline. Anthracyclines interact with several different cellular targets, most importantly topoisomerase II. By inhibiting this DNA re-ligation enzyme, anthracyclines exert their cytotoxic effect. Another doxorubicin mechanism that leads to cell death is known as DNA intercalation, in which the anthracycline molecule inserts itself between base pairs. This phenomenon causes single- and double-stranded breaks in DNA that inhibit cell proliferation. The free radicals generated by the reductive metabolism of anthracyclines may also damage cellular structures such as proteins and DNA.

CURRENT THERAPIES

•

167

Cisplatin is a platinum agent. Platinum agents generate highly reactive charged platinum complexes that bind covalently to DNA, leading to intrastrand, interstrand, and protein cross-linking of DNA that results in inhibition of transcription and/or DNA replication mechanisms.

Clinical Performance. Investigators at the Memorial Sloan-Kettering Cancer Center in New York Þrst described the standard schedule for MVAC in 1985 (Sternberg CN, 1985). Among 83 evaluable patients with advanced BC, 69% achieved an objective response, including a CR rate of 37% (Sternberg CN, 1988). In a multicenter French study of 70 patients with advanced BC, investigators reported an objective response rate of 57% (38 of 67 patients), with a CR rate of 19% (Boutan-Laroze A, 1991). Randomized trials have conÞrmed the superiority of the MVAC regimen, in terms of objective response rates and overall survival, over single-agent cisplatin or cisplatin combined with cyclophosphamide and doxorubicin. A randomized trial investigating MVAC compared with cisplatin alone found MVAC to be superior to single-agent cisplatin with respect to response rate, duration of remission, and overall survival (Loehrer PJ, 1992). Of the 269 patients with advanced urothelial cancer enrolled between October 1984 and May 1989, 246 were randomized to receive cisplatin alone (126) or MVAC (120). The response rate was far better for MVAC (39%) than for cisplatin (12%). The progression-free survival (10 months versus 4.3 months) and overall survival (12.5 months versus 8.2 months) were signiÞcantly greater for the MVAC-treated patients. However, the MVAC regimen was associated with more toxic side effects, particularly leukopenia, mucositis, granulocytopenic fever, and drug-related mortality. Long-term follow-up of the Phase III intergroup study of advanced urothelial cancer patients treated with MVAC conÞrmed that, with the MVAC regimen, survival is superior to cisplatin: 3.7% of MVAC patients experienced more than six years of disease-free survival compared with only 1.6% of patients treated with single-agent cisplatin (Saxman SB, 1997). A randomized, multinational Phase III trial compared GC with MVAC in 405 chemotherapy-naive patients with ABC (von der Maase H, 2000). (See the previous section on the GC regimen for details of the trial.) The researchers concluded that the standard of care for patients with ABC should be changed from MVAC to GC because the latter provides a survival advantage similar to that of MVAC but has a better safety proÞle and tolerability. In the trial, the drug toxicity-related mortality rate was lower for the GC arm (1%) than for the MVAC arm (3%), though the difference was not statistically signiÞcant. To improve survival rates achievable with the MVAC regimen, the EORTC conducted a randomized Phase III trial comparing high-dose-intensity MVAC (HD-MVAC) and granulocyte colony-stimulating factor (G-CSF) versus classic MVAC in advanced urothelial tract tumors (Sternberg CN, 2001). The trial randomized 263 chemotherapy-naive patients with metastatic or advanced TCC to either HD-MVAC (two-week cycle) or MVAC (four-week cycle) between June 1993 and November 1998. Using an intent-to-treat analysis over a median

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follow-up of 38 months, the overall response rate in the HD-MVAC arm was 62%: 21% CR and 41% PR. The overall response rate in the MVAC arm was 50%: 9% CR and 41% PR. Progression-free survival was signiÞcantly better with HD-MVAC. The median progression-free survival time was 9.1 months in the HD-MVAC arm and 8.2 months in the standard MVAC arm. The two-year progression-free survival was 24.7% for HD-MVAC compared with 11.6% for standard MVAC. Researchers concluded that, with HD-MVAC, it was possible to deliver twice the doses of cisplatin and doxorubicin in half the time, resulting in fewer dose delays and less toxicity (Sternberg CN, 2001). The clinical beneÞts, however, are not considered large enough to justify adopting this regimen as standard therapy (Hussain SA, 2003). However, both the HD-MVAC and GC regimens have emerged as promising candidates to end the dominance of MVAC in the treatment of advanced BC (Culine S, 2002). A randomized Phase III intergroup study compared neoadjuvant chemotherapy plus cystectomy with cystectomy alone in patients with MIBC (stage T2–T4a) (Grossman HB, 2003). The primary objective was the comparison of survival; the secondary objective was to quantify the effect of neoadjuvant MVAC on tumor stage (downstaging of the tumor). Patients were randomly assigned to radical cystectomy alone (154 patients) or three cycles of MVAC followed by radical cystectomy (153 patients with adequate renal function) between August 1987 and July 1998. Radical cystectomy included a bilateral pelvic lymphadenectomy, and urinary diversions were performed. Investigators found a signiÞcant and clinically meaningful improvement in survival among patients who received neoadjuvant chemotherapy. Over a median follow-up of 8.7 years, 90 deaths occurred in the MVAC and cystectomy arm; over a median follow-up of 8.4 years, 100 deaths occurred in the cystectomy alone arm. The estimated risk of death was reduced by 25% in the MVAC and cystectomy arm compared with the cystectomy-alone arm. Using an intention-totreat analysis, the median survival was 77 months among patients in the MVAC and cystectomy arm and 46 months among patients in the cystectomy alone arm. Approximately 57% of patients in the MVAC and cystectomy arm were alive at Þve years, compared with 43% of patients in the cystectomy-alone arm. Investigators reported that 38% of the surgical specimens of patients in the MVAC and cystectomy arm were pathologically free of cancer (pT0) at the time of surgery, compared with 15% of patients in the cystectomy-alone arm. The overall adverse effects were moderate: at least one-third of patients had severe hematologic or gastrointestinal toxicities, although all the patients recovered and no treatmentrelated deaths occurred. Furthermore, MVAC did not adversely affect a patient’s chance of undergoing radical cystectomy; nor did it increase the risk of death or complications related to the surgery. MVAC can therefore be administered safely before radical cystectomy to patients with locally advanced BC. Investigators concluded that the use of neoadjuvant MVAC followed by radical cystectomy is associated with improved survival in patients with locally advanced BC.

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CMV Overview. The regimen known as CMV combines cisplatin (Bristol-Myers Squibb’s Platinol-AQ, Nippon Kayaku’s Randa, generics), methotrexate (generics), and vinblastine (Eli Lilly’s Velbe, Eli Lilly/Shionogi’s Exal, generics). CMV is superior to methotrexate and vinblastine (Mead GM, 1998) and has shown response rates similar to those of MVAC. It was widely used in Europe for patients with metastatic TTC of the bladder before the introduction of the GC regimen. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

•

•

Cisplatin is a platinum agent. Platinum agents generate highly reactive charged platinum complexes that bind covalently to DNA, leading to intrastrand, interstrand, and protein cross-linking of DNA that results in inhibition of transcription and/or DNA replication mechanisms. Methotrexate, a structural analogue of folate, is an antimetabolite. Antimetabolites block normal DNA synthesis by inhibiting several key enzymes. Methotrexate inhibits the enzyme dihydrofolate reductase. This inhibition interferes with the maintenance of the reduced folate pool. Reduced folates are essential for de novo synthesis of thymidylate and purine nucleotides, which is required for DNA synthesis and cell replication. Thus, by inhibiting dihydrofolate reductase, methotrexate stops the replication of rapidly proliferating cells such as tumor cells. Vinblastine is a vinca alkaloid. Vinca alkaloids interact with tubulin and disrupt microtubular function in the mitotic spindle. This action leads to metaphase arrest, resulting in mitotic arrest and cell death.

Clinical Performance. A multicenter randomized trial conducted by the British Medical Research Council Advanced Bladder Cancer Working Party compared modiÞed methotrexate and vinblastine (MV) with CMV in patients with advanced or metastatic TCC (Mead GM, 1998). The trial enrolled 214 patients between April 1991 and June 1995. A total of 108 patients were randomized to receive six cycles of CMV and 106 patients to receive six cycles of MV. Of the 88 evaluable patients in the CMV arm, 10% achieved CR and 36% achieved PR. Of the 93 evaluable patients in the MV arm, 7% achieved CR and 12% achieved PR. The hazard ratio (relative risk of dying) was 0.68 in favor of CMV. This Þnding translates to an absolute improvement in one-year survival of 13%: 16% in MV and 29% in CMV. The median survival was 7 months for CMV and 4.5 months for MV. CMV was associated with more toxicity than MV. In the CMV arm, Þve treatment-related deaths occurred, as did grade III leukopenia and thrombocytopenia. Neutropenic fever occurred in 11 CMV-treated patients and grade I renal toxicity in 19 CMV-treated patients, compared with only 2 and 4, respectively, among MV-treated patients. Investigators concluded that adding cisplatin

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to methotrexate and vinblastine should be considered in patients with TCC, taking into account the increased toxicity. The study strengthened the belief that cisplatin is an essential component of combination chemotherapy regimens for patients with adequate renal function. ´ Bacillus Calmette-Guerin Overview. Used for more than 20 years, bacillus Calmette-Gu´erin (BCG) (Organon’s Tice BCG/OncoTICE, SanoÞ-Aventis’s Theracys/ImmuCyst; Nippon Kayaku’s ImmuCyst, Japan BCG’s Immunobladder, generics) remains the most active agent for SBC therapy (Bassi P, 2002). BCG is approved for both intravesical use in the treatment and prophylaxis of primary and recurrent carcinoma in situ (CIS) of the urinary bladder and for prophylaxis following TUR of primary or recurrent stage Ta and/or T1 papillary tumors. Intravesical BCG is certainly the best available intravesical agent for the treatment of CIS (Bassi P, 2002; Baselli EC, 2001). Mechanism of Action. •

BCG is a potent stimulator of host defense mechanisms and can induce tumor regression in immunocompetent hosts. Its exact mechanism of action is unknown, though it appears that repeated instillation of BCG triggers a strong inßammatory reaction in the bladder, which correlates with an antitumor response.

Clinical Performance. A Southwest Oncology Group (SWOG) study compared BCG with 20 mg mitomycin C (Figure 8) using a six-week (followed by monthly) treatment schedule. However, the study was terminated at the Þrst interim analysis because of the highly signiÞcant advantage of BCG (Lamm DL, 1991). Overall recurrence fell from 32.6% with mitomycin C to 19.5% with BCG, while the median time to recurrence was prolonged from 20 months to more than 36 months. Another SWOG study evaluating maintenance therapy in 660 patients with rapidly recurring stage Ta or T1 lesions or CIS demonstrated a long-term diseasefree advantage for BCG maintenance therapy compared with BCG induction therapy alone over an average follow-up of 3.2 years (Lamm DL, 1995). Patients

FIGURE 8. Structure of mitomycin C.

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with CIS were treated with three additional weekly BCG treatments at three months, and their CR increased from 73% to 87%. In patients with stage Ta and T1 BC who were disease-free, BCG maintenance therapy (three weekly treatments of BCG at six-month intervals) resulted in a long-term disease-free status of 83%, compared with 50% in the induction group. An excellent 86% survival observed at four years in 391 randomized patients receiving induction therapy improved to 92% in patients receiving maintenance therapy. This result supports the superiority of continued maintenance treatment in patients with BC. Symptoms associated with BCG include urinary frequency and burning, mild malaise, and low-grade fever. The most dangerous complication is systemic sepsis and/or hypersensitivity reactions characterized by chills, fever, hypotension, and progressive multisystem organ failure (Lamm DL, 2000). Clinical trials frequently study the sequential administration of BCG and mitomycin C. A prospective controlled study presented at ASCO 2004 found that, in the treatment of high-risk BC, intravesical sequential BCG and electromotive delivery of mitomycin C are superior to BCG alone (Di Stasi SM, 2004). The trial randomized 175 patients with T1 BC into two arms following TUR and multiple biopsies. Arm 1 received six weekly instillations of BCG at 81 mg; arm 2 received sequential BCG and electromotive (intravesical electric current; 20 mA for 30 minutes) of mitomycin C at weekly intervals. Nonresponders received repeat courses at three months, and all complete responders underwent maintenance regimens of monthly instillations. The median follow-ups for arm 1 and arm 2 were 64 and 71 months, respectively. The recurrence rate was 47% for arm 1 and 28% for arm 2, and the rates of progression were 20% and 14%, respectively. The time to progression was 17 months for arm 1 and 46 months for arm 2, and the Þve-year mortality rates were 9.3 and 1.1, respectively. Researchers reported no treatment-related deaths, episodes of serious illness, or bladder contractures. These results demonstrate that intravesical sequential BCG/electromotive mitomycin C is superior to BCG alone in the treatment of high-risk disease. Researchers recently reported a meta-analysis of comparative studies in SBC that demonstrated a statistically signiÞcant superiority for BCG compared with single-agent mitomycin C in the prevention of tumor progression, but only if BCG maintenance therapy was provided (Bohle A, 2004). (See “Other Intravesical Agents” further on for details of this meta-analysis.) Additionally, the combination of BCG and interferon-alpha-2b (IFN-α-2b [Schering-Plough’s Intron A]) has been developed based on reported evidence of synergistic activity between the two agents. The two agents are said to be biocompatible and can be administered simultaneously. By combining IFN-α2b with a markedly reduced dose of BCG, researchers reduced toxicity while maintaining anticancer efÞcacy (Keane TE, 2001). This regimen is an effective alternative for patients at high risk of disease recurrence and/or progression and in whom BCG therapy has previously failed. A 56% disease-free survival was achieved at 24 months (O’Donnell MA, 2000). BCG and IFN-α-2b combinations are being studied in clinical trials.

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Interim results from a national multicenter Phase II trial of BCG in combination with IFN-α-2b in BCG-naive (BCG-N) and previous BCG failure (BCG-F) patients with SBC provides a benchmark for the efÞcacy and safety of the combination of BCG and IFN-α-2b (O’Donnell MA, 2004). Between May 1999 and May 2000, the trial enrolled 490 patients with primary or recurrent SBC, stage Ta, T1, CIS, and all grades. Approximately 50% of the patients had CIS or stage T1, and more than 75% had intermediate- to high-grade disease. The BCG-N group (259 patients) received a six-week induction course (six treatments provided once weekly) of standard-dose BCG plus 50 million units of IFN-α-2b followed by three-week maintenance cycles of reduced-dose BCG (1/3 to 1/10) plus 50 million units IFN-α-2b at 3, 9, and 15 months after induction. The BCGF group (231 patients) was treated similarly, except induction therapy began at a lower (1/3 to 1/10) BCG dose. Over a median follow-up of 24 months, the simple tumor recurrence rates for the BCG-N and BCG-F groups were 40% and 52%, respectively, and disease-free rates were 57% and 42%, respectively. Progression to muscle invasion occurred in 5% and 4.3%, respectively, while metastasis occurred in 2.3% and 2.6%, respectively. About 3.9% of patients in each group underwent cystectomy, and two patients died of BC in each group. Serious adverse events occurred in 5.5% of the patients; infection-related serious adverse events were less prevalent in the BCG-F group (2.6%) than in the BCG-N group (5.4%). Moderate-to-severe local side effects during induction were higher in the BCG-F group (6.2%) than in the BCG-N group (16.9%) but equilibrated during maintenance therapy. The study could not determine the incremental value of IFN. Therefore, randomized Phase III trials need to establish if there is any beneÞt to adding IFN to BCG. Other Intravesical Agents Overview. Mitomycin (generics) is the most common chemotherapeutic agent for patients with low- or intermediate-risk SBC. Studies comparing BCG with mitomycin have been controversial: mitomycin has proved equivalent or superior to BCG for reducing recurrences in patients with papillary disease in some studies and has shown efÞcacy in treating CIS and higher-grade lesions in others (Dalbagni G, 2000). Postoperative instillations of mitomycin C can reduce recurrence rates in newly diagnosed patients with SBC. Physicians generally administer adjuvant mitomycin to patients with low- or intermediate-risk SBC and adjuvant intravesical BCG to patients with high-risk disease. Mitomycin is used in patients with SBC who are unable to tolerate BCG therapy because of toxicity or whose disease is refractory to BCG (Baselli EC, 2001). In this section, we also brießy discuss three other intravesical agents used to treat BC: doxorubicin (Figure 7), epirubicin (Figure 9), and thiotepa. None has demonstrated efÞcacy superior to that of BCG. Mechanism of Action. •

Mitomycin and thiotepa are alkylating agents. Alkylating agents cross-link with DNA during cell reproduction, thereby preventing synthesis of secondgeneration DNA. Mitomycin is nonspeciÞc to the cell-cycle phase.

CURRENT THERAPIES

O

OH

173

O OH OH

H3CO

O H3C HO

OH

O

O NH2

FIGURE 9. Structure of epirubicin. •

Doxorubicin and epirubicin are anthracyclines. Anthracyclines interact with several different cellular targets, most importantly topoisomerase II. By inhibiting this DNA religation enzyme, anthracyclines exert their cytotoxic effect. Another mechanism that leads to cell death is known as DNA intercalation, in which the anthracycline molecule inserts itself between base pairs. This phenomenon causes single- and double-stranded breaks in DNA that inhibit cell proliferation. The free radicals generated by the reductive metabolism of anthracyclines may also damage cellular structures such as proteins and DNA.

Clinical Performance. Researchers recently reported a meta-analysis of comparative studies on tumor progression in SBC (Bohle A, 2004). Tumor progression was deÞned as progression to a higher tumor stage or the development of metastatic disease. The main objective was to compare the therapeutic efÞcacy of adjuvant intravesical BCG with adjuvant intravesical mitomycin C on progression of stage Ta and T1 BC (all grades). In nine clinical trials, 1,277 patients were treated with BCG and 1,133 with mitomycin C. With an overall median follow-up of 26 months, 7.67% of patients in the BCG group and 9.44% of the patients in the mitomycin C group developed tumor progression. Of the nine studies analyzed, the combined result of the Þve studies on BCG maintenance showed a statistically signiÞcant superiority of BCG over mitomycin C. In the remaining four studies without BCG maintenance, the combined result indicated no statistically signiÞcant difference between the two treatments. This meta-analysis demonstrates that adjuvant intravesical BCG is statistically superior to adjuvant intravesical mitomycin C for the prevention of tumor progression, but only if BCG maintenance therapy is provided. Another intravesical agent used to treat BC is doxorubicin (PÞzer’s Adriamycin/Adriblastine, Kyowa’s Adriacin, generics), though evidence suggests that its efÞcacy is inferior to BCG’s regardless of tumor grade or stage (Baselli EC, 2000). Furthermore, treatment with doxorubicin causes serious toxicity from chemotherapy-induced cystitis, resulting in permanent bladder contracture in a small but signiÞcant and unpredictable number of patients (Lamm DL, 1988). Epirubicin (PÞzer’s Ellence/Farmorubicin, PÞzer/Kyowa’s Farmorubicin) is approved for adjuvant treatment (in combination regimens) of patients with early

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node-positive breast cancer and has demonstrated an ability to alter the natural history of primary SBC. Administration of a single dose (100 mg in 100 mL of saline) of epirubicin immediately following TUR in patients with primary SBC produced a statistically signiÞcant recurrence-free rate of 66% after two years (Rajala P, 1999). Its adverse effects are usually limited to chemical cystitis. However, long-term reduction of tumor recurrence (greater than or equal to Þve years) has been difÞcult to conÞrm (Lamm DL, 2000). Another chemotherapeutic agent used to manage SBC is thiotepa (Amgen/Wyeth Lederle’s Thioplex, Sumitomo’s Tespamin, generics), but few clinicians recommend it as a Þrst-line alternative to BCG despite its approval for the treatment of superÞcial papillary carcinoma of the urinary bladder (Baselli EC, 2000). More important, direct randomized comparison has demonstrated that BCG prophylaxis is superior to thiotepa, doxorubicin, or mitomycin C in patients with SBC (Lamm DL, 2000). Nonpharmacological Approaches Surgery is the mainstay of treatment for SBC. In particular, TUR of all visible tumors remains the ultimate method for diagnosing and initially managing SBC (Baselli EC, 2001). Tumor specimens taken during TUR are analyzed histologically to determine their pathological grade, and this procedure is essential for determining the clinical stage of the bladder tumor. Patients with SBC, many of whom are at risk for disease recurrence and progression, can be managed with adjuvant intravesical therapy following TUR. If recurrence develops or pathologic assessment places the patient at a higher risk for recurrence and/or progression to MIBC, then adjuvant intravesical maintenance therapy is warranted (Baselli EC, 2001). As we have discussed, studies show that mitomycin and other adjuvant intravesical chemotherapy regimens are inferior to BCG, especially in patients at high risk of tumor recurrence. Cystectomy should be considered in patients who fail to have their disease eradicated following a reasonable course of intravesical treatment or at the Þrst sign of tumor progression (Bochner BH, 2001). Radical cystectomy is the standard treatment for patients with MIBC in the United States and Europe (Rodel C, 2002). Studies have investigated the longterm effect of radical cystectomy in the treatment of invasive BC. A study involving 1,054 patients with TTC of the bladder uniformly treated with radical cystectomy with bilateral pelvic iliac lymphadenectomy and urinary diversion (with or without adjuvant radiation or chemotherapy) supports the aggressive surgical management of invasive BC (Stein JP, 2001). Over a median followup of 10.2 years, results demonstrated that excellent long-term survival can be achieved with a low incidence of pelvic recurrence (9%). The chance of survival at Þve years without evidence of disease was 69%, and overall survival at Þve years was 60%. In addition, investigators note that quality of life after cystectomy has improved because lower urinary tract options have evolved into an orthotopic form of diversion, thus allowing most patients to store urine without the need for catheterization.

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Radiotherapy may be used to either cure locally advanced BC or provide palliative treatment (Nilsson S, 2001). A new, promising treatment to preserve the bladder is extensive TUR followed by intense chemoradiotherapy. Researchers have found that this method offers a Þve-year survival rate similar to that of radical cystectomy (Nilsson S, 2001). Researchers have also found that radiochemotherapy is more efÞcacious than radiotherapy. In a study involving 428 patients with MIBC (T2–T4) or high-risk T1 BC, 126 patients were treated with radiotherapy and 302 with radiochemotherapy after TUR of the tumor (Rodel C, 2002). Over a median follow-up of 60 months, the researchers found radiochemotherapy to be more effective than radiotherapy in terms of CR and survival. The ideal treatment for MIBC would be therapies that preserve the bladder and eradicate the tumor without compromising survival. EMERGING THERAPIES Little progress has been made over the past decade in the drug treatment of bladder cancer (BC), primarily because recruiting sufÞcient numbers of patients to Phase III BC trials is notoriously difÞcult. There are several reasons for this difÞculty. First, the incidence of BC is relatively low compared with that of other cancers, such as non-small-cell lung cancer and colorectal cancer. Second, because the average age of the patient population is 70 years, many patients are not Þt enough to meet trial entry criteria. Third, many pharmaceutical companies believe that the cost of conducting trials outweighs the potential beneÞts of assessing a drug for a market such as BC, which is small compared with those for breast cancer, non-small-cell lung cancer, and prostate cancer. Consequently, signiÞcant clinical data are lacking for most agents in clinical development for the treatment of BC. Table 6 summarizes the drug therapies in development for BC. Unless indicated, our deÞnition of BC is largely interchangeable with transitional cell cancer (TCC) of the urothelium. BC and TCC are not pathologically the same, but we grouped all forms of BC together when evaluating clinical data because TCC represents more than 90% of all BCs, and because other pathological types of BC are treated similarly (staging and grading being more important determinants of treatment and patient stratiÞcations in clinical trials). Epidermal Growth Factor Receptor Inhibitors Overview. Epidermal growth factor receptor (EGFR) is a transmembrane receptor tyrosine kinase expressed in a variety of human malignancies, including BC. The EGFR, also known as c-erbB-1, belongs to the EGFR subfamily of receptor tyrosine kinases, which includes c-erbB-2/HER-2/neu. The natural ligands for EGFR, tumor growth factor–alpha and EGF, have both been immunohistochemically demonstrated to be present in BC (Mellon JK, 1996). The expression of EGFR has been studied extensively in human BC tissue in which a broad range (0–100%) of expression has been observed (Small EJ, 2003). Part of the variation may be explained by the different EGFR antibodies used, the criteria employed to deÞne overexpression, and the stage of BC studied.

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TABLE 6. Emerging Therapies in Development for Bladder Cancer Compound Lapatinib United States Europe Japan Gefitinib United States Europe Japan Trastuzumab United States Europe Japan Erlotinib (Tarceva) United States Europe Japan Paclitaxel United States Europe Japan Vinflunine (Javlor) United States Europe Japan Irinotecan (Camtosar) United States Europe Japan Celecoxib United States Europe Japan

Development Phase

Marketing Company

— II —

— GlaxoSmithKline —

II — —

AstraZeneca — —

II — —

Genentech/Roche/Chugai — —

II — —

Genentech/Roche/OSI Pharmaceuticals — —

IIb IIb —

Bristol-Myers Squibb Bristol-Myers Squibb —

— III —

— Pierre Fabre —

II — —

Pfizer — —

III — —

Pfizer — —

Apoptosis stimulators Mycobacterial cell-wall complex (Regressin) United States III Europe — Japan — Arsenic trioxide (Trisonex) United States II Europe — Japan — Tipifarnib (Zarnestra) United States II Europe — Japan —

Bioniche — — Cell Therapeutics — — Janssen Pharmaceutica/Johnson & Johnson — —

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TABLE 6. (continued) Compound

Development Phase

Marketing Company

Antifolates Pemetrexed (Alimta) United States Europe Japan Immunotherapies IDM-2 (Bexidem) United States Europe Japan

II — —

Eli Lilly — —

— II/III —

— Immuno-Designed Molecules —

Alkylating agents Apaziquone (EOquin) United States Europe Japan

— II —

— Spectrum Pharmaceuticals —

Expression of EGFR at any level was seen in the majority of invasive (T2, T3, or T4) BCs and averaged 64% across 250 patients from select studies for which tumor-stage subset data were available (Small EJ, 2003). The frequency of EGFR expression in metastatic sites has not been studied extensively, although in one study, 65% of metastatic samples expressed EGFR, similar to what was seen in the corresponding primary tumor (Bue P, 1998). A separate study indicated that the frequency of EGFR expression was 78% in 33 patients with T3 or T4 tumors (Neal DE, 1990). A signiÞcant positive correlation has been noted between the primary tumor stage (T3 or T4 versus T1 or T2) and the proportion of metastatic tumors expressing EGFR. EGFR-positive BCs are signiÞcantly more likely to recur after resection, to progress to muscle-invasive disease (MIBC), and to cause death. Therefore, EGFR expression is a prognostic factor in BC. A vast amount of research and development (R&D) is ongoing in the area of EGFR inhibition, signifying its commercial viability. Two main approaches are being investigated for therapeutic intervention in BC: the speciÞc inhibition of EGFR tyrosine kinase and monoclonal antibodies (MAbs) directed at the external domain of the EGFR. No randomized data exist demonstrating the efÞcacy of EGFR inhibitors in the treatment of BC. Nevertheless, the extensive R&D in this area as a whole and the rationale for developing these agents to treat BC warrants their inclusion. Mechanism of Action. The binding of EGF to the receptor activates the receptor’s intracellular enzymatic portion and, in turn, triggers a cascade of intracellular events that stimulate growth and survival of the tumor cell. Researchers believe that overexpression of EGFR and overactivity of its downstream signal transduction pathway are signiÞcant factors in the growth of tumors in a subset of

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BC patients. Likewise, the HER-2 gene is overexpressed in a variety of cancers, including BC, and can lead to dysregulated cell growth. In addition to its effects on cell proliferation and the cell cycle, EGFR activation may inßuence several cellular processes, including angiogenesis, apoptosis, motility, adhesion, invasion, and metastasis. Lapatinib. GlaxoSmithKline’s lapatinib is an orally administered EGFR tyrosine kinase inhibitor that has the added beneÞt of blocking ErbB-2/HER2 tyrosine kinase. The compound is in Phase II trials for BC in Europe. Preliminary Phase II data were presented at the 2004 European Society for Medical Oncology meeting (WulÞng C, 2004). The trial evaluated single-agent lapatinib as a second-line treatment for patients with advanced BC (ABC, locally advanced or metastatic, incurable by surgery alone) after Þrst-line treatment with a platinum-based regimen (63%) or other treatments (37%). Of the 58 patients enrolled, only 30 patients were evaluable for the preliminary analysis; 3 patients (10%) had a partial response (PR), but only one PR was sustainable for eight weeks or longer. Eight patients (27%) had stable disease (SD). A clinical beneÞt (SD for six months or more) was observed in three patients. Survival data are premature. Six patients experienced serious adverse events (grading not given), of which one was determined to be treatment-related. The rationale also exists for combining two EGFR inhibitors that target two speciÞc EGFRs, but no signiÞcant clinical data on this approach have emerged. Gefitinib. GeÞtinib (AstraZeneca’s Iressa) is an orally active, quinazolinederived selective EGFR tyrosine kinase inhibitor that blocks signal transduction. GeÞtinib launched in the United States and Japan as a monotherapy for the treatment of non-small-cell lung cancer after patients fail on both platinum-based and docetaxel chemotherapies. The compound is also undergoing numerous other clinical trials in various other cancer indications, including BC. No signiÞcant clinical data exist for geÞtinib in BC. However, numerous preclinical data and preliminary Phase II data provide the rationale for further development and commercial viability. GeÞtinib’s dose-dependent growth-inhibitory and apoptotic effects on several human BC cell lines correlate with the degree of expression of EGFR (Meye A, 2001). Preclinical studies in athymic mice indicate potentiation of cytotoxic agents by geÞtinib in human tumor xenografts derived from various cancer cell lines (Sirotnak FM, 2000). In this system, geÞtinib alone had growth-retarding effects, which signiÞcantly enhanced the antitumor activity of platinum compounds, taxanes, and other cytotoxic agents. It is noteworthy that geÞtinib’s tumor growth-inhibitory activity in combination with chemotherapy is evident even without high levels of EGFR expression. A Phase I trial of geÞtinib (250 or 500 mg) in combination with 1,250 mg/m2 gemcitabine (Eli Lilly’s Gemzar)/cisplatin (generics) has been completed (Small EJ, 2003). No signiÞcant increase in toxicity attributable to chemotherapy was apparent. Furthermore, large variations in the pharmacokinetics of gemcitabine were not observed at the two dose levels of geÞtinib, suggesting that a clinically signiÞcant pharmacokinetic interaction does not occur.

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Because a Þxed-dose-rate infusion of gemcitabine appears to be more effective than standard dosing for the treatment of pancreatic cancer, the Cancer and Leukemia Group B (CALGB) initiated a Phase II study, called CALGB 90102, to test the efÞcacy of cisplatin, Þxed-dose-rate gemcitabine, and geÞtinib in the treatment of patients with BC that had metastasized to lymph nodes or other sites. A total of 27 patients with N2, N3, or M1 disease (performance status 0–2) were enrolled before the study was halted because the dose-limiting toxicity (DLT) exceeded preestablished stopping rules. Preliminary data were presented at the 2004 American Society of Clinical Oncology (ASCO) meeting (Philips G, 2004). The DLT events observed were two grade V (one neutropenic infection, one cerebral vascular accident) and three grade IV nonhematologic toxicities (one hyperuricemia, one fatigue, one dyspnea). Researchers also observed grade 3 diarrhea and skin rash in two (8%) and four (16%) patients, respectively. Of 24 evaluable patients, 12 responded to treatment, representing an overall response rate of 50%. Median time to progression was 6.9 months. This regimen was associated with excessive toxicity, possibly because of the Þxed-dose-rate infusion of gemcitabine, so the researchers now use a standard gemcitabine dosing schedule. CALGB has also initiated a trial for patients with metastatic BC who have renal insufÞciency and therefore cannot tolerate cisplatin. This trial is using carboplatin instead of cisplatin, in combination with gemcitabine and geÞtinib, because carboplatin is better tolerated in patients with renal insufÞciency. Trastuzumab. Trastuzumab (Genentech/Roche/Chugai’s Herceptin) is a recombinant humanized MAb that binds to the HER2 receptor. In addition to binding the extracellular domain of the receptor, trastuzumab may also induce antibody-dependent cytotoxicity against target tumors. Single-agent trastuzumab is indicated for the treatment of patients with metastatic breast cancer (CaB) whose tumors overexpress HER2 and who have received prior chemotherapy for their disease. Trastuzumab is undergoing Phase II trials in the United States for BC. The HER-2 proto-oncogene encodes a transmembrane receptor tyrosine kinase with substantial homology to the other members of the class I receptor tyrosine kinase family, namely EGFR, HER3, and HER4. The degree of HER2 receptor expression in tumors is represented using a scale of 0 (low or negative) to 3 (strongly positive). High expression of the HER2 receptor is demonstrated in 25–30% of CaB patients, making them eligible for treatment with trastuzumab. High HER2 expression increases the proliferative rate of tumor cells, resulting in poor prognosis. Studies indicate that HER2 expression rates in BC specimens range from 2% to 74% (Small EJ, 2003); part of the variation may be explained by the different HER2 antibodies used, the criteria employed to deÞne overexpression, and the stage of BC studied. Both overexpression without gene ampliÞcation and heterogeneity in HER2 expression are more common in BC than in CaB (Underwood M, 1995; Lee SE, 1994). A recent analysis of a well-deÞned set of MIBC

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specimens found a 28% incidence of HER2 positivity by immunohistochemistry, whereas the incidence of 3+ (high HER2 expression) tumors in the primary specimens was 18% (Jimenez RE, 2001). This study also demonstrated that HER2 overexpression in the primary tumors consistently predicted its overexpression in regional or distant metastatic sites. Conversely, some HER2 -negative primary tumors also expressed HER2 in their corresponding metastases. A higher percentage of HER2 positivity was found in lymph-node metastases (53% were 2+/3+, 31% were 3+) when compared with the primary sites. These data provide rationale for developing trastuzumab for the treatment of BC. At the time of composing this reference, no clinical data exist for trastuzumab in treating BC, but the CALGB has opened a trial of trastuzumab for the treatment of BC. The Phase II study will evaluate trastuzumab, paclitaxel, carboplatin, and gemcitabine in patients with locally recurrent or metastatic urothelial carcinoma overexpressing HER2. Erlotinib. Erlotinib (Tarceva) is under development by OSI Pharmaceuticals in alliance with Genentech and Roche. This orally active quinazoline derivative is in Phase II U.S. trials for the prevention of BC recurrence after surgery. Like geÞtinib, erlotinib is an oral, once-daily small molecule that inhibits the intracellular tyrosine kinase domain of the EGFR, thereby blocking receptor activity. This compound’s lead indication is non-small-cell lung cancer, for which it is awaiting U.S. and European approval. No clinical data exist on erlotinib in BC. A Phase II randomized study of adjuvant erlotinib and a green tea extract (polyphenon E) for preventing recurrence and progression of BC in former smokers with resected high-grade superÞcial BC (SBC) is ongoing. At the time of composing this reference, the clinical trial with green tea extract is ongoing. Patients are stratiÞed according to disease stage (Ta versus T1 versus carcinoma in situ [CIS]) and randomized to one of three treatment arms: arm I patients receive erlotinib and placebo once daily; arm II patients receive oral green tea extract and placebo once daily; and arm III patients receive placebo. In all arms, treatment continues for nine months in the absence of disease recurrence or unacceptable toxicity. Patients are followed every three months for 15 months and then every six months for three years. A total of 330 patients (110 per treatment arm) have been expected to be enrolled for this study within three years. Microtubule-Targeting Agents Overview. Microtubules—major structural components in cells—are the target of a large and diverse group of anticancer drugs, including taxanes, epothilones, and vinca alkaloids. Taxanes are potent antitumor agents; members of this class are marketed for numerous forms of cancer. As single agents and combined with other chemotherapeutic agents, taxanes have shown activity against a range of solid tumors. The most promising taxane in development for the treatment of BC is paclitaxel (Bristol-Myers Squibb’s Taxol, generics). In fact, it is already used off-label

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based on results of multiple clinical trials showing promising efÞcacy. In a Phase II clinical trial, Sonus is investigating an emulsion formulation of paclitaxel that incorporates the company’s Tocosol drug delivery technology. In a related development, Bristol-Myers Squibb is investigating the epothilone ixabepilone in early clinical trials for BC. Ixabepilone has anticancer effects similar to those of taxanes. Although clinical data for BC are not yet available for ixabepilone or for paclitaxel delivered via Tocosol, we do not completely exclude them from the list of potential BC agents. Mechanism of Action. Microtubules are protein polymers that are responsible for various aspects of cellular shape and movement. The major component of microtubules is the protein polymer tubulin. Taxanes and epothilones promote microtubule polymerization and inhibit tubulin depolymerization, arresting mitotic cell division at the metaphase/anaphase transition and inducing cell death. Vinca alkaloids inhibit microtubule polymerization at high drug concentrations. Importantly, considerable evidence indicates that, at lower concentrations, taxanes and epothilones have a common mechanism of action: they suppress the dynamics of microtubules without appreciably changing the mass of microtubules in the cell. The drugs bind to diverse sites on tubulin and at different positions within the microtubule and exert a variety of effects on microtubule dynamics. However, by their common mechanism of suppressing microtubule dynamics, they all block mitosis at the metaphase/anaphase transition and induce cell death. Paclitaxel. Paclitaxel is used commonly to treat other cancers. For BC, paclitaxel has reached Phase III clinical evaluation in the United States and Europe. Phase III data comparing carboplatin/paclitaxel (CP) with methotrexate/ vinblastine/doxorubicin/cisplatin (MVAC) in ABC patients are not encouraging (Dreicer R, 2004). Eighty-Þve patients were randomized to the respective treatment regimens (41 to CP and 44 to MVAC). Response rates and overall survival were similar in both treatment arms. Patients treated with CP had an overall response rate of 28.2%, compared with 35.9% for the MVAC arm. Median progression-free survival among MVAC-treated patients was 8.7 months, compared with 5.2 months for CP-treated patients. At a median follow-up of 32.5 months, the median survival for patients treated with MVAC was 15.4 months versus 13.8 months for patients treated with CP. Patients treated with MVAC had more severe worst-degree toxicities than did patients treated with CP. No signiÞcant differences with regard to quality of life, as assessed by the Functional Assessment of Cancer Therapy–Bladder (FACT-BL) instrument, were observed. An international collaborative group that includes the European Organization for Research and Treatment of Cancer (EORTC) and the Southwest Oncology Group (SWOG) is conducting a randomized, multicenter Phase III trial to compare GC (gemcitabine [Eli Lilly’s Gemzar)]/cisplatin [generics]) with the paclitaxel/cisplatin/gemcitabine (TCG) regimen in previously untreated ABC patients. The trial, with a sample size of 630 patients, was closed in mid 2004.

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Results are still anticipated. A U.S. trial is evaluating adjuvant doxorubicin and gemcitabine followed by paclitaxel and cisplatin versus adjuvant cisplatin and gemcitabine in patients with completely resected locally advanced BC. Clinical trials evaluating paclitaxel combined with cisplatin or carboplatin and/or gemcitabine have yielded some promising results. A Phase II trial provided evidence for the therapeutic beneÞt of paclitaxel and carboplatin as a Þrst-line treatment of ABC (Redman BG, 1998). Patients (performance status of 2 or less) received paclitaxel 200 mg/m2 followed by carboplatin (area under the curve [AUC] 5) every 21 days. Thirty-Þve patients were assessable for response. Researchers administered a total of 184 cycles (median, six cycles per patient). They observed 7 complete responses (CRs) and 11 PRs, giving an overall response rate of 51.5%. Median response durations for CR and PR were six and four months, respectively. Overall median survival was 9.5 months. Nine patients required one dose reduction, and seven patients required two dose reductions; only one episode of febrile neutropenia and sepsis occurred. Myalgias and arthralgias of grades I/II occurred in 16 patients and usually lasted two to three days after treatment. No treatment delays because of toxicity were noted. Researchers have also evaluated the triplet combination of carboplatin, paclitaxel, and gemcitabine in patients with ABC (Hussain M, 2001). Patients (performance status 2 or less) received the same 21-day regimen as in the previously discussed trial but with the addition of gemcitabine (800 mg/m2 ) on days 1 and 8. Forty-nine patients (44 men and 5 women) were enrolled, 47 of whom were assessable for response. A total of 272 cycles were administered (median, six cycles). Fifteen (32%) patients experienced a CR and 17 (36%) patients experienced a PR, for an overall response rate of 68%. Responses were seen in all sites, including 15 (68%) of 22 patients with visceral metastases. The median survival was 14.7 months, with a one-year survival rate of 59%. Major toxicities were grade III and IV neutropenia in 17 and 19 patients, respectively; grade III and IV thrombocytopenia in 15 and 6 patients, respectively; grade III and IV anemia in 10 and 2 patients, respectively; grade III neuropathy in 4 patients; and diarrhea in 2 patients. The incidence of febrile neutropenia was 1.4%; no patients died of drug toxicity. Paclitaxel and cisplatin have been evaluated in the treatment of metastatic BC (Burch PA, 1999). Cycles (135 mg/m2 paclitaxel; 70 mg/m2 cisplatin) were repeated every 21 days until progression or a maximum of six cycles. Twentynine patients (performance status 2 or less; 26 males and 3 females) with untreated metastatic BC participated in the study. Researchers observed an overall response rate of 72% and a CR rate of 34%. Eighteen patients progressed; median time to progression was eight months. When these data were released, 14 patients were still alive, and their median survival was 13 months. Grade III/IV toxicities included anorexia (two patients), nausea (Þve patients), vomiting (four patients), and fatigue (one patient). Six patients experienced grade II or higher neurotoxicity. Paclitaxel and cisplatin have also been evaluated as neoadjuvant therapy in locally advanced BC to determine whether those who experience a CR can avoid

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radical cystectomy. Some encouraging Phase II data are available (Mel JR, 2001). In this trial, patients received paclitaxel (175 mg/m2 ) and cisplatin (75 mg/m2 ) every 21 days for three cycles. Patients were then re-staged by computed tomography, cystoscopy, and multiple biopsies. Patients with a PR, stable disease (SD), or progressive disease (PD) underwent radical cystectomy. Patients with a CR underwent radiation therapy to avoid radical cystectomy. Forty-four patients (40 males, 4 females; performance status 2 or less) were evaluable for response and toxicity evaluations; of this group, 20.5% were stage II and 79.5% were stage III. Researchers observed 24 (54.5%) CRs, 11 PRs (25%), 6 SDs (11.4%), and 4 PDs (9.1%). After a median follow-up of 15.5 months (median response duration was 14 months), four patients had local relapses and four patients had distant relapses. Neutropenia was the main hematologic toxicity (grade III/IV neutropenia occurred in 31.8% of patients). No toxic deaths and no febrile neutropenia occurred. Other grade III/IV toxicities included nausea and vomiting, which occurred in 9.1%. Toxicity did not cause any treatment delays. Based on the aforementioned trials, paclitaxel is likely to play an important role in BC treatment, especially among patients with recurrent or nonresponding disease. Vinflunine. Vinßunine (Pierre Fabre’s Javlor) (Figure 10), a semisynthetic vinca alkaloid derived from vinorelbine, is in Phase III clinical trials for the treatment of BC in Europe and other, non-major-market territories. The Phase III study will enroll 330 patients who have either not responded to treatment with a platinumbased regimen or relapsed afterward. These patients will be drawn from 90 centers in 18 countries, including Mexico, Argentina, South Africa, and most countries in Europe. The main criterion for evaluation is overall survival. Results of a Phase II study of vinßunine in patients with ABC who had progressed after at least one line of platinum-containing therapy were presented at the 2003 ASCO meeting (Bui B, 2003). Among 48 evaluable patients, researchers observed 9 PRs (19%), 23 SDs (48%), and 16 PDs. Grade III/IV neutropenia, leukopenia, anemia, thrombocytopenia, constipation, myalgia, and vomiting occurred in 67%, 45%, 14%, 6%, 12%, 4%, and 6% of patients, respectively.

N

CF2CH3

N N H CH3O2C CH3O

CH3 N

OCOCH3 HO CO2CH3 CH3

FIGURE 10. Structure of vinflunine.

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Although vinßunine has shown some promising results in the aforementioned Phase II trial, Phase III data are needed. No effective treatment in the second-line setting exists for BC, particularly for patients whose disease is refractory to or who cannot tolerate gemcitabine-based therapy, so any therapy that could demonstrate even a marginal beneÞt over best supportive care would be a welcome option. Topoisomerase I Inhibitors Overview. Cytotoxic agents targeting topoisomerase I are commonly used to treat cancer. Camptothecin (CPT, generics; an alkyloid agent derived from the Campotheca acuminate tree) (Figure 11) was the Þrst topoisomerase I inhibitor to enter trials for the management of cancer. However, unacceptable toxicity, including severe myelosuppression, enteritis, and hemorrhagic cystitis, led to the discontinuation of initial trials. A new generation of topoisomerase I inhibitors has emerged, including irinotecan (PÞzer’s Camptosar, Yakult/SanoÞ-Aventis’s Campto, Daiichi’s Topotecin) (Figure 12), discussed further on. Mechanism of Action. Topoisomerase I catalyzes the conversion of DNA strands into single-strand DNA breaks—a process necessary to relieve chain

FIGURE 11. Structure of camptothecin (R1 = R2 = R3 = H).

H3C O

O

O N

N N O N

HO H3C

FIGURE 12. Structure of irinotecan.

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tension during the DNA replication required for cell division—by nicking and then reannealing the DNA strand. Camptothecin analogues exert their cytotoxic effect by binding to topoisomerase I and stabilizing the intermediate enzymeDNA cleavable complex, generating DNA strand breaks that are lethal to the cell. Irinotecan. Irinotecan (PÞzer’s Camptosar, Yakult/SanoÞ-Aventis’s Campto, Daiichi’s Topotecin) (Figure 12) is a parenteral topoisomerase I inhibitor approved for the treatment of metastatic colorectal cancer. It is in development for a number of cancers, including BC. At the time of composing this reference, no data on irinotecan for the treatment of BC are available. Two Phase II trials are under way in the United States. One trial is evaluating single-agent irinotecan in the second-line setting; the other is evaluating the combination of irinotecan and gemcitabine in ABC. Given irinotecan’s demonstrated clinical activity in colorectal cancer and many other solid tumors, it has potential in the Þeld of BC as well. Cyclooxygenase-2 Inhibitors Overview. Nonsteroidal anti-inßammatory drugs (NSAIDS) that inhibit cyclooxygenase-1 and -2 (COX-1 and -2) are active in preventing cancers (Sabichi AL, 2004). However, the use of nonspeciÞc NSAIDs such as aspirin (generics) in cancer prevention and treatment is limited because of the risk of upper gastrointestinal (GI) side effects. The recent disclosure that a COX-2 inhibitor increased the risk of heart attacks and strokes will make clinicians less likely to prescribe agents in this class for a relatively curable cancer. Mechanism of Action. The mechanism of anticancer activity of COX-2 inhibitors is not fully understood, although it may involve anti-inßammatory effects. COX-2 is expressed strongly in invasive BC. The degree of COX-2 expression correlates with tumor grade and stage (Yoshimura R, 2001). COX-2 immunostaining is positive in 31–80% of invasive bladder cancers (Shirahama T, 2000). Thus, high levels of COX-2 may promote tumorigenesis through enhanced angiogenesis, increased tumor invasiveness, resistance to apoptosis, and reduced host-immune surveillance. Celecoxib. Celecoxib (PÞzer’s Celebrex) (Figure 13) is already approved in the United States and is awaiting European approval as an adjuvant to surgery in treating familial adenomatous polyposis (FAP), a precursor to colorectal cancer. No randomized data on the role of celecoxib in BC treatment or prevention exist. However, the results of a study using data from a Canadian database containing prescription and physician service claims were disclosed at the 2003 ASCO meeting (Sheehy OE, 2003). Study patients were newly diagnosed with SBC and had surgical endoscopic treatment for bladder malignancies or cystectomy. Patients must have had one follow-up visit for diagnostic and therapeutic urethro-cystoscopy after the initial procedure. Eligible patients were grouped into

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O

H2N S O

N

N CF3

H3C

FIGURE 13. Structure of celecoxib (R1 = NH2 , R2 = CH3 ).

cohorts of celecoxib, NSAIDs, acetaminophen, and non-users. Researchers compared SBC recurrence (as signiÞed by receipt of another surgical procedure to remove bladder tumor) among study cohorts. Of the 1,172 patients evaluable, 55 received celecoxib, 382 received NSAIDs, 208 received acetaminophen, and 527 received nothing. Survival analysis showed cumulative recurrence rates at one year of 61.5% among non-users and 56.7% in the acetaminophen group, but approximately 30% for the celecoxib and NSAID groups. After adjusting the aforementioned variables, the COX-regression model indicates that celecoxib and NSAID users had a signiÞcantly lower risk of SBC than non-users. A Phase III U.S. trial examining celecoxib in the chemopreventive setting is under way. The trial will compare the time to recurrence after treatment with celecoxib versus placebo in patients with SBC after resection who have a high risk for recurrence (i.e., stage Ta [grade 3 or multifocal or at least two occurrences, including current tumor, within the past 12 months] or stage T1 or CIS [any grade]). A total of 152 patients will be recruited. As mentioned above, but needs to be repeated, the recent disclosure that a COX-2 inhibitor increased the risk of heart attacks and strokes will make clinicians less likely to prescribe agents in this class for a relatively curable cancer. Apoptosis Stimulators Overview. Apoptosis, or programmed cell death, represents a universal and exquisitely efÞcient cellular suicide pathway. As the understanding of its vital role in normal development has deepened, researchers have identiÞed numerous genes that encode apoptotic regulators, some of which represent familiar oncogenes or tumor-suppressor genes. Targeting apoptosis is successful only if the therapeutic index is good enough to selectively destroy cancer cells rather than normal cells. In drug-curable malignancies, such as common pediatric leukemias and certain solid tumors, apoptosis is a prominent mechanism associated with the induction of tumor remission. Many cytotoxic agents’ ultimate mechanism of action works via the general apoptotic pathway; we introduce this category here to cover the agents described further on because it is the most appropriate class in which to group them.

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Mechanism of Action. Apoptotic cell death is triggered by intracellular cues such as DNA damage and osmotic stress, and by extracellular cues such as growth factor withdrawal, matrix detachment, and direct cytokine-mediated killing. Two central pathways are involved in the process of apoptotic cell death: one activates the caspase proteases and the other involves a mitochondrial pathway. A drug that activates apoptosis might achieve a suitable therapeutic index in several ways. First, it might activate a death cascade via a drug target that is uniquely expressed in a cancer cell. Alternatively, it might be delivered to the target tissue in a manner that is selective for the cancer cell. A third possibility—perhaps the most promising one—is to exploit a pathway that is activated by oncogenes in order to provoke apoptosis selectively in cancer cells. It is now clear that oncoproteins can interact with apoptotic regulatory pathways. Thus, overexpression of myc sensitizes cells to a wide assortment of apoptotic triggers, probably reßecting the role of apoptosis in the intracellular immunity that prevents normal cells from persisting in the body once they acquire cancer-causing genetic defects. However, many human tumors that overexpress myc are highly resistant to apoptotic triggers, probably owing to a variety of downstream lesions that blunt the death pathway. Still, the recognition that oncogenes can sensitize cells to proapoptotic treatments suggests that if such lesions can be circumvented, drugs that induce cell death could prove highly selective for cancer cells. Mycobacterial Cell-Wall Complex. Mycobacterial cell-wall complex (MCC) (Bioniche’s Regressin, Urocidin) is a DNA oligonucleotide complex that actively inhibits tumor cell growth and triggers apoptosis. Its lead indication is BC, for which Phase II trials have been completed in the United States. In February 2006, the protocol was under review by the FDA, which had cleared the company to start a smaller open-label phase III bladder cancer trial. Enrollment is planned to begin in the second quarter of 2006. Final Phase I/II data evaluating MCC in SBC were presented at the 2004 American Urological Association meeting (Filion MC, 2004). The trial involved 55 patients with CIS refractory to traditional treatments. They received MCC (4 or 8 mg) and were evaluated at 3, 6, 12, and 18 months. CRs at 12 months ranged from 43% to 64% of patients. Three patients had to stop treatment with MCC. At the time these data were disclosed, Bioniche was working on the design of a pivotal, multicenter program in North America and Europe to conÞrm the efÞcacy and safety of MCC in patients with high-risk SBC. Data were also presented from a study of the immune response following intravesical administration of MCC emulsion in patients with BC (Morales A, 2004). MCC induced an immune response against both MCC and bacillus Calmette-Gu´erin (BCG, generics), as deÞned by antibodies in these patients. Arsenic Trioxide. Arsenic trioxide (Cell Therapeutics’ Trisenox) is in Phase II trials in the United States as a single agent in the treatment of BC. Several clinical trials are evaluating the agent for various tumor types, but lead indications are hematologic cancers.

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Arsenic trioxide has been shown to inhibit growth and induce apoptosis in preclinical models, although its mechanism of action has not been fully elucidated. Preliminary preclinical data suggest it may be active against cells expressing the multidrug resistance (MDR) gene. Dose-limiting toxicities with arsenic trioxide include weight gain and peripheral neuropathy. The most common side effects have been dizziness during infusion, skin reactions, hyperglycemia, and musculoskeletal pain. Arsenic trioxide has activity against certain solid tumor cell lines, among which BC cell lines were the most sensitive to its effects (Small EJ, 2003), providing the rationale for the completed Phase II trial. This trial evaluated arsenic trioxide in 35 patients with recurrent urothelial carcinoma of the bladder, urethra, ureter, or renal pelvis (after failure of one previous line of chemotherapy). Bortezomib. Bortezomib (Millennium Pharmaceuticals’ Velcade) is a smallmolecule proteasome inhibitor that works through multiple pathways, including those inßuencing apoptosis and angiogenesis. Its lead indication is multiple myeloma, for which it is approved in the United States. Bortezomib is in Phase II trials in the United States for the treatment of BC. Proteasomes are enzyme complexes responsible for breaking down proteins, including those that regulate cell division, most notably nuclear factor kappabeta (NF-κβ). NF-κβ is implicated in BC (Karashima T, 2003). Bortezomib has high selectivity for the proteasome over other proteases (e.g., thrombin) and has demonstrated in vitro cytotoxicity against a wide range of tumor cell lines. Preclinical data show that bortezomib inhibited cell growth in a concentrationdependent fashion and augmented the growth-inhibitory effects of gemcitabine in vitro (Kamat AM, 2004), but no clinical data in BC seem to have yet emerged. Farnesyl Transferase Inhibitors Overview. The proto-oncogene Ras, which encodes a small GTP-binding protein, is required for both normal intracellular signaling via receptor tyrosine kinases (such as EGFR tyrosine kinases) and appropriate control of cell division. Mutated Ras occurs in up to 20% of BC cases (Quek ML, 2003), and substantial research has been invested in identifying inhibitors of Ras function. Farnesyl transferase inhibitors (FTIs) have emerged as potential therapies for the treatment of BC and other neoplastic diseases because they catalyze a critical post-translational modiÞcation step required for Ras activity. In preclinical models, unmodiÞed Ras molecules were unable to stimulate mitogen-activated protein kinase (MAPK), thereby inhibiting cell division (McGeady P, 1995). The only FTI in clinical trials for BC is Janssen Pharmaceutica’s tipifarnib (Zarnestra, previously R-115777), which we discuss further on. Mechanism of Action. Ras normally transduces intracellular signals from receptor tyrosine kinases; Ras signaling activates MAPK, thereby stimulating the promitogenic transcription factors Fos and Jun and promoting cell proliferation. Mutations of Ras stimulate uncontrolled cell proliferation by constitutive activation of MAPK. Ras function requires the addition of a farnesyl (lipid) group to

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a cysteine residue at the carboxy-terminus of the protein. This post-translational modiÞcation is catalyzed by farnesyl transferase; small-molecule inhibitors of this enzyme antagonize the modiÞcation of Ras, which in turn generates a naive form of Ras that is incapable of stimulating MAP-kinase-associated cell division. Tipifarnib. Janssen and its parent company, Johnson & Johnson, are developing the oral FTI tipifarnib. A Phase II trial will evaluate tipifarnib in recurred BC after at least one line of intravesical therapy. In the meantime, no clinical data on tipifarnib in the treatment of BC are available. Data available for tipifarnib in other cancers (pancreatic, breast, colorectal) have been disappointing, and overall, the FTIs show modest single-agent activity in various cancer trials and are associated with signiÞcant toxicity. Antifolates Overview. New-generation antifolate drugs are targeting multiple steps in the folate metabolism pathway, resulting in the inhibition of both purine and pyrimidine de novo synthesis. Of these antifolates, pemetrexed (Eli Lilly’s Alimta) is the only one in development for BC. Mechanism of Action. The folate pathway is involved in the metabolism and synthesis of the DNA precursors (purines); antifolates that inhibit this pathway interfere with DNA synthesis, causing tumor cell death. Previous generations of antifolates were directed principally at the enzyme thymidylate synthase, which is a key enzyme in the folate pathway. New generations of antifolates inhibit additional key enzymes involved in the folate pathway. Pemetrexed. Pemetrexed (Eli Lilly’s Alimta) (Figure 14) is a new-generation, multitargeted antifolate, which, at the time of composing this reference, is still in Phase II trials in the United States for the treatment of BC. Pemetrexed inhibits the enzymes involved in DNA synthesis, namely thymidylate synthase, as well as dihydrofolate reductase and glycinamideribonucleotide (GAR) formyltransferase. Promising Phase II data were disclosed at the 1998 European Society of Medical Oncology conference (Paz-Ares L, 1998). Researchers were able to evaluate 20 of 25 patients with metastatic BC (performance status less than or equal to

FIGURE 14. Structure of pemetrexed.

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2) who received pemetrexed (500–600 mg/m2 every three weeks). The median number of cycles per patient was three. Seven courses (10%) were dose-reduced and seven (10%) delayed. Grade III/IV hematological toxicity occurred in 15 patients (75%); Þve patients developed neutropenic fever. Nonhematologic toxicities included skin rash (grade III; one patient), stomatitis (grade III; one patient), and diarrhea (grade III; two patients). Two toxic deaths occurred. Seven patients (35%) experienced PR, Þve (25%) SD, and seven (35%) PD. On the basis of these results, a single-agent pemetrexed trial that delivers 500 mg/m2 with vitamin supplementation has been initiated in pretreated patients in several centers in the United States. A Phase II U.S. trial evaluating pemetrexed and gemcitabine in treating ABC patients is also ongoing. No signiÞcant data on this compound have been reported in recent years. Immunotherapies Overview. Several immunotherapies are being investigated for use in cancer treatment and/or prophylaxis, although only limited R&D is ongoing in BC. We have grouped all interesting immunotherapies in this section. Note that no signiÞcant positive data have been made available on these agents in BC. Mechanism of Action. The body’s immune system normally rids itself of aberrant cells before they have a chance to multiply, colonize, or metastasize. Immunotherapy takes advantage of these natural host defense mechanisms via the following agents: • • •

Monoclonal antibodies that target aberrant proteins on cancer cells. Cytokines that increase or activate host immune cells, such as T cells, macrophages, and natural killer cells. Vaccines that activate speciÞc host T-cell or antibody responses that attack the cancer cells.

These agents, also known as biological response modiÞers, include several marketed compounds, such as cytokines (interleukins [ILs] and interferons [IFs]), colony-stimulating factors (CSFs), monoclonal antibodies (MAbs), and vaccines. Immunotherapy is a general term encompassing a wide variety of mechanisms of action, but its fundamental mechanism is the production of an immune response to treat or prevent disease. IDM-2. IDM-2 (Immuno-Designed Molecules’ Bexidem) is a combination of anticancer agents incorporating macrophage-activated killer (MAK) cells. In March 2004, Immuno-Designed Molecules (IDM) began a Phase II/III European trial of IDM-2 in BC. IDM-2 is designed to destroy residual cancer cells after transurethral resection of SBC, much as BCG does; the MAK cells are produced by activating the patient’s own white blood cells ex vivo. MAK cells are designed to be reinjected into the patient and have the capacity to recognize and destroy cancer cells. IDM has two products based on MAK.

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The randomized trial will be carried out primarily in France, Belgium, Luxembourg, and Germany. Half of the patients will receive 12 intrabladder instillations of IDM-2 over a six-month period. The other half will receive BCG. Researchers will use an intermediate analysis of the results on the Þrst 138 patients to deÞne the exact number of patients for Phase III trials. Phase I/II data were presented at the European Perspectives on Bladder Cancer meeting in 2001 (IDM press release, November 2001). Seventeen patients with SBC (TaG3 or relapsed TaG2) having a high probability of relapse within a year received six weekly intravesical infusions of IDM-2 based on MAK obtained from autologous mononuclear cells. Five patients received maintenance therapy at three-month intervals. All patients were followed up for at least one year before IDM-2 treatment and two years after the Þrst IDM-2 injection to observe how many times their disease recurred before and after treatment. During the Þrst year after treatment initiation, eight recurrences were observed, compared with 34 recurrences during the year before IDM-2 treatment. During the second year of follow-up, ten recurrences took place. Only one patient experienced PD that necessitated a cystectomy, and only 12 minor (grade I or II) adverse events possibly related to the protocol were reported. Immunotherapy could eventually play a role in earlier stages of BC and/or after tumor resection, when tumor burden is low, to prevent or reduce the likelihood of recurrence. Although BCG is reasonably effective in this setting, there is room for a more effective agent, for an agent with a different side-effect proÞle, and, indeed, for an agent that can be used in combination with BCG. Alkylating Agents Overview. Alkylating agents are so named because of their ability to add alkyl groups to many electronegative groups under conditions present in cells. Many alkylating agents are already well-established treatments for various types of cancer but have not yet found a role in BC treatment. Mechanism of Action. Alkylating agents involve reactions with guanine in DNA. These drugs add methyl or other alkyl groups onto molecules where they do not belong. This action, in turn, inhibits their correct utilization by base pairing and causes a miscoding of DNA. The Þrst mechanism by which an alkylating agent damages DNA involves attaching alkyl groups to DNA bases. This alteration results in the DNA being fragmented by repair enzymes in their attempts to replace the alkylated bases. A second mechanism by which alkylating agents damage DNA is by forming crossbridges—bonds between atoms in the DNA. In this process, two bases are linked together by an alkylating agent that has two DNA binding sites. Cross-linking prevents DNA from being separated for synthesis or transcription. Alkylating agents’ third mechanism of action causes the mispairing of the nucleotides, leading to mutations. Apaziquone. Spectrum Pharmaceuticals is developing apaziquone (EOquin) for the potential treatment of BC. A Phase II trial in Europe is ongoing.

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The pro-drug is metabolized in human and murine cells to the alkylating agent by the reductive enzyme DT-diaphorase (DTD), which is preferentially expressed in cancer cells. The Phase II trial is testing 4 mg/40 mL of the compound for six weeks in SBC patients. At the time of composing this reference, the trials are still ongoing. Encouraging preliminary toxicology data for this trial were presented at the 15th AACR-NCI-EORTC meeting in 2003 (Puri R, 2003). Two weeks after tumor removal, patients (n = 6) were administered apaziquone intravesically on a weekly schedule for six weeks. Doses commenced at 0.5 mg/40 mL and doubled until toxicity or maximum dosage (16 mg/40 mL) was achieved. All patients tolerated doses of 4 mg/40 mL, and only minor, reversible side effects were experienced at higher doses. Two patients were able to tolerate the maximum dose. No recurrences have been observed after one year in the Þrst patient treated, nor after six months for the remaining patients. Apaziquone may have an advantage over other alkylating agents because it is selectively activated by DT-diaphorase. Consequently, it is more likely to play a potential role in BC therapy. REFERENCES Aben KK, Kiemeney LA. Epidemiology of bladder cancer. European Urology. 1999;36:6 (Curriculum in Urology 6.4;1–13). Adrian PM, et al. Intravesical instillation of epirubicin, bacillus Calmette-Gu´erin and bacillus Calmette-Gu´erin plus isoniazid for intermediate- and high-risk Ta, T1 papillary carcinoma of the bladder: a European Organization for Research and Treatment of Cancer Genitourinary Group randomized Phase III trial. Journal of Urology. 2001;166:476–481. Advanced Bladder Cancer (ABC) Meta-Analysis Collaboration. Neoadjuvant chemotherapy in invasive bladder cancer: a systemic review and meta-analysis. Lancet. 2003;361: 1927–1934. Al-Sukhun S, Hussain M. Current understanding of the biology of advanced bladder cancer. Cancer. 2003;97(suppl 8):2064–2075. American Joint Committee on Cancer Staging Manual . 5 th Edition. Lippincott William & Wilkins, Philadelphia, PA, 1997. Baselli EC, Greenberg RE. Intravesical therapy for superÞcial bladder cancer. Oncology (Huntington). 2000;14(5):719–729; discussion 729–731, 734, 737. Baselli EC, Greenberg RE. Maintenance therapy for superÞcial bladder cancer. Oncology (Huntington). 2001;15(1):85–91. Bassi P. BCG (Bacillus Calmette Guerin) therapy of high-risk superÞcial bladder cancer. Surgical Oncology. 2002;11(1–2):77–83. Bellmunt J, et al. A feasibility study of carboplatin with Þxed-dose gemcitabine in unÞt patients with advanced bladder cancer. European Journal of Cancer. 2001;37: 2212–2215. Bergman AM, et al. Synergistic interaction between cisplatin and gemcitabine in vitro. Clinical Cancer Research. 1996;2(3):521–530.

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Raitamen MP, et al. Impact of tumor grade, stage, number, size, smoking and sex on survival in patients with transitional-cell carcinoma of the bladder. British Journal of Urology. 1995;76(4):470–474. Rajala P, et al. Transurethral resection with perioperative instillation of interferon-alpha or epirubicin for the prophylaxis of recurrent primary superÞcial bladder cancer: a prospective randomized multicenter study—Finnbladder III. Journal of Urology. 1999;161:1133–1136. Redman BG, et al. Phase II trial of paclitaxel and carboplatin in the treatment of advanced urothelial carcinoma. Journal of Clinical Oncology. 1998;16(5):1844–1848. Ries LAG, et al., eds. SEER Cancer Statistics Review, 1975–2000 [electronic]. National Cancer Institute: Bethesda, MD; 2003. http://seer.cancer.gov/csr/1975–2000. Rischmann P, et al. AFU recommendations 1998. Committee on Cancer of the French Association of Urology. Progres en urologie: journal de l’Association francaise d’urologie et de la Societe francaise d’urologie. 1998;8(3):25. Rodel C, et al. Apoptosis, p53, bcl-2, and Ki-67 in invasive bladder carcinoma: possible predictors for response to radiochemotherapy and successful bladder preservation. International Journal of Radiation Oncology, Biology, Physics. 2000;46:1213–1221. Rodel C, et al. Combined-modality treatment and selective organ preservation in invasive bladder cancer: long-term results. Journal of Clinical Oncology. 2002;20(14): 3061–3071. Sabichi AL, Lippman SM. COX-2 inhibitors and other nonsteroidal anti-inßammatory drugs in genitourinary cancer. Seminars in Oncology. 2004;31(2 S7):36–44. Saxman SB, et al. Long-term follow-up of a phase III intergroup study of cisplatin alone or in combination with methotrexate, vinblastine, and doxorubicin in patients with metastatic urothelial carcinoma: a cooperative group study. Journal of Clinical Oncology. 1997;15:2564–2569. Schenkman E, Lamm DL. SuperÞcial bladder cancer therapy. ScientiÞc World Journal. 2004;4(suppl 1):387–399. Schottenfeld D, Fraumeni-JF J, eds. Cancer Epidemiology and Prevention. 2nd ed. Oxford: Oxford University Press; 1996. Schuster TG, et al. Pelvic recurrences postcystectomy: current treatment strategies. Seminars in Urologic Oncology. 2001;19(1):45–50. Sch¨uz J, et al. Cancer registration in Germany: current status perspectives and trends in cancer incidence 1973–1993. Journal of Epidemiology and Biostatistics. 2000;5: 99–107. Shariat SF, et al. p53, p21, pRB, and p16 expression predict clinical outcome in cystectomy with bladder cancer. Journal of Clinical Oncology. 2004;22(6):1014–1024. Sheehy OE, et al. Celecoxib associated with reduced risk of superÞcial bladder cancer (SBC) recurrence. Proceedings of the American Society of Clinical Oncology. 2003. Abstract 1539. Shelley MD, et al. A meta-analysis of randomised trials suggests a survival beneÞt for combined radiotherapy and radical cystectomy compared with radical radiotherapy for invasive bladder cancer: are these data relevant to modern practice? Clinical Oncology (Royal College of Radiologists [Great Britain]). 2004;16(3):166–171. Shiina H, et al. Immunohistochemical analysis of bcl-2 expression in transitional-cell carcinoma of the bladder. Journal of Clinical Pathology. 1996;49:395–399.

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Breast Cancer

ETIOLOGY AND PATHOPHYSIOLOGY Nosology Breast cancer (CaB) is the most common cancer in women worldwide. It arises from the epithelium that lines the lactiferous ducts (the conduits for milk) and lobules in the breast and is classiÞed as either carcinoma in situor invasive carcinoma. Carcinoma in situ(which is outside the scope of this report) is the term applied to lesions that express some characteristics of invasive cancers (abnormal nuclei, for example) but have not penetrated the mammary duct and entered the supportive tissue of the breast. Carcinoma in situ is treated by surgery with or without radiotherapy. The majority of primary breast cancers are adenocarcinomas, and approximately 50% are located in the upper outer quadrant of the breast. Invasive carcinoma (the subject of this study) usually arises in the ductal epithelium, with inÞltrating ductal carcinoma accounting for 75% of CaB cases. InÞltrating ductal carcinoma is associated with a high propensity to spread to lymph nodes and to distant organs, resulting in a poor prognosis. InÞltrating lobular carcinoma is the next most common histological type of CaB, accounting for approximately 10% of cases. Certain variants of this type of cancer are also associated with poor outcome, although metastatic disease is slightly less common with inÞltrating lobular CaB than it is with ductal CaB. The remaining types of breast cancer are relatively infrequent (accounting for less than 10% of cases), tend to remain localized or metastasize late, and have more favorable outcomes. At diagnosis, Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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CaB—by deÞnition—has begun to spread into surrounding breast tissue. Cells that separate from the tumor are the source of regional metastases (to the local lymph nodes and skin) and distant metastases (to other organs). Anatomy The main functional purpose of the breast in any mammal is the production of milk for sustaining offspring. Each breast comprises 15–20 lobes, which are composed of numerous tiny glands or lobules, termed alveoli or acini (Figure 1). The alveoli produce milk and other substances during lactation and are linked by thin tubes called the milk or lactiferous ducts. These ducts lead to the nipple in the center of the breast, which is within a dark area of skin called the areola. Adipose and connective tissues surround the lobes of glandular tissue. The amount of fatty tissue depends on many factors, including age, percentage of body fat, and heredity. Each breast is well vascularized by both the blood and the lymphatic system. The lymph vessels drain into the lymph nodes, clusters of which are found under the arm, above the collarbone, and in the chest, as well as in many other parts of the body. Both the lymphatic system and the blood system are responsible for the dissemination of tumor cells from a primary cancer in the breast. There are no muscles within the breast, but muscles lie under each breast and cover the ribs. Cooper’s ligaments connect the chest wall to the skin of the breast, giving the breast its shape and elasticity. Etiology Risk Factors. Many risk factors for CaB have been identiÞed to a greater or lesser degree of causal certainty. These risk factors fall into two classes: unavoidable risk factors and modiÞable risk factors that relate to lifestyle. Aside from

FIGURE 1. Structure of the breast.

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a genetic predisposition, the common factor in many of these risks is increased exposure to endogenous estrogen over a long period of time. In several large studies, researchers have established the signiÞcant risk factors for CaB that are detailed in Table 1. The “Germline Mutations” section details one of the most important risk factors listed in Table 1. These mutations are a focus because information about them could help clinicians target preventive therapy or prophylactic treatment to individual patients. Germline Mutations. Germline mutations are inherited and are present in all cells in the body. Experts estimate that up to 10% of all CaBs are linked to heritable factors (Sng JH, 2000). Mutations of the cancer susceptibility genes breast cancer 1 (BRCA1 ) and breast cancer 2 (BRCA2 ) are thought to be associated with most heritable cases of CaB (up to 80% of familial cases, 5–6% of the total number of cases). Other germline mutations associated with CaB include the following: •

•

• •

p53 mutations (such as in Li-Fraumeni syndrome, an inherited mutation of the p53 tumor suppressor gene causing high incidence of cancers in afßicted individuals); PTEN mutations (such as Cowden’s disease, in which sufferers have a germline mutation in the PTEN gene producing increased susceptibility to polyps and cancers); ATM (ataxia-telangiectasia mutation, which increases susceptibility to developing cancers by impairing the immune system); and Susceptible HRAS1 polymorphisms.

BRCA1, BRCA2, p53, and PTEN mutations are all highly penetrant (likely to be expressed), and women with any of these mutations are quite likely to develop CaB. For example, the risk of developing CaB for BRCA1 carriers is estimated to be as high as 85% by age 55 (Ford, D 1994); however, the risk associated is mutation-speciÞc, and as such, may be family-speciÞc. A signiÞcant fraction of the population may carry an ATM or a susceptible HRAS1 polymorphism, but these mutations have low penetrance; because few carriers actually develop CaB, it is difÞcult to determine the precise number of CaB cases triggered by these abnormalities. Population studies suggest that other, as yet unidentiÞed, genes are implicated in familial CaB. BRCA1 and BRCA2. Expressed in normal breast tissues, BRCA1 and BRCA2 are tumor suppressor genes that regulate the proliferation and differentiation of breast epithelial cells. Mutation of the BRCA genes confers a much higher susceptibility to CaB (Table 2). Familial CaB resulting from BRCA mutations tends to be aggressive and to have an early onset: the mean age at diagnosis is approximately 43 years and 47 years for BRCA1 and BRCA2 mutations, respectively. Women with BRCA mutations are also at risk of developing second primary breast tumors

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TABLE 1. Risk Factors Associated with Breast Cancer Age

Incidence of breast cancer (CaB) increases with age (McPherson K, 2000). The majority of cases occur in women aged 50 years or older (Ries LAG, 2004). Some studies have indicated that age may be a prognostic factor (Kim SH, 1998; Fowble BL, 1994; de la Rochefordiere A, 1993), and that younger women tend to have more aggressive cancer. Other studies indicate no differences. Exposure to Estrogen exposure increases with the number of ovulatory cycles. Earlier estrogen age at menarche (younger than 12 years), low or null parity, later age at menopause (older than 55 years), and later age (older than 30 years) at first full-term pregnancy are associated with increased risk of CaB (Adami HO, 1998; Huang WY, 2000). Use of oral contraceptives is also associated with increased risk, as is obesity, as high amounts of fat tissue can increase estrogen levels. Recent data from the United Kingdom Million Women Study claim an increased risk in CaB related to HRT usage—increased risk with duration of therapy and use of progesterone. However, the study has been criticized as being flawed, so any real risk of HRT remains uncertain and controversial. History of benign Evidence from clinical follow-up studies indicates a relationship between breast disease histologically proven benign breast disease and CaB risk and that risk varies according to the histological category of benign breast disease and hormonal status (Schnitt SJ, 2001). Three studies have reported that when biopsy revealed proliferative disease without atypia, subsequent risk was approximately 1.5 times greater; when biopsy revealed atypical hyperplasia (AH), risk was approximately 4.5 times greater. When patients with AH had a family history of CaB, subsequent risk approached that of patients with in situ carcinoma: approximately 8–10 times greater (Connolly JL, 1993). Race (related to African-American women aged 30–39 have higher incidence of CaB than familial risk) their white counterparts (Johnson ET, 2002). Mutations of the tumor suppressor genes BRCA1 and BRCA2 occur in 1/500 in the general population but in 1/40 in the Ashkenazi population (Winer EP, 2001). Heavy alcohol Consuming two to ten alcoholic beverages per week may confer a consumption 30–60% increase in risk of CaB (Gapstur SM, 1995). Alcohol appears to increase blood levels of estradiol. Other studies suggest an effect only in women who are also taking HRT (Ginsburg ES, 1996). Ionizing radiation Exposure to ionizing radiation increases CaB risk. Because of the long latency period for radiation-induced CaB and greater sensitivity of the developing breast to mutagenic damage, radiation exposure before the age of 40 incurs the highest risk; exposure after the age of 40 incurs minimal increase in risk. A marked increase in CaB has been reported in women who received mantle irradiation for treatment of Hodgkin’s lymphoma before the age of 15 (Winer EP, 2001). Familial risk Up to 1–6% of all cases are considered hereditary. A woman whose mother or sister had CaB has a 20% chance of developing CaB by the age of 40 and an 80% chance by the age of 70 (Colditz GA, 1996). Inherited germline Approximately 10% of CaB cases are attributable to BRCA1 and BRCA2 mutations mutations (Heisey RE, 1999; Hopper JL, 1999); mutations of these genes trigger a cascade of additional mutations that result in CaB. Lifestyle factors A high-fat, low antioxidant diet and a lack of exercise are generally associated with increased risk for most types of cancer. Note: Full source citations appear in ‘‘References.’’

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TABLE 2. BRCA1 or BRCA2 Mutation Carriers: Lifetime Breast and Ovarian Cancer Risk Genotype Normal (BRCA1, BRCA2) BRCA1 mutation carrier at age 50 BRCA1 mutation carrier at age 70 BRCA2 mutation carrier at age 70 Male BRCA2 mutation carrier

Lifetime Risk of Breast Cancer (%) 11–12 50 82 70 5–7

Lifetime Risk of Ovarian Cancer (%) 1.3 23 up to 50 up to 10 N.A.

N.A. = Not applicable.

and, particularly for BRCA1 mutations, ovarian cancer. These patients may also be more susceptible to other cancers, including stomach cancer and leukemia (Risch HA, 2001). Identifying all carriers of the BRCA genes in the general population is impractical because the test is expensive and may cause unnecessary concern to the individual and her family. However, physicians are increasingly screening and counseling women in families with a history of CaB. For those women identiÞed as high risk because of a high family incidence of cancer and conÞrmed by gene analysis, physicians usually recommend regular mammograms. Other options are prophylactic bilateral mastectomy and, for women with BRCA1 mutation who have do not wish to have any (further) children, an oophorectomy (removal of the ovaries) to eliminate this source of estrogen. The BRCA1 gene is located on chromosome 17q21. The BRCA1 protein comprises 1,863 amino acids. Scientists have identiÞed more than 300 different mutations of this gene. Two speciÞc mutations (a deletion and an insertion in the protein-coding sequence) appear in 22% of carriers; the remaining mutations are unique to different families. The BRCA2 gene is located on chromosome 13q12-13. The BRCA2 protein comprises 3,418 amino acids. Approximately 100 different mutations for the gene, with varying incidence, have been described to date. Mutations of BRCA genes in spontaneous, rather than inherited CaB, were thought to be rare, but recent evidence suggests otherwise. Alterations in BRCA1 function, as a result of a variety of causes including gene mutation, may play a signiÞcant role in the pathophysiology of sporadic CaB incidence and aggressiveness, particularly in sporadic invasive ductal carcinoma (Fraser JA, 2003). BRCA1 and BRCA2 proteins are also thought to play a role in response to DNA damage. This interaction could inßuence future treatment choice (Tassone P, 2003), although molecular proÞling data are not yet sufÞcient to have a major impact on this approach. Two recent publications report selective activity of poly(ADP-ribose) polymerase (PARP) inhibitors in BRCA1- and BRCA2deÞcient cells (Bryant S, 2005; Farmer H, 2005). PARP is an enzyme responsible for repairing DNA damage. The selectivity to BRCA-deÞcient cells is a result of the cells’ deÞciency in homologous recombination, making them acutely sensitive to PARP inhibitors because DNA damage is unable to be repaired. Treatment with

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PARP inhibitors is likely to be highly tumor speciÞc in BRCA-deÞcient patients; this approach is under preclinical evaluation. A third BRCA gene, BRCA3, is also associated with germline mutations that may confer a moderate-to-high risk of CaB. The gene is localized to chromosome 13q21, but evidence of this gene’s role in the etiology of CaB remains controversial (Kainu T, 2000; Thompson D, 2002). ATM. Mutations in ATM genes are known to increase the risk of CaB. This gene’s precise function is still unknown, but its sequence is similar to that of genes that encode proteins that regulate telomere length and cell-cycle progression. Patients who are homozygous possess two mutations of the ATM gene and are at high risk of developing ataxia-telangiectasia (AT, a rare inherited disorder associated with neurological disorders (and immune system deterioration); vascular lesions of the skin and eyes (telangiectasias); and radiation sensitivity. Possibly as a result, AT patients are at Þve to seven times higher risk than the general population of developing CaB; however, most AT patients die in their teenage years or early 20s, before reaching the age at which CaB usually manifests. Heterozygous ATM mutations (only one copy of ATM is mutant) have low penetrance and a low likelihood of the gene being expressed: up to 1% of the population may have a single mutant copy of ATM, but very few women in this subgroup develop CaB. This low penetrance also complicates analysis of the number of CaB cases that are attributable to inherited ATM mutations, because they may be considered sporadic cases: researchers estimate that ATM is a factor in 3.4–10% of all CaBs and could therefore be a more important factor than the BRCA1 and BRCA2 genes. They hypothesize that two classes of ATM mutations (truncating and missense) confer different risks of developing CaB (Meyn MS, 1999). Sporadic ATM mutations are less common than inherited mutations, but physicians may still be able to exploit the ATM genotype in patients with sporadic mutations to guide treatment decisions in CaB and other cancers. Because loss of ATM function is associated with radiation sensitivity, these patients may be better treated with aggressive surgery instead of breast-conserving surgery followed by radiotherapy. Researchers speculate that radiation exposure (e.g., through mammography) could also confer a higher risk for CaB in these patients (Werneke U, 1997; Bebb G, 1997). PTEN. The protein PTEN (phosphate tensin homologue/mutated in multiple advanced cancers) is a phosphatase involved in transmitting growth-inhibiting signals initiated by the transforming growth factor beta (TGF-β) signaling pathway. A somatic mutation in the PTEN gene causes Cowden’s disease; although this mutation is rare, 30–50% of women with this aberrant allele will develop CaB and are likely to develop bilateral breast tumors. Inherited PTEN mutations probably account for less than 5% of all CaB cases. Spontaneous mutations are rare, accounting for less than 0.3% of all CaB cases.

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HRAS1 Polymorphism. A regulatory DNA region (known as the HRAS1 minisatellite region), located downstream of HRAS1, could be responsible for a predisposition to CaB. Women who lack one of the four common minisatellite alleles at the HRAS1 locus are at increased risk of CaB. As with ATM gene mutations, the susceptible alleles have low penetrance but occur with high frequency and so are common in the general population—6% of the population may be carriers. Although rigorous studies are lacking, this mutation could contribute up to 9% of all CaB cases. Somatic Mutations. Somatic mutations occur randomly in individual cells throughout the body as a result of exposure to carcinogens or errors in DNA replication or DNA repair; a number of somatic mutations associated with CaB are listed in Table 3. Several mutations are needed to transform a cell, but in women who have an inherited mutation, fewer somatic mutations are needed. A large number of gene mutations have been identiÞed in CaB samples. This section discusses the most prevalent mutations and those that have attracted interest as potential targets for therapeutic intervention. Cell-Cycle Regulators. A number of cell cycle regulators—molecules that control the progression of cells through the cell cycle, such as cyclin D1 (CCND1), cyclin-dependent kinases, RB-1, and p16—have been shown to be mutant or have altered gene expression in some breast tumors. The protein products of these genes are key to cell-cycle regulation and are potential targets for chemotherapeutic intervention via such approaches as small-molecule, antibody, or antisense therapy. The gene CCND1 encodes cyclin D1, a protein that has a regulatory function in cell-cycle control. CCND1 is often overexpressed in CaB—13–24% of breast tumors have extra copies of CCND1, and an additional 13–50% have excess CCND1 without a gene ampliÞcation. CaB patients with estrogen receptor (ER)–negative and CCND1 -overexpressing tumors have a poor prognosis

TABLE 3. Somatic Mutations in Breast Cancer Genes BCL-2 HER1 Cyclin D1 (CCND1) p16 HER2 p53 RB-1 myc Ras PTEN DBC2 (early data on cell lines)

Prevalence (%) 63 9–51 13–50 40 10–40 22–30 15–20 5–20 12 5 60

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(Umekita Y, 2002). Data support a role for CCND1 as an ER-responsive or ERcoactivator gene in CaB; hence, it may play a role in selecting therapy in cases of sporadic CaB. Proteins needed during the S-phase of cell replication are repressed by pRb (RB-1 ), whereas p16 antagonizes CCND1. Although these genes are not prognostic factors, they are mutant in 15–20% and 40% of CaBs, respectively (Dickson RB, 2001; Han S, 2001). Human Epidermal Growth Factors. The human epidermal growth factor receptor (EGFR) HER2 is a transmembrane glycoprotein with intrinsic tyrosine kinase activity. Normally present as inactive monomers on the cell surface membrane, HER2 molecules form homodimers (pairs of HER2 receptors) or heterodimers with other HER receptors (HER1, for example). This interaction stimulates phosphorylation of the intracellular portion of HER2; strongly activates multiple intracellular signal-transduction pathways, including the Ras/MAP kinase pathway; and promotes cell proliferation. In normal cells, binding of the ligand to the extracellular portion of HER is necessary to stabilize the receptor dimers and maximally stimulate signal transduction. When EGFRs are involved in oncogenesis, they are usually overexpressed and are potent proliferative stimuli. Overexpression of HER2 is associated with several important phenomena: • • •

•

Aggressive tumors. Resistance to chemotherapy and radiotherapy. Ligand-independent cell division. Scientists believe that when HER2 is overexpressed, its homodimers can form spontaneously—in the absence of ligand—in numbers sufÞcient to stimulate signal transduction pathways that drive cell division. AmpliÞed ligand-dependent cell growth. Overexpression of HER2 ampliÞes ligand-dependent cell-growth signaling mediated through HER1, possibly by inhibiting the normal ligand-dependent loss of HER1 from the cell surface.

Scientists have known for several years that HER2 is overexpressed in approximately 25–28% of early-stage CaBs. In about 95% of cases, overexpression is caused by the presence of multiple copies of the HER2 gene. In the remaining cases, scientists believe, overexpression of HER2 is caused by abnormally high levels of transcription (production of HER2 messenger RNA) or translation (production of HER2 protein) because cells contain the normal two copies of the HER2 gene. Scientists do not yet understand why ampliÞcation of the HER2 gene occurs or why the product of the nonampliÞed gene is sometimes overexpressed, but other regulatory factors appear to be implicated in the process (Vernimmen D, 2003). HER2 protein overexpression in CaB is associated with shortened diseasefree survival and reduced overall survival in lymph node–positive disease and is therefore widely accepted as a marker for disease prognosis. The evidence

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for any prognostic signiÞcance of HER2 overexpression in node-negative CaB is less compelling. Some evidence suggests that HER2 overexpression can also predict the efÞcacy of adjuvant chemotherapy and hormonal therapy regimens used to treat CaB. For example, data published by A.D. Thor suggest that HER2 overexpression predicts response to a dose-intensive regimen of 5-ßuorouracil (5-FU), doxorubicin, and cyclophosphamide (FAC) in lymph node–positive CaB patients (Thor AD, 1998). These data revealed that patients with HER2-overexpressing tumors who were randomly assigned to dose-intensive FAC survived longer than patients assigned to intermediate or low doses. No dose-survival relationship was observed among patients with breast tumors that expressed normal levels of HER2; these patients did equally well regardless of FAC dose. Clinicians interpret these results to indicate that dose-intensive FAC should be considered only for treating tumors that overexpress HER2. Some experts have also concluded from this and related studies that an anthracycline (such as doxorubicin) should be included in adjuvant regimens for HER2-overexpressing CaB. However, scientists have yet to uncover the mechanisms that determine this apparent sensitivity to these chemotherapy agents. The development of the HER2-directed antibody trastuzumab (Roche and Genentech’s Herceptin), which blocks HER2-mediated signaling and has demonstrated a survival beneÞt in metastatic CaB, has driven the characterization of tumors for HER2 expression. There is a good correlation between degree of HER2 overexpression and response to trastuzumab. Assessment of HER2 expression, a factor for poor prognosis, at time of disease diagnosis is becoming increasingly routine in medical practice and is affecting treatment selection. The importance of the epidermal growth factor HER1 is still unclear, but if the prevalence of HER1 overexpression is as common as some studies suggest (up to 51% of all tumors), this protein may be a valuable drug target. Indeed, it is the major target for several emerging therapies, including geÞtinib (AstraZeneca’s Iressa) and erlotinib (Osi/Genentech/Roche’s Tarceva). However, despite preclinical evidence, no convincing demonstration of these agents’ clinical efÞcacy in CaB has been reported. Preclinical studies also suggest a possible preventive role for geÞtinib in a mouse model of cancer (Lu C, 2003; Konecny GE, 2003). Programmed Cell Death Regulators. Mutation of programmed cell death (apoptosis) pathway regulators contributes to tumorigenesis by allowing cells to continue dividing in the presence of growth-inhibiting conditions (absence of growth factors, hypoxia, severe DNA damage). Programmed cell death is activated by the p53 protein; the bcl-2 protein protects cells from death. In CaB, p53 gene mutations are relatively common (22–30% of all tumors), but bcl-2 gene mutations that cause overexpression of bcl-2 are more common (63% of all tumors). Both genes have prognostic signiÞcance and implications for therapy selection in CaB for the following reasons: •

p53 -negative tumors are unlikely to respond to doxorubicin treatment.

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• •

•

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p53 -negative tumors are more sensitive than p53 -positive tumors to taxane treatment. In estrogen-dependent tumors, exposure to this hormone upregulates BCL-2 protein expression, which also reduces the effectiveness of doxorubicin treatment. BCL-2 —positive tumors are usually ER-expressing and p53 -positive, both of which indicate a good prognosis.

Telomere Regulation. Telomeres, the ends of the strands of DNA that protect chromosome ends, shorten with every division. When telomeres are sufÞciently shortened, programmed cell death pathways are activated. In normal cells, this mechanism limits the number of cell divisions. Telomerase, an RNA-dependent polymerase that repairs telomeres after chromosome replication, is usually not expressed in normal cells, but cancer cells often express this enzyme. Many CaB tumors (up to 90%) express telomerase (Herbert BS, 2001); the prognostic signiÞcance of telomerase expression is unclear. Experimental studies to investigate the effects of antisense on telomerase are ongoing (Zhang X, 2003). Microarray Analysis. Microarrays are collections of nucleic acid combinations representing genes that are spotted or synthesized onto a substrate and then tested against a tissue sample. Microarray analysis measures mRNA levels and, thus, gene expression. Microarray analyses enable researchers to examine many different genes and discern expression patterns: for example, in diseased versus normal tissue, or in early-stage disease tissue versus late-stage disease tissue. These gene expression patterns have recently started to be used in diagnosis, treatment decisions, and target identiÞcation for new drug development. Somatic mutations in certain genes may be predictive of clinical outcome. However, the large number of genes implicated make prediction extremely complex. Reports indicate that using multimicroarray analysis to proÞle large numbers of genes could increase prognostic value in early disease compared with conventional prognostic indicators such as age, ER status, and HER2 expression (Van de Vijver, 2002). Knowledge and implication of gene expression in patients with CaB could guide treatment decisions, especially when gene-based therapies reach the clinic. Several clinical studies have been initiated that assign patients to either standard or aggressive therapy based on a gene scan of up to 90 genes. The outcome of these studies will determine whether gene signatures will be used in the future as a routine diagnostic method and for stratifying CaB patient populations to receive the optimal treatment regimen. Several companies are developing microarray tests to enable routine characterization of gene expression that would contribute to a more detailed diagnosis and selection of appropriate therapy according to the degree of risk of relapse. For example, the Institut Paoli-Calmettes in France has tested the Breast Cancer ProÞleChip (BCPC) (Borie N, 2004). Tumor samples from 220 anthracyclinetreated patients underwent gene analysis for 9,000 different genes. Signatures for

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estrogen receptor, progesterone receptor, bcl2, EGFR, VEGFA, and ErbB2 were analyzed and found to be quantitative, sensitive, and reproducible. One recently published study describes data generated from tumor samples from 447 patients (Paik S, 2003). From a collection of 250 genes, 16 cancerrelated genes were predictive for recurrence. Researchers created a formula that generates a “recurrence score” based on the expression patterns of these genes in a tumor sample. Ranging from 1 (lowest risk) to 100 (highest risk), the recurrence score is a measure of the risk that a given cancer will recur. A further 668 tumor samples were analyzed and assigned into low, intermediate, and high risk of recurrence groups based on their gene expression proÞle. The low-risk group contained 51% of the samples (with a score of less than 18); 22% were at intermediate risk (recurrence score 18 or higher but less than 31); 27% were at high risk (a score of 31 or higher). These risk-group divisions correlated well with the actual rates of recurrence of breast cancer after ten years. There was a signiÞcant difference in recurrence rates between women in the low- and highrisk groups. In the low-risk group, there was only a 7% rate of recurrence at ten years; in the intermediate- and high-risk categories, these rates were 14% and 31%, respectively. Up to a recurrence score of 50, rates of recurrence increased continuously as the recurrence score increased. These trends held true across age-groups and tumor size. Microarray analysis has the potential to change medical practice, enabling chemotherapy to be targeted to those women most likely to beneÞt from it. It could also spare women with low risk of recurrence, and who would not beneÞt from chemotherapy, the side effects and adverse impact on quality of life. Pathophysiology A number of factors have been identiÞed that inßuence the response to treatment, progression, and subsequent prognosis of CaB. The most important and established of these factors are described here, along with more experimental parameters that could become future biomarkers or targets for drug intervention. Tumor Stage. The TNM (primary tumor/regional lymph nodes/distant metastasis) staging system (Tables 4 and 5) is widely used to guide treatment. Stage of the tumor at diagnosis is the key determinant of prognosis. The TNM system classiÞes stage of disease according to tumor size (T), nodal involvement (N), and presence of metastases (M); see Table 4 for TNM staging deÞnitions. The size of the primary tumor, the presence and extent of lymph node involvement, and the presence of distant metastases all have implications for the selection of appropriate treatment and for prognosis. In addition, evaluation of ER and PR status is obligatory when gauging a patient’s candidacy for hormonal therapy. The presence of certain germline and somatic cell mutations and HER2 protein overexpression are associated with a poor prognosis (Dickson RB, 2001). Since HER2-directed therapy (Genentech/Roche’s Herceptin, Þrst approved by the FDA in 1998) became available, HER2 expression is more often assessed during diagnosis and affects treatment selection. Stage of disease is also an important prognostic indicator.

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TABLE 4. Definitions of TNM Staging Classifications of Breast Cancer Primary tumor (T) TX T0 Tis T1 T1mic T1a T1b T1c T2 T3 T4

T4a T4b T4c T4d

Primary tumor cannot be assessed. No evidence of primary tumor. Carcinoma in situ; intraductal carcinoma, lobular carcinoma in situ, or Paget’s disease of the nipple with no associated tumor. Tumor 2.0 cm or less in greatest dimension. Microinvasion 0.1 cm or less in greatest dimension. Tumor more than 0.1 cm but not more than 0.5 cm in greatest dimension. Tumor more than 0.5 cm but not more than 1.0 cm in greatest dimension. Tumor more than 1.0 cm but not more than 2.0 cm in greatest dimension. Tumor more than 2.0 cm but not more than 5.0 cm in greatest dimension. Tumor more than 5.0 cm in greatest dimension. Tumor of any size with direct extension to chest wall or skin. (Note: Chest wall includes ribs, intercostal muscles, and serratus anterior muscle but not pectoral muscle.) Extension to chest wall not including pectoral muscle. Edema (including peau d’orange) or ulceration of the skin of the breast or satellite skin nodules confined to the same breast. Both T4a and T4b. Inflammatory carcinoma.

Regional lymph nodes (N) NX N0 N1 N2

N2a N2b N3

N3a N3b N3c

Regional lymph nodes cannot be assessed (e.g., previously removed). No regional lymph node metastasis. Metastasis to movable ipsilateral axillary lymph node(s). Metastasis to ipsilateral axillary lymph node(s) fixed or matted or in clinically apparenta ipsilateral internal mammary nodes in the absence of clinically evident axillary lymph node metastasis. Metastasis in ipsilateral axillary lymph nodes fixed to one another or to other structures. Metastasis only in clinically apparenta ipsilateral internal mammary nodes and in the absence of clinically apparent axillary lymph node metastasis. Metastasis to ipsilateral infraclavicular lymph node(s) with or without axillary lymph node involvement or in clinically apparenta ipsilateral internal mammary lymph node(s) and in the presence of clinically evident axillary lymph node metastasis or metastasis in ipsilateral supraclavicular lymph node(s) with or without axillary or internal mammary lymph node involvement. Metastasis to ipsilateral infraclavicular lymph node(s). Metastasis to ipsilateral internal mammary lymph node(s) and axillary lymph node(s). Metastasis to ipsilateral supraclavicular lymph node(s).

Distant metastasis (M) MX M0 M1

Presence of distant metastasis cannot be assessed. No distant metastasis. Distant metastasis present.

a ‘‘Clinically apparent’’ is defined as detected by imaging studies (excluding lymphoscintography) or by

clinical examination or grossly visible pathologically. Source: American Joint Committee on Cancer; International Union Against Cancer; National Comprehensive Cancer Network, 2003.

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TABLE 5. TNM Staging System for Classification of Breast Cancer Stage of Disease

Tumor (T)

Node (N)

Metastasis (M)

0

Tis

N0

M0

I

T1a

N0

M0

IIA

T0 T1a T2

N1 N1 N0

M0 M0 M0

IIB

T2 T3

N1 N0

M0 M0

IIIA

T0 T1a T2 T3 T3

N2 N2 N2 N1 N2

M0 M0 M0 M0 M0

IIIB

T4

N0, N1, N2

M0

IIIC

Any T

N3

M0

IV

Any T

Any N

M1

a Including T1mic.

Note: TNM (primary tumor/regional lymph nodes/distant metastasis) is the internationally recognized classification system of the American Joint Committee on Cancer (AJCC) and the International Union Against Cancer (UICC). See Table 3 for definitions of the TNM staging classifications.

Tumor Grade. The tumor’s histological grade has a bearing on its aggressiveness and on consequent treatment and prognosis. Three features of the tumor are assessed when assigning a cancer’s grade: the frequency of cell mitosis (rate of cell division); tubule formation (percentage of cancer composed of tubular structures); and nuclear pleomorphism (change in cell size and uniformity). Each of these features is assigned a score ranging from 1 to 3 (1 indicating slower cell growth and 3 indicating faster cell growth). The scores of each of the cells’ features are then added together for a Þnal sum that will range from 3 to 9. A tumor with a Þnal score of 3, 4, or 5 is considered a grade 1 tumor (well differentiated). A score of 6 or 7 is considered a grade 2 tumor (moderately differentiated), and a score of 8 or 9 is a grade 3 tumor (poorly differentiated). Poorer survival rates are associated with higher grade tumors: Þve-year survival for grade 3 tumors is 50% compared with 75% for grade 2 tumors and 95% for grade 1 tumors. Local Spread to Lymph Nodes. As described in the preceding section, the number of tumor-positive lymph nodes detected in a patient affects both the prognosis and treatment of CaB. Data from 24,740 CaB patients recorded in the Surveillance, Epidemiology, and End Results (SEER) Program of the National Cancer Institute (NCI) were used to evaluate the breast cancer survival experience in a representative sample of U.S. women (Carter C, 1989). The Þndings from this study were used in the development of the treatment guidelines drawn

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TABLE 6. Breast Cancer Recurrence Rates and Number of Positive Lymph Nodes Number of Positive Nodes 0 1–3 4–6 7–9 10+

Recurrence at Five Years (%) 25 40 49 58 78

Source: Adapted from Nemoto T, 1980. Full source citations appear in ‘‘References.’’

up by the National Comprehensive Cancer Network (NCCN). Survival rates in the study varied from 46% (for large tumors with diameters of at least 5 cm and with positive axillary nodes) to 96% (for small tumors, less than 2 cm, with no involved nodes). Tumor diameter and lymph node status were found to act as independent but additive prognostic indicators. As tumor size increases, survival decreases, regardless of lymph node status; as lymph node involvement increases, survival status also decreases, regardless of tumor size. A linear relation was found between tumor diameter and the percentage of cases with positive lymph node involvement. However, for patients with BRCA1 -related CaB, this correlation does not hold true, implying that these tumors may behave differently (Foulkes WD, 2003). In a second large pivotal study, a total of 24,136 female patients with histologically conÞrmed breast cancer were studied (Nemoto T, 1980). In these patients, Þve-year survival rates were 73% for localized disease and 49% for regional disease. The number of metastatic nodes in the axilla was investigated, and it was demonstrated that patients with one or more positive nodes had reduced cure and survival rates compared with those with negative axillary nodes. With the increase in the number of positive nodes, a continuing associated decline could be seen in survival and cure. The clinical size of the tumor also correlated well with the prognosis. Along with increased tumor size came a gradual increase in the probability of axillary nodal involvement. However, axillary metastasis occurred in 25% of patients with tumor size smaller than 1 cm. When the axillary nodes were involved, the cure rate in those patients was not signiÞcantly better than for those with larger primary tumors in this study. These Þndings are summarized in Table 6. Micrometastases. Micrometastases—small clumps of cells that have separated from the primary tumor and entered other organs—command signiÞcant research interest because of the danger they pose to patients. Even patients whose cancers are completely resected may relapse months or years after initial treatment, presumably because of occult micrometastatic disease that escaped treatment. In CaB, common sites of metastasis include the lymph nodes, brain, bone, lung, liver, and skin (Pantel K, 1999). In many cases, though, these

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micrometastases do not exhibit the characteristics of rapidly proliferating cells, suggesting that they are not all destined to develop into secondary tumors. As the technology for detecting the presence of micrometastases in the blood, bone, and organs improves, physicians will be able to apply chemotherapy optimally in patients who are at the highest risk of relapse. It must be noted, however, that the inability to identify relevant markers remains a major obstacle to this approach, and even if micrometastases are identiÞed, researchers are not yet able to differentiate between those that will grow to become clinically relevant and those that will not. Patterns of Metastatic Spread. The site of metastatic disease in CaB will dictate the treatment given to the patient. The site and likelihood of developing metastatic disease depends on factors that include histological diagnosis and HER2 status (Porter GJ, 2004). Common sites of metastasis are shown in Figure 2. For example, although painful, and with a high incidence of bone complication, bone metastases are not typically life threatening. Bone is the most common site of spread in CaB: as many as 70% of patients with metastatic inÞltrating ductal carcinoma are affected. The lung or pleura, liver, and brain are also common sites of metastatic spread and ultimately the cause of death in CaB. Hormone-Receptor Activation. Both normal and cancerous breast tissue contain ERs and PRs. Activation of these receptors by the steroids estrogen and progesterone causes cell proliferation by modulating the expression of various growth factors. Even before ERs were identiÞed, physicians recognized the potential for arresting breast tumor growth by estrogen manipulation (originally by oophorectomy, and later by treatment with luteinizing hormone–releasing hormone [LHRH] antagonists or tamoxifen therapy). Oncologists can test the hormone-receptor (HR) status of breast tumors: patients with HR-positive (ER and/or PR) tumors are most likely to respond to estrogen-ablation therapies; patients with HRnegative tumors rarely respond to antiestrogen treatment. Approximately 70% of patients treated for breast cancer have ER-receptor-positive tumors, and of these patients, 60% will respond to endocrine therapy. In general, hormone expression is more common and more intense in postmenopausal women than in premenopausal women. Hormone status can also be used to assess a patient’s prognosis. HR-positive tumors are associated with slower growth and better prognosis. Patients who are HR-negative are more frequently HER2-positive (a characteristic of aggressive disease). In addition, HR-negative tumors often overexpress the signal transducer protein kinase C. This protein mitigates estrogen’s growth-promoting effect on the tumor. Between the diagnosis of early-stage disease and the development of metastatic CaB, receptor status can change—this fact has implications for the choice of treatment for metastatic disease (Holdaway IM, 1983). Cathepsin D. Cathepsin D is an estrogen-dependent lysosomal protease that is synthesized by normal tissues but overexpressed and secreted by some breast

ETIOLOGY AND PATHOPHYSIOLOGY

FIGURE 2. Patterns of spread in breast cancer.

219

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cancers. It is suspected that cathepsin D facilitates invasion and metastasis of breast cancer; indeed, levels of cathepsin D tend to be higher in node-positive cases of breast cancer. Overexpression of cathepsin D in breast cancer is associated with high risk of recurrence and poor survival. In a study of 199 patients with node-negative disease and 198 with node-positive disease, high levels of cathepsin D proved to be a signiÞcant predictor of reduced disease-free survival in the node-negative population only (median follow-up = 64 months) (Tandon AK, 1990). Relating the level of cathepsin D to other prognostic factors in the patients with node-negative disease, the researchers found an association with aneuploidy (more or less than the diploid number of chromosomes) but no association with estrogen or progesterone receptors, tumor size, or patient age. In multivariate analyses, a high level of cathepsin D was the most important independent factor for predicting shorter disease-free and overall survival in patients with node-negative disease. Cathepsin D distinguishes node-negative patients whose disease may recur (and therefore should receive adjuvant chemotherapy) from node-negative patients, whose disease probably will not recur. Elevation of cathepsin D indicates a poorer prognosis. E-Cadherin. E-cadherin is a transmembrane glycoprotein that mediates epithelial cell-to-cell adhesion. The loss of E-cadherin can result in the disruption of cell clusters. It is therefore postulated that E-cadherin may function as a tumor suppressor protein. The loss of E-cadherin has been associated with metastasis and poor prognosis in invasive CaB and can help differentiate pathologically between ductal and lobular neoplasms of the breast. Loss of E-cadherin appears to be a later step in the metastatic process compared with angiogenesis and the loss of nm23-H1 expression. Researchers found that E-cadherin is a useful marker for identifying patients with poor prognosis; in a retrospective analysis of E-cadherin in tumor samples, the 14-year disease-free survival was shown to be 84%, 80%, and 56% in patients with high, intermediate, and low E-cadherin, respectively (Heimann R, 2000). Soluble Urokinase Plasminogen Activator Inhibitor-1. Soluble urokinase plasminogen activator inhibitor-1 (uPA) is secreted as a virtually inactive singlechain pro-enzyme (pro-uPA), which can be converted into active uPA by limited proteolysis. The binding of uPA to the receptor uPAR strongly enhances and localizes the activation of surface-bound plasminogen into plasmin, which, with broad speciÞcity, degrades most components of the extracellular matrix and basement membranes and is thought to play a pivotal role in the metastatic process. Researchers have reported that the concentration of a serum-soluble form of the receptor (suPAR) correlates with CaB patient outcome (Risbro R, 2002). In a group of patients with low suPAR, only 11% experienced a relapse during the observation period, compared with 40% in the group with high suPAR levels. Similarly, in the analysis of overall survival, only 17% in the low suPAR group experienced an early death compared with 49% in the group with high suPAR levels. Thus, suPAR may become a prognostic marker that could aid in treatment selection.

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CURRENT THERAPIES The treatment of CaB employs a multimodal approach that comprises hormone therapy, chemotherapy, biological agents, radiotherapy, and surgery. Treatment selection is tied primarily to disease stage, estrogen and progesterone receptor (ER and PR) status, performance status, and, increasingly, HER2 expression. Hormone therapy and/or chemotherapy are given in the following circumstances: • • •

Neoadjuvant therapy (prior to surgery) to reduce tumor size and facilitate surgery. Adjuvant therapy (postsurgery) to prevent recurrence (both local and distant). Palliative treatment of metastatic disease, where it might also be used to prolong survival. At this time, treatment of metastatic CaB is not considered to be curative, although treatment, and survival, can be long term in a minority of patients.

Treatment choice reßects the speciÞc patient and tumor characteristics and the likelihood of relapse. These factors include the patient’s age, menopausal status, performance status, ER/PR status, tumor histology, level of HER2 expression, lymph node involvement, and presence of metastatic disease. A large number of drugs—given alone, in combination, or in sequence—have demonstrated clinical beneÞt in CaB patients and have been adopted into clinical practice. Generally, neoadjuvant and adjuvant chemotherapy uses combinations of drugs—each with a different mechanism of action and complementary toxicity proÞle—to maximize efÞcacy while minimizing toxicity. Adjuvant chemotherapy is usually followed by hormonal therapy in patients who are ER- and PR-positive. In cases of early-stage disease, hormonal agents are used in the majority of ER- and/or PR-positive patients as long-term (routinely Þve years) adjuvant therapy. Hormonal agents are also the preferred treatment option in appropriate metastatic CaB patients because these agents’ efÞcacy is comparable to that of chemotherapy in this setting; hormonal agents also possess excellent side-effect proÞles, which are beneÞcial for long-term therapies. Traditionally, the selective estrogen-receptor modulator (SERM) tamoxifen (AstraZeneca’s Nolvadex, generics) has been the cornerstone of hormonal CaB treatment. However, in recent years, aromatase inhibitors (AIs) have become the treatment of choice for postmenopausal women with metastatic disease because of apparent superior efÞcacy. Data also show that adjuvant anastrozole (AstraZeneca’s Arimidex) is more effective than adjuvant tamoxifen at extending disease-free survival; thus, anastrozole and other AIs are challenging tamoxifen’s monopoly of the hormone adjuvant market, particularly in the United States. A meta-analysis of 47 adjuvant polychemotherapy trials clearly demonstrated the beneÞt of adjuvant cytotoxic therapy for early-stage (stages I and II) CaB patients (Early Breast Cancer Trialists’ Collaborative Group [EBCTCG], 1998),

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and this approach has been widely adopted. Patients who received chemotherapy had lower mortality compared with patients who received no chemotherapy, regardless of nodal or ER status or tamoxifen therapy. The degree of beneÞt gained from chemotherapy varied depending on patient age, menopausal status, and nodal involvement. The consensus is that certain subpopulations of patients will respond better to certain therapies, but the optimal regimens are yet to be determined, and criteria for selecting appropriate patients to receive chemotherapy remain unconÞrmed. The absolute reduction in the risk of recurrence using polychemotherapy (multiple chemotherapeutic drugs) in women younger than 50 years was 10.4% in women with node-negative disease and 15.4% in women with node-positive disease. The absolute reduction in mortality rates in women younger than 50 years with the use of adjuvant chemotherapy was 5.7% in patients with axillary nodenegative disease and 12.4% in patients with axillary node-positive disease. The corresponding absolute reduction in recurrence in women aged 50 to 69 years was 5.7% and 5.4% for patients with axillary node-negative disease and axillary node-positive disease, respectively. The absolute reduction in mortality rates was 6.4% and 2.3% for patients with axillary node-negative disease and axillary node-positive disease, respectively. More recently, taxanes have been introduced into neoadjuvant and adjuvant chemotherapy treatment regimens, primarily for high-risk (typically nodepositive) patients. To date, mature data are available from three large trials in which patients were randomized to receive either a taxane-containing regimen or a non-taxane-containing regimen. Studies show that both combination and sequential therapy (sequential lines of various single-agent chemotherapies) have their place in the treatment of metastatic CaB. Given the heterogeneity of CaB, physicians must be ßexible in their approach to treating the disease. Thus, treatment of patients with metastatic disease tends to be very individualized; optimal treatment regimens have yet to be determined. Sequential therapy may be particularly appropriate for older patients or those with reduced performance status because it enables the optimal delivery and management of single-drug therapy and potentially reduces the risk of toxicity without compromising quality of life. Newer drug combinations, such as trastuzumab/paclitaxel (Roche/Genentech/ Chugai’s Herceptin; Bristol-Myers Squibb’s Taxol, generics) or docetaxel/ capecitabine (SanoÞ-Aventis’s Taxotere; Roche/Chugai’s Xeloda), confer survival advantages over single-agent therapy in metastatic CaB and have manageable side-effect proÞles. Such combination treatments may be preferable to sequential therapy among patients who require immediate reduction in their tumor burden. In the clinic, combination regimens are favored for Þrst-line treatment of metastatic disease, particularly for patients whose cancer is rapidly progressing. Although trastuzumab is appropriate for the treatment of only 25–28% of the patients with tumors overexpressing the human epidermal growth factor receptor2 (HER2), its excellent clinical proÞle and beneÞt for this poor-prognosis group of patients make it worthy of additional mention. The availability of HER2 testing

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is much more widespread than in previous years, and testing to determine HER2 status is now recommended, and performed, at Þrst diagnosis, regardless of stage. Trastuzumab is approved only for the treatment of metastatic CaB (either alone or in combination with a taxane) in HER2-overexpressing patients, depending on previous treatments and performance status. However, the rationale exists for introducing adjuvant trastuzumab into earlier stages of disease (Slamon D, 2001). So far, three large clinical trials investigating the beneÞt of trastuzumab in the adjuvant and neoadjuvant settings, have conÞrmed: Trastuzumab demonstrates substantial beneÞt in adjuvant setting. It is under regulatory review in both the US and Europe. Trials are NSABP and NCCTG Trials in the US and HERA in Europe. BCIRG Trial is the 4th trial, which is less advanced. Retrospective studies of randomized clinical trials have revealed a correlation between the overexpression of HER2 and resistance to cyclophosphamide/ methotrexate/5-ßuorouracil (CMF) chemotherapy and tamoxifen. Other studies have suggested an interaction between HER2 overexpression and enhanced response to dose-intensive doxorubicin-containing regimens in lymphnode–positive patients (although cardiotoxicity is problematic with this combination). This Þnding has led to interest in combining trastuzumab with liposomal formulations of doxorubicin, reportedly associated with reduced cardiotoxicity. Optimizing trastuzumab combinations with other agents and improving the trastuzumab schedule are ongoing efforts. Ongoing clinical trials are testing different combinations and sequences of existing agents. Indeed, interest has grown in the use of platinum agents as second- and subsequent-line treatments in metastatic disease, but they have not yet been established as standard treatment. Additionally, cancer patients with metastatic bone disease have been treated for many years with bisphosphonates, to allay these complications. These agents do not have a proven direct anti-tumor effect. However, many trials are ongoing to determine whether bisphosphonates can slow disease progression and whether they can play a preventive role when given to cancer patients not yet suffering from bone involvement. Herein, the focus is on the regimens in regular clinical use. Table 7 lists the leading monotherapies and combination therapies used to treat CaB. The evaluation of new agents and regimens is a slow process. Because CaB is a slowly progressing disease, particularly in postmenopausal patients, it takes many years of follow-up in large clinical trials to conÞrm any improvement in response rate or overall survival. Furthermore, many agents or combinations of agents may be producing only modest, incremental improvements that are difÞcult to ascertain with statistical accuracy. Cyclophosphamide/Methotrexate/5-Fluorouracil (CMF) Regimen Overview. The CMF regimen—cyclophosphamide/methotrexate/5-FU (generics)—was the Þrst to become established as adjuvant chemotherapy for CaB and was standard care for premenopausal, node-positive patients until its replacement by anthracycline-containing regimens. The CMF regimen represented the

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TABLE 7. Current Regimens Used for Breast Cancer Regimen or Class CMF

FAC

FEC

Regimen Components Agent

Availability

Dose

Cyclophosphamide US, F, G, I, (Bristol-Myers S, UK, J Squibb’s Cytoxan, Baxter’s Endoxan/ Endoxana, generics)

Cyclophosphamide: 100 mg/m2 /day on days 1–14 of a 28-day cycle for a total of 6 cycles. PO.

Methotrexate (Pfizer’s Farmitrexat, generics) 5-fluorouracil (generics)

US, F, G, I, S, UK, J

Methotrexate: 40 mg/m2 /day on days 1 & 8. IV.

US, F, G, I, S, UK, J

5-fluorouracil (generics)

US, F, G, I, S, UK, J

5-fluorouracil: 600 mg/m2 /d on days 1 & 8. IV. 5-fluorouracil: 500 mg/m2 /day on day 1 of a 21-day cycle for a total of 4 cycles.

Doxorubicin US, F, G, I, (Pfizer’s S, UK, J Adriamycin/ Adriblastine, Kyowa Hakko’s Adriacin, generics) Cyclophosphamide US, F, G, I, (Bristol-Myers S, UK, J Squibb’s Cytoxan, Baxter’s Endoxan/ Endoxana, generics) 5-fluorouracil US, F, G, I, (generics) S, UK, J

Common Toxicities • Myelosuppression. • Renal toxicity. • Alopecia. • Gastrointestinal toxicity. • Transient elevation of liver enzymes (methotrexate).

• Myelosuppression. • Cardiomyopathy. • Alopecia. • Gastrointestinal toxicity. • Skin rash, fever, chills (doxorubicin).

Doxorubicin: 50 mg/m2 /day on day 1.

Cyclophosphamide: 500 mg/m2 /day on day 1. All IV.

5-fluorouracil: 500 mg/m2 /day on days 1 & 8 of a 21-day cycle for a total of 4–6 cycles IV.

• Myelosuppression. • Cardiomyopathy. • Alopecia. • Gastrointestinal toxicity.

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TABLE 7. (continued) Regimen or Class

Regimen Components Agent

Availability

Epirubicin (Pfizer’s US, F, G, I, Ellence/ S, UK, J Farmarubicin, Kenfarma/Pfizer/ Kyowa Hakko’s Farmarubicin, generics) CyclophosUS, F, G, I, phamidea S, UK, J (Bristol-Myers Squibb’s Cytoxan, Baxter’s Endoxan/ Endoxana, generics) Doxorubicin/ Doxorubicin US, F, G, I, cyclophos(Pfizer’s S, UK, J phamide Adriamycin/ followed by Adriblastine, a taxane Kyowa Hakko’s Adriacin, generics)

Cyclophosphamide US, F, G, I, (Bristol-Myers S, UK, J Squibb’s Cytoxan, Baxter’s Endoxan/ Endoxana, generics) Paclitaxel US, F, G, I, (Bristol-Myers S, UK, J Squibb’s Taxol, generics)

TAC

Docetaxel US, F, G, I, (Sanofi-Aventis’s S, UK, J Taxotere)

Dose

Common Toxicities

Epirubicin: 50–100 mg/m2 / day on days 1 & 8. IV.

Cyclophosphamide: 75–100 mg/m2 / day on days 1–14. PO.

Doxorubicin: One of three doses—60 mg/m2 on day 1, or 75 and 90 mg/m2 on days 1 and 2. Every three weeks for 4 cycles.

• Granulocytopenia. • Infection. • Nausea and vomiting. • Hypersensitivity reactions. • Cardiotoxicity.

Cyclophosphamide: 600 mg/m2 IV on day 1.

Paclitaxel: 175 mg/m2. Every 3 weeks for 4 cycles after the completion of 4 cycles of doxorubicin/ cyclophosphamide. Docetaxel: 75 mg/m2. Every 3 weeks for 6 cycles.

• Anemia. • Neutropenia. • Amenorrhea. • Fever in absence of infection. • Hypersensitivity reactions. • Peripheral edema. • Neurosensory. • Skin events.

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TABLE 7. (continued) Regimen or Class

Regimen Components Agent

Availability

Doxorubicin US, F, G, I, (Pfizer’s S, UK, J Adriamycin/ Adriblastine, Kyowa’s Adriacin, generics) Cyclophosphamide US, F, G, I, (Bristol-Myers S, UK, J Squibb’s Cytoxan, Baxter’s Endoxan/ Endoxana, generics) Trastuzumab, Trastuzumab US, F, G, I, single agent (GenenS, UK, J tech/Roche/ Chugai’s Herceptin)

Trastuzumab/ Trastuzumab paclitaxel (Genentech/Roche/ Chugai’s Herceptin)

Paclitaxel (Bristol-Myers Squibb’s Taxol, generics)

Dose

Common Toxicities

Doxorubicin: 50 mg/m2.

Cyclophosphamide: 500 mg/m2.

Trastuzumab loading dose: 4 mg/kg, given on first day of treatment; maintenance dose: 2 mg/kg/day every 7 days. IV.

US, F, G, I, S, UK, J

Trastuzumab loading dose: 4 mg/kg, given on first day of treatment; maintenance dose: 2 mg/kg/day every 7 days. IV.

US, F, G, I, S, UK, J

Paclitaxel: 175 mg/m2 / day every 21 days. IV.

• Myelosuppression. • Neurotoxicity. • Hypersensitivity. • Cardiac toxicity. • Gastrointestinal toxicity. • Rash. • Pulmonary toxicity. • Fever. • Myelosuppression (neutropenia/ neutropenic infection). • Neuropathy/ neurotoxicity. • Hypersensitivity. • Cardiac toxicity. • Gastrointestinal toxicity. • Alopecia (paclitaxel). • Rash (trastuzumab). • Fever (trastuzumab).

CURRENT THERAPIES

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TABLE 7. (continued) Regimen or Class

Regimen Components Agent

Trastuzumab/ Trastuzumab docetaxel (Genentech/Roche/ Chugai’s Herceptin)

Availability US, F, G, I, S, UK, J

Docetaxel US, F, G, I, (Sanofi-Aventis’s S, UK, J Taxotere) Docetaxel/ Docetaxel US, F, G, I, capecitabine (Sanofi-Aventis’s S, UK, J Taxotere)

Capecitabine US, F, G, I, (Roche/Chugai’s S, UK, J Xeloda)

Docetaxel, Docetaxel US, F, G, I, single agent (Sanofi-Aventis’s S, UK, J Taxotere)

Dose Trastuzumab: loading dose: 4 mg/kg in week 1; maintenance dose: 2 mg/kg weekly. Docetaxel: 100 mg/m2 every 3 weeks for 6 cycles. Docetaxel: 75 mg/m2 /day every 21 days. IV.

Capecitabine: 2,500 mg/m2 / dayb , 14 days then 7 days rest. PO. Docetaxel: 100 mg/m2 /day every 21 days. IV. Premedication required.

Common Toxicities • Cardiac toxicity. • Febrile neutropenia. • Neutropenic sepsis.

• Myelosuppression (neutropenia and neutropenic fever). • Gastrointestinal toxicity. • Alopecia. • Cardiac toxicity. • Rash/pruritus. • Hyperbilirubinemia. • Stomatitis. • Hypersensitivity (docetaxel). • Hand-foot syndrome (capecitabine).

• Myelosuppression (leukopenia/ neutropenia)/ neutropenic infection. • Neurosensory toxicity. • Hypersensitivity. • Gastrointestinal toxicity. • Rash/pruritus. • Edema. • Fatigue.

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TABLE 7. (continued) Regimen or Class

Regimen Components Agent

Nanoparticle Paclitaxel paclitaxel, (Bristol-Myers single agent Squibb’s Taxol, generics)

Availability US

Dose Nanoparticle Paclitaxel: 260 mg/m2 administered every three weeks.

Common Toxicities • Thrombocytopenia. • Neutropenia. • Mild nausea. • Fatigue. • Alopecia. • Peripheral neuropathy.

Capecitabine, Capecitabine US, F, G, Capecitabine: single agent (Roche/Chugai’s I, S, 1,250 mg/m2 twice daily, Xeloda) UK, J 14 days, then 7 days rest. PO.

Gemcitabine/ paclitaxel

Gemcitabine (Eli Lilly’s Gemzar)

US, F, G, Gemcitabine: I, S, 1,250 mg/m2 /day on days 1 and 8 UK, J of a 21 day cycle. IV.

Paclitaxel US, F, G, Paclitaxel: (Bristol-Myers I, S, 175 mg/m2 on day 1. IV. Squibb’s Taxol, UK, J generics) Vinorelbine, Vinorelbine (Glaxo- US, F, G, Vinorelbine: single agent SmithKline/Pierre I, S, 30 mg/m2 /day every 7 days. IV. Fabre/Ellem/ UK, J Kyowa’s Navelbine)

Tamoxifen, Tamoxifen single agent (AstraZeneca’s Nolvadex, Pfizer’s Kessar, generics)

US, F, G, Tamoxifen: 20 mg I, S, daily. Toremifene: UK, J 60 mg daily. PO.

• Gastrointestinal toxicity. • Fatigue. • Thrombocytopenia. • Hand-foot syndrome. • Elevated liver enzymes. • • • •

Thrombocytopenia. Neutropenia. Neuropathy. Myalgia and fatigue.

• Myelosuppression (leukopenia/neutropenia). • Neuropathy. • Alopecia. • Gastrointestinal toxicity. • Fever. • Thromboembolic events. • Increased risk of endometrial cancer (2–7 times). • Hot flashes.

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TABLE 7. (continued) Regimen or Class

Regimen Components Agent

Availability

Dose

Common Toxicities • Gastrointestinal toxicity. • Transient increase in bone pain or swelling.

Anastrozole, Anastrozole single agent (AstraZeneca’s Arimidex)

US, F, G, I, S, UK, J

Anastrozole: 1 mg daily. Exemestane: 25 mg daily. PO.

Letrozole, Letrozole US, F, G, I, Letrozole: 2.5 mg single agent (Novartis’s S, UK daily until tumor Femora/Femara) (Preregistraprogression is evident. PO. tion in Japan)

Exemestane, Exemestane single agent (Pfizer’s Aromasin)

US, F, G, I, S, UK, J

Exemestane: 25 mg daily until tumor progression is evident. PO.

Fulvestrant, Fulvestrant single agent (AstraZeneca’s Faslodex)

US

250 mg intramuscular injection every 28 days.

Goserelin, Goserelin acetate single agent (AstraZeneca’s Zoladex)

US, F, G, I, S, UK, J

Goserelin acetate: 3.6 mg every 28 days. IV.

• Fatigue. • Asthenia. • Rash. • Hot flashes. • Headache. • Dyspnea.

• Asthenia. • Rash. • Hot flashes. • Headache. • Hot flashes. • Nausea. • Fatigue. • Headache. • Pain and inflammation at injection site. • Vasodilation (hot flashes). • Nausea and vomiting. • Hot flashes. • Impotence. • Decreased libido.

a Cyclophosphamide may be administered by IV infusion 200 mg/m2 on days 1 and 8 or 500–600 mg/m2 on day 1. b This dose is recommended, but clinicians generally use a dose of 2,000 mg/m2 /day because many patients do not tolerate the higher dose. US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan. IV = Intravenous; PO = Oral.

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prototype of combination chemotherapy that signiÞcantly improved early and long-term results. For decades, six cycles of CMF was the gold standard of adjuvant chemotherapy in CaB. CMF is largely being replaced with more-aggressive anthracycline and/or taxane-containing chemotherapy regimens, and the former regimen’s use is now restricted mainly to patients at low risk of relapse or those with poor performance status, such as the elderly, who are unable to tolerate more-aggressive regimens. CMF remains suitable for patients for whom anthracycline therapy is contraindicated, such as the elderly and those with a preexisting cardiac myopathy. It also demonstrates beneÞt in patients with a relatively low risk of recurrence, such as those with tumors that are ER positive, with negative or low expression of HER2, and with negative (uninvolved) lymph nodes. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

•

•

Cyclophosphamide (Figure 3) is an alkylating agent. These agents alkylate DNA bases, thereby producing “cross-links” that covalently link the two DNA strands and prevent cell replication. Methotrexate (Figure 4), a structural analogue of folate, is an antimetabolite. Antimetabolites block normal DNA synthesis by inhibiting several key enzymes. Methotrexate inhibits the enzyme dihydrofolate reductase. This inhibition interferes with the maintenance of the reduced folate pool. Reduced folates are essential for de novo synthesis of thymidylate and purine nucleotides—requirements for DNA synthesis and cell replication. Thus, by inhibiting dihydrofolate reductase, methotrexate stops the cell replication of rapidly proliferating cells, such as tumor cells. 5-FU (Figure 5) is a ßuoropyrimidine antifolate. Antifolates exert their cytotoxic effect via a number of mechanisms. One of the most important targets for 5-FU is thymidylate synthase (TS), a key enzyme in de novo thymidine synthesis. This pyrimidine, which requires folate cofactors, is an essential requirement for normal DNA synthesis. By inhibiting TS, 5-FU inhibits DNA synthesis and causes cytotoxicity. In addition, because 5-FU is a structural analogue of uracil, it is misincorporated into RNA in place of uracil

FIGURE 3. Structure of cyclophosphamide.

CURRENT THERAPIES

H2N

N

231

N CH3

N

N N H N

NH2 O

COOH COOH

FIGURE 4. Structure of methotrexate.

FIGURE 5. Structure of 5-fluorouracil (5-FU).

and so interferes with normal RNA function, an action that also triggers cell death. Clinical Performance. An early clinical trial, now with more than 20 years’ follow-up, convincingly demonstrated a survival advantage of CMF adjuvant chemotherapy compared with no adjuvant treatment. The trial recruited 386 women with node-positive CaB who were randomized to receive either no further treatment or 12 months of CMF treatment following surgery. The 20-year survival rates were 47% for those treated postoperatively with CMF versus 22% for the group receiving no chemotherapy. This beneÞt was particularly marked in patients younger than 50 years (Bonadonna G, 1995). The CMF regimen has been shown inferior to anthracycline-containing regimens in numerous studies in pre- and postmenopausal women, particularly those patients with poor prognostic factors such as lymph node involvement (see the following discussions of the FAC and FEC regimens). In the future, if patients who would beneÞt from anthracycline therapy can be identiÞed, the use of CMF in the remaining, lower-risk patients may increase and prevent unnecessary exposure of patients to anthracyclines’ cardiotoxic effects. Studies have been performed combining CMF with doxorubicin (PÞzer’s Adriamycin, generics), using various numbers of cycles and administering doxorubicin either before or after CMF. At this stage, although some data suggest an improved disease-free and overall survival following treatment with four cycles of doxorubicin followed by eight cycles of CMF, it remains uncertain whether combining both regimens offers an overall advantage. Currently, interest in the CMF regimen is waning, so further studies are unlikely. A large clinical study is under way in the United Kingdom to investigate the addition of epirubicin (PÞzer’s Ellence/Farmarubicin, generics) to CMF.

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The National Epirubicin Adjuvant Trial (NEAT) and Scottish Cancer Trials Breast Group (SCTBG BR9601) are analyzing 2,391 patients with early CaB; 2,021 eligible patients were entered into NEAT and 370 into SCTBG BR9601 between April 1996 and July 2001 (Poole C, 2003). NEAT compared epirubicin (100 mg/m2 × 4 cycles) followed by classical (c)CMF (×4 cycles) with six cycles of cCMF. The SCTBG BR9601 study compared epirubicin (100 mg/m2 × 4 cycles) followed by IV three-times weekly dose-modiÞed CMF (750 : 50 : 600 mg/m2 ) × 4 cycles) with three-times-weekly IV CMF (×8 cycles). The incidence of toxicity was higher in the patients receiving epirubicin in addition to CMF. Despite lower than predicted recurrence rates in the CMF arm, disease-free survival and overall survival were both superior in the epirubicin arm. The study indicates that this combination is an active regimen in this patient population, offering a highly signiÞcant improvement in both relapsefree and overall survival rates. These data add to the body of evidence that anthracyclines are a beneÞcial addition to adjuvant treatment and support the notion that this regimen has the potential to become an established treatment option. 5-Fluorouracil/Doxorubicin/Cyclophosphamide (FAC) Regimen Overview. Combination regimens that include 5-FU and cyclophosphamide are well established in the treatment of CaB, primarily as neoadjuvant and adjuvant therapy. Combination with an anthracycline in place of the antimetabolite methotrexate has been shown to have superior effects on long-term survival. Doxorubicin is now routinely used in combination regimens for the treatment of CaB—both in the adjuvant setting and in advanced disease. This use continues despite the increasing use of epirubicin since its introduction in 1985 (see the following section on the FEC Regimen). A consensus statement issued by the National Institutes of Health (NIH) in 2000 concluded that doxorubicin has a threshold dose effect in the adjuvant setting. Dose levels below this threshold demonstrate no clinical beneÞt, and once the threshold is reached, no further clinical beneÞt can be gained from exceeding it. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

5-FU (Figure 5) is a ßuoropyrimidine antifolate. Antifolates exert their cytotoxic effect via a number of mechanisms. One of the most important targets for 5-FU is thymidylate synthase (TS), a key enzyme in de novo thymidine synthesis. This pyrimidine, which requires folate cofactors, is an essential requirement for normal DNA synthesis. By inhibiting TS, 5-FU inhibits DNA synthesis and causes cytotoxicity. In addition, because 5-FU is a structural analogue of uracil, it is misincorporated into RNA in place of uracil and so interferes with normal RNA function, an action that also triggers cell death.

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233

FIGURE 6. Structure of doxorubicin (R = OCH3 , R1 = OH, R2 = H, R3 = H, R4 = OH).

•

•

Doxorubicin (Figure 6) is an anthracycline. Anthracyclines interact with several different cellular targets, most importantly topoisomerase II. By inhibiting this DNA religation enzyme, anthracyclines exert their cytotoxic effect. Another mechanism leading to cell death, known as DNA intercalation, involves insertion of the anthracycline molecule between base pairs. This phenomenon causes single- and double-stranded breaks in DNA that inhibit cell proliferation. The free radicals generated by the reductive metabolism of anthracyclines may also damage cellular structures. Cyclophosphamide (Figure 3) is an alkylating agent. These agents alkylate DNA bases, thereby producing “cross-links” that covalently link the two DNA strands and prevent cell replication.

Clinical Performance. Extensive evidence shows that including an anthracycline in adjuvant therapy regimens improves disease-free survival and overall survival (see the following section on the FEC Regimen). In a European study, 985 women with either node-negative or node-positive CaB were randomized to receive six cycles of either FAC or CMF (Martin M, 2003). The data indicated a nonsigniÞcant trend toward improved and overall survival with FAC. In a prospectively formed subset of node-negative patients, disease-free survival and overall survival were statistically superior in the FAC treatment arm. This advantage was not observed in the subset of node-positive patients. Although the clinical toxicity of FAC was greater than that of CMF, the levels were considered manageable and clinically acceptable. A meta-analysis of 11 randomized trials involving 5,942 patients compared anthracycline-containing chemotherapy with CMF (Kaklamani V, 2003). The investigators found a 12% reduction in recurrence with anthracycline-containing chemotherapy compared with CMF and an 11% reduction in the risk of mortality. Another important Þnding showed that anthracycline-containing regimens were superior to CMF only when they were given for a minimum of three months.

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Researchers remain unsure whether there are subpopulations of CaB patients who would beneÞt from anthracycline treatment. If these patients could be identiÞed, it would prevent other patients from being “overtreated” with toxic therapies. Retrospective subset analyses show that node-positive patients who also have high HER2 expression in their tumors (i.e., patients with poor prognosis) beneÞted the most from the FAC regimen. 5-Fluorouracil/Epirubicin/Cyclophosphamide (FEC) Regimen Overview. Combination regimens that include 5-FU (generics) and cyclophosphamide (generics) are well established in the treatment of CaB, primarily as neoadjuvant and adjuvant therapy (see the previously discussed FAC regimen). Regimens containing an anthracycline such as epirubicin, in place of the antimetabolite methotrexate, have been shown superior for long-term survival. The beneÞt of epirubicin treatment, as with other anthracyclines, in early-stage CaB may depend on patients receiving a sufÞciently high dose of epirubicin in each cycle and/or the cumulative dose. Therefore, clinicians generally extend the period between cycles or use growth factor stimulants (to support the bone marrow) rather than reduce the epirubicin dose per cycle in patients who experience serious side effects. Note, however, that the NIH consensus statement on adjuvant therapy for CaB endorses the view that increasing the dose of epirubicin (or another anthracycline) above the threshold level does not improve adjuvant treatment outcomes (Montemurro F, 2000). Studies suggest that epirubicin, the 4’-epimer of doxorubicin, has a better safety proÞle than its parent at equivalent dose levels (French Epirubicin Study Group, 1988; Italian Multicenter Breast Study with Epirubicin, 1988). Equitoxic doses are 60 mg/m2 of doxorubicin and 90 mg/m2 of epirubicin (Perez D, 1991). This evidence may encourage the use of FEC over FAC. Because of the cardiomyopathy associated with anthracyclines, these agents have a maximum lifetime dose that limits their extended use. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

•

5-FU (Figure 5) is a ßuoropyrimidine antifolate. Antifolates exert their cytotoxic effect via a number of mechanisms. One of the most important targets for 5-FU is thymidylate synthase (TS), a key enzyme in de novo thymidine synthesis. This pyrimidine, which requires folate cofactors, is an essential requirement for normal DNA synthesis. By inhibiting TS, 5-FU inhibits DNA synthesis and causes cytotoxicity. In addition, because 5-FU is a structural analogue of uracil, it is misincorporated into RNA in place of uracil and so interferes with normal RNA function, an action that also leads to cell death. Epirubicin (Figure 7) is an anthracycline. Anthracyclines interact with several different cellular targets, most importantly topoisomerase II. By inhibiting this DNA religation enzyme, anthracyclines exert their cytotoxic effect.

CURRENT THERAPIES

O

OH

235

O OH OH

H3CO

O H3C HO

OH

O

O NH2

FIGURE 7. Structure of epirubicin.

•

Another mechanism leading to cell death, known as DNA intercalation, involves insertion of the anthracycline molecule between base pairs. This phenomenon causes single- and double-stranded breaks in DNA that inhibit cell proliferation. The free radicals generated by the reductive metabolism of anthracyclines may also damage cellular structures. Cyclophosphamide (Figure 3) is an alkylating agent. These agents alkylate DNA bases, thereby producing “cross-links” that covalently link the two DNA strands and prevent cell replication.

Clinical Performance. A meta-analysis of 47 clinical studies involving more than 18,000 patients with early-stage CaB showed that adjuvant anthracyclinebased regimens (e.g., doxorubicin, epirubicin) are more effective than adjuvant nonanthracycline regimens (CMF was the principal comparator in the trials) (Early Breast Cancer Trialists’ Group, 1998). Anthracycline-treated patients had a 12% reduction in risk of recurrence and 11% reduction in risk of death compared with patients who received nonanthracycline regimens. The authors of the metaanalysis noted that these Þndings should be considered preliminary because the follow-up on the trials was short, the effect on survival was only marginally signiÞcant, and any increase in efÞcacy associated with the anthracycline regimens must be weighed against their increased toxicity, particularly in the adjuvant setting (Goldhirsch A, 2001). In the same analysis, the percentage of patients free from disease at Þve years ranged from 57% to 76% when an anthracycline was included in the adjuvant treatment regimen. A ten-year follow-up of a study directly comparing FEC with CMF in premenopausal, node-positive patients found that 52% of patients who underwent the FEC regimen were disease-free versus 45% given CMF. In addition, 62% of the FEC-treated patients had improved overall survival with reduced incidence of severe side effects compared with 58% of the CMF-treated group. Studies of epirubicin in metastatic CaB found that the higher dose, 100 mg/m2 , was more effective than the lower dose, 50 mg/m2 (French Adjuvant Study Group, 2001). These studies also found that in the adjuvant setting, node-positive CaB patients have a statistically signiÞcant superior Þve-year disease-free survival (66% versus 55%) and overall survival (77% versus 65%) with the 100 mg/m2

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(FEC100) regimen compared with 50 mg/m2 (both given for six cycles). This beneÞt is most pronounced in patients with more than three positive nodes (i.e., higher-risk patients). An update presented at the American Society of Clinical Oncology (ASCO) annual meeting in June 2003 showed that ten-year overall survival remains superior for the 100 mg/m2 group (54.8% versus 50.0%) (Bonneterre J, 2003). Because of an increase in the incidence and severity of toxicity, the FEC100 regimen is recommended only for patients at high risk of relapse. Doxorubicin/Cyclophosphamide Followed by a Taxane Regimen Overview. With their unique structure and mechanism of action, taxanes have demonstrated antitumor activity against a broad range of cancers. Incorporation of the taxane paclitaxel (Bristol-Myers Squibb’s Taxol, generics) into treatment regimens for metastatic CaB has demonstrated greater beneÞt than standard chemotherapy regimens. Researchers eagerly await further trials as well as follow-up of existing trials to help determine whether docetaxel is superior to paclitaxel and whether treatment with docetaxel is best given sequentially to, or concurrently with, doxorubicin or epirubicin (Hudis C, 2002). Paclitaxel is indicated for adjuvant treatment of node-positive CaB when administered sequentially to standard doxorubicin-containing combination chemotherapy. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

•

•

Doxorubicin (Figure 6) is an anthracycline. Anthracyclines interact with several different cellular targets, most importantly topoisomerase II. By inhibiting this DNA re-ligation enzyme, anthracyclines exert their cytotoxic effect. Another mechanism leading to cell death, known as DNA intercalation, involves insertion of the anthracycline molecule between base pairs. This phenomenon causes single- and double-stranded breaks in DNA that inhibit cell proliferation. The free radicals generated by the reductive metabolism of anthracyclines may also damage cellular structures. Cyclophosphamide (Figure 3) is an alkylating agent. These agents alkylate DNA bases, thereby producing “cross-links” that covalently link the two DNA strands and prevent cell replication. Paclitaxel (Figure 8) and docetaxel (Figure 9) are taxanes. Taxanes act by stabilizing the mitotic spindle microtubules, thereby preventing depolymerization. This stability inhibits the dynamic reorganization of the microtubules, resulting in a mitotic block at interphase with a consequential inhibition of cell proliferation and eventual cell death.

Clinical Performance. The Cancer and Leukemia Group B (CALGB) 9344 study enrolled 3,121 node-positive CaB patients with CaB between 1994 and 1999 at 516 study sites. Patients were randomized to receive either paclitaxel or

CURRENT THERAPIES

237

FIGURE 8. Structure of paclitaxel.

FIGURE 9. Structure of docetaxel.

no therapy following four cycles of AC. At a median of 69 months’ follow-up, the researchers found that the paclitaxel-treated group experienced a 17% reduction in the risk of recurrence and an 18% reduction on the risk of death (Henderson I, 2003). Both of these differences were statistically signiÞcant at this analysis point. At Þve years’ follow-up, there was a 3% improvement in overall survival: 80% for those patients receiving paclitaxel versus 77% for those not receiving paclitaxel. A subset analysis of patients revealed that those with hormone receptor (HR)–negative disease beneÞted more than the HR-positive patient population from the inclusion of paclitaxel into their treatment regimen. In the National Surgical Adjuvant Breast and Bowel Project’s (NSABP) B-28 trial, 3,060 node-positive patients were randomized to receive four cycles of AC alone or followed by four cycles of paclitaxel at 225 mg/m2 (every three weeks) (Crown J, 2004). With a median follow-up of 64 months, disease-free survival was signiÞcantly improved by adding the taxane—76% versus 72%; however, this improvement did not translate into improved overall survival in this study. A second NSABP trial (B-30) is comparing the addition of docetaxel to anthracyclines and cyclophosphamide (Crown J, 2004). The three arms include a combination of docetaxel and doxorubicin; a combination of docetaxel, doxorubicin, and cyclophosphamide; and a combination of doxorubicin and cyclophosphamide followed sequentially by docetaxel (AC followed by docetaxel). Researchers hope to accrue 4,000 node-positive CaB patients into the study; half of this target number had been reached in 2004 and, at the time of

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composing this reference, the Þnal data are still not available. A quality-of-life question is being asked regarding all patients. Many other randomized trials designed to evaluate the efÞcacy of adjuvant taxane therapy are ongoing or have recently completed recruitment. Until longterm survival data are available from these trials, some clinicians remain uncertain about the role of taxanes in adjuvant therapy, particularly in light of the perceived increase in toxicities. However, in the aforementioned trial, the majority of severe toxicities were observed during the AC part of treatment: for example, 62% of patients experienced granulocytopenia during AC treatment and only 16% during paclitaxel therapy, suggesting that these perceptions are unfounded. Docetaxel/Doxorubicin/Cyclophosphamide (TAC) Regimen Overview. In August 2004, the FDA approved injectable docetaxel (SanoÞ-Aventis’s Taxotere) for use in combination with doxorubicin and cyclophosphamide for the adjuvant treatment of women with operable nodepositive breast cancer. Approval followed in Europe in January 2005 for the same indication. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

•

•

Doxorubicin (Figure 6) is an anthracycline. Anthracyclines interact with several different cellular targets, most importantly topoisomerase II. By inhibiting this DNA re-ligation enzyme, anthracyclines exert their cytotoxic effect. Another mechanism leading to cell death, known as DNA intercalation, involves insertion of the anthracycline molecule between base pairs. This phenomenon causes single- and double-stranded breaks in DNA that inhibit cell proliferation. The free radicals generated by the reductive metabolism of anthracyclines may also damage cellular structures. Cyclophosphamide (Figure 3) is an alkylating agent. These agents alkylate DNA bases, thereby producing “cross-links” that covalently link the two DNA strands and prevent cell replication. Docetaxel (Figure 9) is a taxane. Taxanes act by stabilizing the mitotic spindle microtubules, thereby preventing depolymerization. This stability inhibits the dynamic reorganization of the microtubules, resulting in a mitotic block at interphase with a consequential inhibition of cell proliferation and eventual cell death.

Clinical Performance. As reported on the Taxotere prescribing information sheet, investigators enrolled 1,491 women with node-positive operable CaB in an international, multicenter, randomized trial (TAX316). Patients were stratiÞed according to the number of positive axillary lymph nodes (1–3, 4+) and were randomly allocated to receive adjuvant treatment with either docetaxel 75 mg/m2 administered one hour after doxorubicin 50 mg/m2 and cyclophosphamide 500 mg/m2 (TAC arm) or the gold-standard treatment of doxorubicin 50 mg/m2 followed by ßuorouracil 500 mg/m2 and cyclophosphamide

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239

500 mg/m2 (FAC arm). Both regimens were given every three weeks for six cycles. After the last cycle of chemotherapy, patients with positive estrogen and/or progesterone receptors (ER+ and/or PR+) received tamoxifen 20 mg daily for up to Þve years. The primary end point was disease-free survival, which included both local and distant recurrences of CaB and deaths from any cause. At a median followup of 55 months, results from the second interim analysis showed that patients treated with the TAC regimen has signiÞcantly longer disease-free survival than those treated with FAC. This was equivalent to an overall reduction in risk of relapse of 25.7% (hazard ratio = 0.74, stratiÞed log rank p = 0.0047) in favor of the taxane-containing regimen. As per the Clinical Study Report by Aventis Pharmaceuticals, Inc. ((# RP56976V-316); 2004, at the time of the interim analysis, with 55 months of follow-up and based on 219 deaths, overall survival was longer for TAC than FAC, with a reduction in the risk of death of 30% (hazard ratio = 0.69). Further analysis from this trial will be presented when survival data mature. In terms of the toxicity proÞle of TAC, women receiving TAC had an increase in anemia, grade 3 and grade 4 neutropenia, amenorrhea, fever in absence of infection, hypersensitivity reactions, peripheral edema, and neurosensory and skin events compared with those receiving FAC. The toxicity, while signiÞcant, did not cause a large number of patients to withdraw from treatment. As with other anthracycline/cyclophosphamide-containing regimens, long-term serious toxicity for the TAC regimen included a low incidence of leukemia (0.4%) and congestive heart failure (1.6%) (Smith RE, 2002). The approved dose of docetaxel for the adjuvant treatment of operable nodepositive breast cancer is 75 mg/m2 administered one hour after doxorubicin 50 mg/m2 and cyclophosphamide 500 mg/m2 every three weeks for six cycles. Following the success of the TAC regimen in the adjuvant setting, studies have been performed to determine whether this regimen has neoadjuvant activity prior to surgery for CaB. Interim data from 907 patients, reported at ASCO 2004 (Costa S, 2004), conÞrm activity of TAC in this setting. Patients were treated with two cycles of TAC, and responders (>50% reduction in tumor volume as measured by ultrasound) were randomized to receive either four or six further cycles of TAC. Patients not responding to two cycles of TAC were randomized to receive either four further cycles of TAC or four cycles of vinorelbine and capecitabine in combination. The overall response rate in the study was 75%. Neutropenia (grades 3 and 4) was the most common toxicity and was more frequent in patients receiving TAC compared with a combination of vinorelbine and capecitabine: 8.6% versus 1%. Trastuzumab, Single Agent Overview. Trastuzumab (Genentech’s/Roche/Chugai’s Herceptin) is a recombinant humanized monoclonal antibody (MAb) that binds to the HER2 receptor. The degree of HER2 receptor expression in tumors is represented using a scale

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from 0 (low or negative) to 3 (strongly positive). High expression of the HER2 receptor is demonstrated in 25–30% of CaB patients, making them eligible for treatment with trastuzumab. High HER2 expression increases the proliferative rate of tumor cells, resulting in poor prognosis. Trastuzumab’s initial indication was as a single agent for the treatment of patients with metastatic CaB whose tumors overexpress the HER2 protein and who have received chemotherapy for their metastatic disease. In June 2004, trastuzumab also gained approval, in Europe, for Þrst-line treatment, in combination with docetaxel, for patients whose tumors overexpress HER2. Several studies have conÞrmed that patients who do not overexpress HER2 do not beneÞt from trastuzumab. Studies are underway to determine whether there is any beneÞt of continuing to treat patients with trastuzumab, as a single agent, following disease progression. Typically, if a patient suffers disease progression while on chemotherapy, the treatment is ceased. However, when trastuzumab is used in combination with chemotherapy, possibly the tumor has acquired resistance only to the chemotherapy component of the regimen. Because the side-effect proÞle of trastuzumab as a single agent is relatively benign, studies are underway to identify whether patients remaining on trastuzumab gain a survival beneÞt. Currently, several important clinical trials are evaluating the role of trastuzumab in early disease or in the adjuvant setting. An April 26, 2005, press release from Roche and Genentech announced that primary and secondary end points had been met in both disease-free and overall survival. Mechanism of Action. Trastuzumab binds selectively and with high afÞnity to the extracellular domain of the human epidermal growth factor receptor 2 (EGFR2), also known as HER2, which signals cellular growth and proliferation. Downregulation of HER2 occurs when the trastuzumab antibody attaches to HER2 protein receptors on the cell surface. This binding causes some receptors to be internalized into the cell, reducing the signal for cell growth. Clinical Performance. In a single-arm, open-label monotherapy trial, the overall response rate (complete response plus partial response) was 14% for metastatic CaB patients overexpressing HER2 (2+ and 3+) who were pretreated with trastuzumab (Cobleigh MA, 1999). The median duration of response was 9.1 months, and median survival was 12.8 months. Twenty-four percent of patients were progression-free at 5.8 months. The degree of HER2 protein overexpression correlated with the degree of treatment effect, although both HER2 2+ and 3+ graded patients populations showed a beneÞt. As an antibody therapy, trastuzumab offers substantial efÞcacy without the severe toxicities of traditional cytotoxic chemotherapies. The agent’s most serious side effect is cardiac dysfunction similar to that observed with anthracycline treatment (Seidman A, 2002; Keefe D, 2002). This overlapping toxicity proÞle raises concern, particularly in the adjuvant setting, about the administration of protocols that contain both agents. In a Phase II trial of 469 patients, sequential administration of trastuzumab and an anthracycline caused grade 3 or 4

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left-ventricular ejection fraction (LVEF) events in approximately 4% of patients (Sledge GW, 2001). Physicians consider these levels low enough to warrant investigation of this combination for adjuvant therapy. Nevertheless, extensive cardiac monitoring is mandatory for patients in the ongoing Phase III adjuvant therapy trials using trastuzumab. Four ongoing, pivotal Phase III clinical trials are investigating the beneÞt of including trastuzumab in adjuvant treatment (for eligible HER2-positive patients). They are the B-31 adjuvant trastuzumab study—North Central Cancer Treatment Group (NCCTG) and North American Breast Intergroup study number N9831; the National Surgical Adjuvant Breast and Bowel Project (NSABP) study number 31; the Breast Cancer International Research Group (BCIRG) study number 006; and the Breast International Group (BIG) Herceptin Adjuvant (HERA) trial. All the trials have been designed to test one year of treatment (the HERA trial also tests two years) against no therapy at all; they mostly build on a backbone of adjuvant chemotherapy comprising AC followed by a taxane. The HERA clinical trial is a large-scale international clinical trial designed to determine if earlier use of trastuzumab can increase the length of survival without cancer recurrence in women whose early CaB has been diagnosed as HER2 positive, which tends to be less responsive to classical treatment. The BIG and Roche are collaborating in the conduct of the HERA trial, which recruited nearly 4,500 patients between December 2001 and March 2004 and is now closed to recruitment. The Þnal analysis of study results will take place two years after the Þnal patient has been entered into the study, mid-2006. Success of trastuzumab in this setting will likely mean a high uptake by clinicians. An April 25, 2005, press release from the Data Monitoring Committees that are overseeing the combined analysis of two of these adjuvant trastuzumab trials (NSABP-B-31 and NCCTG-N9831) recommended that the results of a recent combined interim analysis, with data from more than 3,000 patients, be made public because the studies had met their primary endpoints of increasing diseasefree survival in patients receiving trastuzumab in combination with chemotherapy. The improvement in overall survival was statistically signiÞcant. Patients in the clinical trials who received trastuzumab following treatment with standard combination chemotherapy (AC followed by T) also had a 52% decrease in disease recurrence compared with patients treated with chemotherapy alone. This difference is highly statistically signiÞcant. Trastuzumab/Paclitaxel Regimen Overview. Researchers have combined trastuzumab with different chemotherapeutic agents in experimental treatment regimens. Trastuzumab in combination with doxorubicin is contraindicated, as both agents have potential for serious cardiac adverse reactions. One established combination is trastuzumab and paclitaxel, which is used to treat patients with metastatic CaB whose tumors overexpress the HER2 protein and who have not previously undergone chemotherapy for their metastatic disease.

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Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

•

Trastuzumab binds selectively and with high afÞnity to the extracellular domain of the human epidermal growth factor receptor 2 (EGFR2), also known as HER2. Down regulation of the growth factor protein receptor HER2 occurs when the trastuzumab antibody attaches to HER2 protein receptors on the cell surface. This binding causes some receptors to be internalized into the cell, reducing the signal for cell growth. Preclinically, researchers have demonstrated trastuzumab’s synergy with several chemotherapies and hormonal therapies (e.g., treatment with trastuzumab prevents DNA repair following the impact of DNA-damaging drugs). Paclitaxel is a taxane. Taxanes act by stabilizing the mitotic spindle microtubules preventing depolymerization. This stability inhibits the dynamic reorganization of the microtubules, resulting in a mitotic block at interphase with a consequential inhibition of cell proliferation and eventual cell death.

Clinical Performance. A clinical trial comparing paclitaxel alone with paclitaxel in combination with trastuzumab as Þrst-line therapy for metastatic CaB demonstrated that the combination resulted in a marked improvement in response rate, time to disease progression, and overall survival (Slamon D, 2001). In another study, the overall response rate induced by trastuzumab/paclitaxel was shown to be independent of ER status: 44% for ER+ and 46% for ER–(Klein P, 2003). The study concluded that ER status is not a determinant of response to trastuzumab/paclitaxel therapy. In trastuzumab’s pivotal Phase III combination therapy trial, 469 previously untreated patients with metastatic CaB and HER2 overexpression at the 2+ or 3+ level received either anthracycline/cyclophosphamide (AC) or paclitaxel, plus either trastuzumab or placebo (Slamon D, 2001). The response rate for patients who received AC plus trastuzumab was 56% versus 42% in the AC/placebo group; the response rate for patients who received paclitaxel plus trastuzumab was 41% versus 17% in the paclitaxel/placebo group. For both the AC and paclitaxel treatment groups combined, the addition of trastuzumab increased the response rate from 32% to 50%, time to progression from 4.6 to 7.4 months, and overall survival from 20.3 to 25.1 months.

Trastuzumab/Docetaxel Regimen Overview. In June 2004, the European Commission approved the use of trastuzumab in combination with docetaxel in the European Union as a Þrstline therapy in HER2-positive metastatic breast cancer patients who have not yet received chemotherapy for their disease. Studies are also under way examining the role of trastuzumab in combination with docetaxel as a neoadjuvant treatment for HER2-positive patients with locally advanced CaB.

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Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

•

Trastuzumab binds selectively and with high afÞnity to the extracellular domain of the human epidermal growth factor receptor 2 (EGFR2), also known as HER2. Down regulation of the growth factor protein receptor HER2 occurs when the trastuzumab antibody attaches to HER2 protein receptors on the cell surface. This binding causes some receptors to be internalized into the cell, reducing the signal for cell growth. Preclinically, researchers have demonstrated trastuzumab’s synergy with several chemotherapies and hormonal therapies (e.g., treatment with trastuzumab prevents DNA repair following the impact of DNA-damaging drugs). Docetaxel is a taxane. Taxanes act by stabilizing the mitotic spindle microtubules, preventing depolymerization. This stability inhibits the dynamic reorganization of the microtubules, resulting in a mitotic block at interphase with a consequential inhibition of cell proliferation and eventual cell death.

Clinical Performance. The approval of docetaxel in combination with trastuzumab is based on study results that showed that, for women with HER2-positive CaB, the combination of trastuzumab and docetaxel signiÞcantly improved median life expectancy by more than one-third (31 months with trastuzumab plus docetaxel versus 22 months for docetaxel alone) (Burris H, 2000). The study also revealed that 61% of patients treated with the combination responded to treatment, compared with 34% of patients who received docetaxel alone. Data from a small Phase II study investigating trastuzumab and docetaxel in combination as a neoadjuvant treatment for patients with locally advanced CaB reported a 72% overall response rate to the combination. A partial response was seen in 47% of patients and a complete response achieved in 25% of patients (Hines J, 2003). Docetaxel/Capecitabine Regimen Overview. Like 5-FU, capecitabine (Roche/Genentech/Chugai’s Xeloda) (Figure 10) is a ßuoropyrimidine antifolate. It is approved in the United States and Europe as a second-line treatment for metastatic CaB, both as a monotherapy and in combination with the taxane docetaxel or as a Þrst-line treatment for patients ineligible for anthracycline therapy. When combined with docetaxel, capecitabine is also approved for the treatment of anthracycline-refractory patients. In Japan, Chugai has Þled for the approval of capecitabine as a third-line treatment for metastatic CaB. Clinical studies to assess the performance of capecitabine in combination regimens and in the adjuvant setting are ongoing. In May 2003, the United Kingdom’s National Institute for Clinical Excellence (NICE) endorsed the addition of capecitabine to docetaxel treatment for patients with locally advanced or metastatic CaB who have failed or are unsuitable for anthracycline therapy.

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O CH3 HN

O F

N O H3C

HO

O

N

OH

FIGURE 10. Structure of capecitabine.

Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

•

Capecitabine is a ßuoropyrimidine. It is converted to its active form—5FU—by thymidine phosphorylase (TP). TP is found in higher levels in cancer cells and therefore produces higher levels of the 5-FU metabolite in cancerous than in normal tissues. Fluoropyrimidines act by inhibiting thymidylate synthase (TS), a key enzyme in DNA synthesis. Docetaxel is a taxane. Taxanes act by stabilizing the mitotic spindle microtubules, thereby preventing depolymerization. This stability inhibits the dynamic reorganization of the microtubules, resulting in a mitotic block at interphase with a consequential inhibition of cell proliferation and eventual cell death.

Clinical Performance. In a randomized study involving patients with metastatic CaB, researchers administered docetaxel either alone or in combination with capecitabine (O’Shaughnessy J, 2002). The authors concluded that adding capecitabine to docetaxel treatment produced a statistically signiÞcant improvement in time to disease progression of 6.1 months compared with 4.2 months. They also observed an improvement in median survival—14.5 months with the combination regimen versus 11.5 months for docetaxel alone. Researchers at ASCO’s annual meeting in June 2003 presented data from two Phase II clinical studies involving patients with metastatic CaB. Patients were treated with docetaxel and capecitabine in combination (Mackey JR, 2003; Ramaswamy B, 2003). The Mackey study concluded that the increase in grade 3 toxicities observed with the combination (nail loss, asthenia, hand-foot syndrome) was difÞcult to manage and did not justify a change from the established dosing regimen of once every three weeks for docetaxel. The ongoing Ramaswamy study described toxicities as manageable and considered the interim response data to be good: of the 21 evaluable patients who were assessed, 14% demonstrated a complete response and 62% demonstrated clinical beneÞt. Other studies also

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observed that the combination of capecitabine and paclitaxel offers excellent efÞcacy (Gradishar WJ, 2001). Docetaxel, Single Agent Overview. The taxanes docetaxel (SanoÞ-Aventis’s Taxotere) (Figure 9) and paclitaxel (Bristol-Myers Squibb’s Taxol) (Figure 8) are both active against metastatic CaB. Clinical data have supported widespread single-agent application of these two taxanes in this population. Emerging clinical data indicate that docetaxel is more effective than paclitaxel, a Þnding that may result in docetaxel becoming the taxane of choice. Single-agent docetaxel is indicated for the treatment of CaB either after combination chemotherapy for metastatic disease fails, or if a relapse occurs within six months of adjuvant chemotherapy. Prior therapy should have included an anthracycline unless clinically contraindicated. Mechanism of Action. Taxanes act by stabilizing the mitotic spindle microtubules, thereby preventing depolymerization. This stability inhibits the dynamic reorganization of the microtubules, resulting in a mitotic block at interphase with a consequential inhibition of cell proliferation and eventual cell death. Clinical Performance. The response rate for single-agent docetaxel in patients with locally advanced or metastatic CaB who have failed Þrst-line therapy is 30–45% compared with 16–32% for single-agent paclitaxel (Mamounas EP, 2002). Response rates are slightly higher when taxanes are used as Þrst-line therapy for metastatic disease; here, too, docetaxel shows superiority over paclitaxel (50% versus 25–33%). Clinicians have observed improved response rates when a taxane is combined with an anthracycline and given as Þrst-line therapy (55–63% for docetaxel; 40–68% for paclitaxel). In a large Phase III randomized trial, patients with anthracycline-pretreated metastatic CaB were assigned to receive either docetaxel monotherapy or the combination of the alkylating agent mitomycin (generics) and the vinca alkaloid vinblastine (generics) (Nabholtz JM, 1999). Docetaxel demonstrated superior response rates to those of mitomycin/vinblastine (28% versus 10%) and superior median survival (11.4 versus 9.1 months). A comparison of docetaxel with doxorubicin in patients previously treated with an alkylating agent demonstrated a superior response rate in the docetaxel-treated arm but no improvement in overall survival (Chan S, 1999). Nanoparticle Paclitaxel, Single Agent Overview. The taxanes docetaxel (SanoÞ-Aventis’s Taxotere) (Figure 9) and paclitaxel (Figure 8) (Bristol-Myers Squibb’s Taxol) are both active against metastatic CaB and are increasingly used in the adjuvant setting, particularly for patients with a high risk of relapse. Clinical data have supported widespread single-agent application of taxanes in patients with metastatic CaB.

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Nanoparticle paclitaxel (American Pharmaceutical Partners’ Abraxane) is a formulation of paclitaxel that is also given intravenously. The nanoparticles of paclitaxel are bound to albumin—a naturally occurring plasma protein. And, in contrast to paclitaxel, the nanoparticle paclitaxel formulation is cremophorfree and has a higher solubility. The lack of cremophor means that nanoparticle paclitaxel does not require intensive steroid premedication, and the agent is associated with a lower risk of treatment-related severe hypersensitivity and potential life-threatening anaphylaxis and a lower incidence of neuropathy. Nanoparticle paclitaxel recently gained FDA approval as a single agent for the treatment of metastatic CaB in patients who have failed combination chemotherapy for metastatic disease or who have relapsed within six months of adjuvant chemotherapy. Prior therapy should have included an anthracycline unless clinically contraindicated. In April 2005, American Pharmaceutical Partners announced it would be initiating a Phase II clinical study of nanoparticle paclitaxel in one of the most widely used paclitaxel combination chemotherapy regimens. Nanoparticle paclitaxel will be used in place of paclitaxel and in combination with trastuzumab as Þrst-line therapy for HER2-overexpressing CaB. Mechanism of Action. Nanoparticle paclitaxel is a taxane. Taxanes act by stabilizing the mitotic spindle microtubules and thereby preventing depolymerization. This stability inhibits the dynamic reorganization of the microtubules, resulting in a mitotic block at interphase with a consequential inhibition of cell proliferation and eventual cell death. Clinical Performance. Data have been released from a Phase II study in 106 poor prognosis patients who had taxane-resistant progressive metastatic CaB and were given weekly nanoparticle paclitaxel (100 mg/m2 over 30 minutes; the typical weekly dose of paclitaxel would be administered over a minimum of three hours) (Blum J, 2003). A 15% overall response rate was achieved in this study, and 40% of patients were free of disease progression for four months; 30% were free of disease progression for six months. Nanoparticle paclitaxel was well tolerated; two patients experienced grade 4 toxicity (one thrombocytopenia and one neutropenia); common adverse events included mild nausea, fatigue, and alopecia. In a pivotal randomized controlled Phase III trial of nanoparticle paclitaxel, the safety and efÞcacy of 260 mg/m2 of nanoparticle paclitaxel was compared with 175 mg/m2 of paclitaxel, administered every three weeks (Perez E, 2005). The study was performed in 460 patients with metastatic CaB. The Phase III trial demonstrated that nanoparticle paclitaxel almost doubled the response rate (responses of speciÞc target lesions), 21.5% compared with 11.1% for paclitaxel, and prolonged time to tumor progression. The investigator reported response rates of 33% and 19% for nanoparticle paclitaxel and paclitaxel, respectively. In addition, the study conÞrmed that nanoparticle paclitaxel could be administered safely over 30 minutes without the need for steroid premedication and, despite

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being used at 50% higher dose than paclitaxel, nanoparticle paclitaxel was not associated with an increased rate of toxicities. Neutropenia (all grades) was experienced by 80% of patients treated with nanoparticle paclitaxel compared with 82% of patients with paclitaxel, with a lower incidence of grade 4 neutropenia in the nanoparticle paclitaxel arm of the study. A higher number of patients treated with nanoparticle paclitaxel experienced peripheral neuropathy (all grades, but no grade 4 reported) compared with those treated with paclitaxel: 71% versus 56%. The majority of patients experienced a rapid improvement in neuropathological symptoms. Capecitabine, Single Agent Overview. Capecitabine (Roche/Chugai’s Xeloda) (Figure 10), an orally administered pro-drug of 5-FU, has been available in all the major markets as a third-line treatment for locally advanced and metastatic CaB since 2002. Its approval was based on its activity in patients pretreated with anthracyclines and taxanes. Capecitabine is approved for the treatment of metastatic CaB as a monotherapy and in combination with docetaxel. Single-agent capecitabine is indicated for patients who are refractory to both paclitaxel and anthracyclines and for paclitaxel-resistant patients for whom further anthracycline therapy is contraindicated. In May 2003, NICE endorsed the use of capecitabine monotherapy treatment for patients who have failed both anthracycline- and taxane-based treatment. Researchers hypothesize that levels of thymidine phosphorylase (TP) enzyme expression may predict response to capecitabine. A retrospective analysis of a large Phase III trial showed that high levels of TP correlated with a favorable outcome in patients receiving 5’-deoxy-5-ßuorouridine (5’-FUDR, an intermediate form of capecitabine) but not in patients who did not receive the compound (Toi M, 2001). If similar results were obtained with capecitabine therapy, and a simple and effective test for TP expression was developed, physicians could target the use of capecitabine to patients who are most likely to respond to the agent, thus preventing unnecessary treatment. Mechanism of Action. Capecitabine is a ßuoropyrimidine. It is converted to its active form—5-FU—by TP. TP is found in higher levels in cancer cells and therefore produces higher levels of the 5-FU metabolite in cancerous tissue than in normal tissues. Fluoropyrimidines act by inhibiting thymidylate synthase (TS), a key enzyme in DNA synthesis. Clinical Performance. As a single agent, capecitabine produces signiÞcant responses in pretreated patients with metastatic disease. In a Phase II trial, 162 patients with metastatic CaB received capecitabine (1,250 mg/m2 twice per day on days 1–14 every 21 days) (Blum JL, 2001). All patients had previously been treated with paclitaxel, and the majority had also received an anthracycline (91%) and/or a 5-FU-based therapy (82%). Twenty percent of the patients

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responded to capecitabine, and an additional 43% achieved stable disease. The median time to disease progression was 3.0 months; median overall survival was 12.8 months. The most common adverse events were lymphopenia (94%), anemia (72%), diarrhea (57%), hand-foot syndrome (57%), nausea (53%), vomiting (37%), dermatitis (37%), neutropenia (26%), and thrombocytopenia (24%). These data were sufÞcient for capecitabine to receive accelerated approval by the FDA. An ongoing randomized adjuvant trial (CALGB-49907) is comparing capecitabine (alone) with AC or CMF in women over the age of 65 years. Oral, single-agent capecitabine is seen as an attractive choice for adjuvant treatment, particularly in low-risk, older, or poor performance status patients who are ineligible for more-aggressive chemotherapy treatment. Gemcitabine/Paclitaxel Overview. Gemcitabine (Eli Lilly’s Gemzar) (Figure 11) has largely replaced 5-FU as Þrst-line therapy for nonresectable or metastatic pancreatic cancer in all the major pharmaceutical markets. It is also approved for Þrst-line use, in combination with cisplatin (Figure 12), for nonresectable, locally advanced or metastatic nonsmall-cell lung cancer. Because of its favorable side-effect proÞle and convincing efÞcacy data, it is used extensively off-label as single-agent second-line or subsequent therapy for metastatic CaB. Several clinical trials are underway to evaluate the use of gemcitabine in combination with other cytotoxic agents (e.g., cisplatin, vinorelbine) as Þrst-line treatment and in anthracycline- and taxane-pretreated CaB patients. Gemcitabine in combination with paclitaxel has recently gained approval by the FDA for Þrst-line treatment of patients with metastatic CaB after they have had chemotherapy with anthracyclines. Patients contraindicated for anthracyclines are also eligible for gemcitabine/paclitaxel treatment. Mechanism of Action. Gemcitabine is an antimetabolite. It is a deßuorinated analogue of deoxycytidine with a novel mechanism of action. Gemcitabine undergoes intracellular metabolism by nucleoside kinases to its active form, leading to cytotoxicity via inhibition of DNA synthesis. Clinical Performance. In nine, mainly small, clinical studies of gemcitabine monotherapy, researchers disclosed response rates of up to 37% in the Þrst-line NH2 N HO

O O

N F

OH

F

FIGURE 11. Structure of gemcitabine.

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FIGURE 12. Structure of cisplatin.

setting, 26% in the second-line setting, and 18% or better in the third-line setting (Heinemann V, 2003). In view of these results, gemcitabine may be regarded as a valuable alternative to the palliative treatment of metastatic CaB because its unique mechanism of action may elicit responses in heavily pretreated patients. In the large pivotal study that formed the basis for gemcitabine’s FDA approval for CaB, 267 patients (median age 53 years) received gemcitabine plus paclitaxel; another 262 patients (median age 52 years) received paclitaxel alone (Albain K, 2004). The median overall survival was 18.5 months for gemcitabine with paclitaxel and 15.8 months for paclitaxel alone. One-year survival was 71% in the group that received the combination therapy, compared with 61% for the group treated with paclitaxel alone. The study found that 55% of patients in the paclitaxel arm stopped therapy because of disease progression, in contrast to only 38% of the combination drug group. This research also found that gemcitabine provides pain relief in symptomatic patients. Vinorelbine, Single Agent Overview. Vinorelbine (GlaxoSmithKline’s/Pierre Fabre’s Navelbine) (Figure 13) is used primarily as a single-agent option for second-line (or subsequent) treatment of locally advanced or metastatic CaB when anthracycline-based treatments either have failed or are unsuitable. Vinorelbine monotherapy has a response rate of about 24% in the pretreated CaB population. Combinations of vinorelbine with other chemotherapeutic agents have failed to show clinical beneÞt. For example, in a randomized comparison of vinorelbine plus doxorubicin with doxorubicin alone, no signiÞcant differences between treatment groups were detected

N

CH3 N

N H CH3O2C CH3O

CH3 N

OCOCH3 HO CO2CH3 CH3

FIGURE 13. Structure of vinorelbine.

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in either response rate (38% versus 30%), duration of response (median 7.2 months versus 6.8 months), or survival (median 13.8 months versus 14.4 months). Vinorelbine monotherapy often Þnds application in older CaB patients because it is a well-tolerated option. In the United Kingdom, NICE endorsed this application of vinorelbine in September 2003. Mechanism of Action. Vinorelbine is a semisynthetic vinca alkaloid. Vinca alkaloids interact with tubulin and disrupt microtubular function in the mitotic spindle. This action leads to metaphase arrest, resulting in mitotic arrest and cell death. Clinical Performance. A noncomparative study using vinorelbine as a Þrstline metastatic treatment in elderly (>60 years) CaB patients yielded an objective response rate of 38%, median duration of response of nine months, and median time to disease progression of six months (Vogel C, 1999). The major doselimiting toxicity was hematologic. Grade 3–4 nonhematologic toxicity consisted of asthenia (7%) as well as nausea and generalized pain (5%). Neurotoxicity and alopecia were grade 1–2 and relatively infrequent. Data that directly compares vinorelbine monotherapy with other monotherapy in pretreated patients with metastatic CaB are limited. However, a randomized controlled trial compared vinorelbine with melphalan (an agent not in current widespread usage for CaB) in 179 patients who failed to respond to anthracyclinecontaining chemotherapy (Jones S, 1995). A statistically signiÞcant survival beneÞt for vinorelbine was found. The median survival time for patients treated with vinorelbine was 35 weeks compared with 31 weeks for those treated with melphalan; the one-year survival rate was 35.7% for the vinorelbine group compared with 21.7% for melphalan. Decisions on treatment selection are based primarily on an agent’s toxicity and the patient’s previous treatments. Vinorelbine exists as one of several treatment options in this setting and is normally indicated for second-line or subsequent treatment in advanced CaB. Tamoxifen, Single Agent Overview. Tamoxifen (AstraZeneca’s Nolvadex, generics) (Figure 14) is the most widely studied SERM, and its role in the treatment of CaB is well established. The agent is approved for the treatment of all stages of CaB; patients who are HR-positive are most likely to respond to treatment. Tamoxifen is not recommended for the treatment of HR-negative CaB. In 1998, the FDA approved tamoxifen as a preventive agent in women at high risk of developing CaB. Tamoxifen is administered orally; the standard dose is 20 mg daily, and in the adjuvant setting, physicians generally prescribe it for Þve years or until relapse. Mechanism of Action. Tamoxifen is a SERM. SERMs act by interfering with estrogen’s signaling of epithelial cell growth and reproduction. As a mixed ER antagonist/agonist, tamoxifen can selectively stimulate or inhibit the ERs of different target tissues: this action is responsible for its protective effect on bone.

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FIGURE 14. Structure of tamoxifen.

Clinical Performance. Tamoxifen has been in widespread routine use for several decades and, until recently, has been virtually unchallenged in both the adjuvant setting and as Þrst-line hormonal treatment for metastatic CaB. Other agents were relegated to second-line or subsequent treatments. Studies have shown that tamoxifen increases the survival of women with CaB. The Early Breast Cancer Trialists’ Collaborative Group overview of randomized trials of adjuvant tamoxifen versus no tamoxifen showed that during approximately ten years of follow-up, the proportional reductions in mortality for one, two, and approximately Þve years of adjuvant tamoxifen therapy in ER-positive patients were 12%, 17%, and 26%, respectively (EBCTCG, 2000). As a result of these data, Þve-year adjuvant treatment with tamoxifen is now recommended. In a meta-analysis of 11 trials involving more than 15,000 patients, tamoxifen was shown to be equivalent in efÞcacy to adjuvant chemotherapy (EBCTCG, 2002). Since the emergence of the third-generation potent and selective aromatase inhibitors (AIs, discussed next), tamoxifen’s dominance has been under threat. Interim analysis has shown that for adjuvant treatment, the AI anastrozole (AstraZeneca’s Arimidex) offers a small but statistically signiÞcant survival advantage over tamoxifen and an improved side-effect proÞle, with the exception of the incidence of bone fractures (Anastrozole, Tamoxifen Alone and in Combination [ATAC] Trialist Group, 2002; Buzdar A, 2003; Baum M, 2003). However, until the outcome of the ATAC trial is elucidated, tamoxifen remains the standard adjuvant treatment for ER-positive patients. This agent will likely remain the preferred therapy for premenopausal patients with CaB because the aromatase route of estrogen production is less important in these patients. Anastrozole, Single Agent Overview. Anastrozole (AstraZeneca’s Arimidex) (Figure 15) is an aromatase inhibitor, or AI, approved in all the major markets for the adjuvant treatment (with or without radiation) of postmenopausal women with HR-positive early CaB. Approval was based on study results from patients taking anastrozole as adjuvant treatment for a median of two and a half years. This study is ongoing; further

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FIGURE 15. Structure of anastrozole.

follow-up of patients will be required to determine long-term results, including the agent’s side-effect proÞle and patient survival. Until these data become available, a complete comparison with tamoxifen is not possible. (Tamoxifen has more than 15 years’ detailed follow-up involving many thousands of patients.) Anastrozole is also approved for the treatment of locally advanced and metastatic CaB—either as a Þrst-line hormonal therapy or for patients currently taking tamoxifen whose disease has progressed. Mechanism of Action. Anastrozole is a selective nonsteroidal AI. AIs act by binding to the aromatase enzyme in place of androgens, inhibiting aromatase activity. Aromatase is the enzyme that converts androgens to estrogens; its inhibition results in the reduction of plasma estrogen levels, a promoter of cell replication in some breast tumors. Clinical Performance. The interim ATAC trial results show that patients treated with adjuvant anastrozole survived longer than those treated with tamoxifen (Baum M, 2001). More than 10,000 postmenopausal, hormonereceptor-positive, early-stage CaB patients were randomized to receive anastrozole, tamoxifen, or both. Accrual to the combination arm was halted early after evidence of inferiority to tamoxifen alone. Early analyses at 33 and 47 months reported that, compared with patients taking tamoxifen, patients taking anastrozole experienced a lower incidence of hot ßashes, weight gain, vaginal bleeding, endometrial cancer, and thromboembolic events. However, anastrozole patients had a higher incidence of arthralgia symptoms and bone fractures (Burstein HJ, 2002). A subsequent update of the ATAC trial, with a median follow-up of 68 months, was presented at the San Antonio CaB conference (Breast Cancer Symposium) in December 2004 (Howell A, 2004). The data showed that differences between anastrozole and tamoxifen in disease-free survival, time to recurrence, the occurrence of new tumors in the opposite breast, and metastases were even more pronounced at 68 months than they were at 47 months. However, no statistically signiÞcant improvement in overall survival between the two arms of the study was seen. Researchers noted that in this postmenopausal population, the course of

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FIGURE 16. Structure of letrozole (R = CN).

CaB can typically span 15 to 20 years, and patients often die of unrelated causes, making it difÞcult to conÞrm an overall survival beneÞt. Nevertheless, clinicians are already changing to anastrozole, in preference to tamoxifen, especially in patients with a high risk of thrombosis. In light of these ATAC data, ASCO has stated that Þve years of tamoxifen is no longer the optimal treatment choice in postmenopausal hormone-receptorpositive patients. The Society recommends that adjuvant therapy should include an AI, either as initial therapy or after treatment with tamoxifen, to reduce risk of recurrence. Letrozole, Single Agent Overview. Letrozole (Novartis’s Femora/Femara) (Figure 16) has been approved for the hormonal treatment of metastatic CaB. The drug was Þrst launched in 1997 in the United Kingdom for the second-line treatment of metastatic CaB. That same year, the FDA cleared letrozole as a once-daily therapy for the treatment of metastatic CaB in postmenopausal women with disease progression following antiestrogen therapy. The drug is also registered as a Þrstline therapy for advanced CaB. Letrozole is now being studied extensively for the treatment of early-stage CaB, the interim data for which are discussed in this section. Mechanism of Action. Letrozole is a selective nonsteroidal AI. AIs act by binding to the aromatase enzyme in place of androgens, thereby inhibiting aromatase activity. Aromatase is the enzyme that converts androgens to estrogens; its inhibition results in the reduction of plasma estrogen levels, a promoter of cell replication in some breast tumors. Clinical Performance. Two clinical studies (551 and 555 patients) evaluated letrozole’s tolerability and efÞcacy compared with that of megestrol acetate (generics) (Dombernowsky P, 1998) and aminoglutethimide (generics) (Gershanovich M, 1998) in patients with advanced CaB who had progressed on antiestrogen therapy. Letrozole showed superiority over both agents in tolerability and efÞcacy, including response rate and duration of response.

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A study comparing letrozole with tamoxifen as a Þrst-line hormonal therapy in patients with advanced CaB concluded that letrozole is signiÞcantly superior to tamoxifen in several efÞcacy parameters, including time to disease progression (41 weeks versus 26 weeks) and overall response rate (30% versus 20%) (Mouridsen H, 2001). Letrozole’s tolerability was reportedly equivalent to that of tamoxifen. In a randomized, double-blind neoadjuvant trial, 337 postmenopausal patients with large HR-positive tumors requiring mastectomy or breast-conserving surgery (BCS) were randomized to receive either preoperative letrozole or tamoxifen for four months (Smith I, 2003). The overall response rate was signiÞcantly better for letrozole (55% for letrozole versus 36% for tamoxifen). Neoadjuvant treatment allowed BCS in 45% of the patients treated with letrozole versus 35% treated with tamoxifen. Again, the improvement observed in the letrozole group was statistically signiÞcant. Interim data from the Breast International Group (BIG) 1–98 study were disclosed at the St. Gallen Primary Therapy of Early Breast Cancer conference in January 2005 (Thurliman {Thurlimann} B, 2005). BIG 1-98 is a multinational Phase III double-blind, randomized multicenter trial that is being conducted in 27 countries and has recruited more than 8,000 postmenopausal women with early CaB who have hormone-receptor-positive tumors. The primary endpoint of the study is to determine if letrozole can reduce the risk of recurrence or death compared with tamoxifen. At a median follow-up of 26 months, the study showed that, compared with tamoxifen, letrozole reduced the risk of such events by 19%. Among the 4,003 patients in the letrozole group, 84% remained alive and disease-free at Þve years compared with 81% of the 4,007 patients in the tamoxifen group. As in other studies with AIs, the incidence of thromboembolytic events and endometrial changes were reportedly lower with letrozole than with tamoxifen. Conversely, letrozole was associated with a higher incidence of bone complications, such as fractures. Exemestane, Single Agent Overview. Exemestane (PÞzer’s Aromasin) (Figure 17) has been approved for the treatment of hormone-receptor-positive, postmenopausal metastatic CaB patients who have progressed on tamoxifen (either in the adjuvant or metastatic setting). Studies investigating exemestane for Þrst-line metastatic treatment and adjuvant treatment of CaB are ongoing. Data showing a beneÞt from switching from adjuvant tamoxifen (following two to three years of treatment) to exemestane for a further two to three years (Coombes RC, 2004) has driven exemestane’s off-label usage in high-risk early-stage patients. Mechanism of Action. Exemestane is a selective steroidal AI. Steroidal AIs act by irreversibly binding to the aromatase enzyme in place of androgens, inhibiting aromatase activity. Aromatase is the enzyme that converts androgens to estrogens; its inhibition results in the reduction of plasma estrogen levels, a promoter of cell replication in some breast tumors.

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FIGURE 17. Structure of exemestane.

Clinical Performance. In a pivotal randomized, double-blind, multicenter, multinational, comparative clinical trial, exemestane was administered once daily to postmenopausal women with advanced breast cancer who experienced disease progression after treatment with tamoxifen for metastatic disease or as adjuvant therapy. Some patients also had received prior cytotoxic therapy, either as adjuvant treatment or for metastatic disease. Three hundred sixty-six patients were randomized to receive exemestane once daily, and 403 patients received megestrol acetate (the comparator of choice at the time of the study) four times daily. In the study, exemestane reduced the relative risk of tumor progression by 16%: patients experienced a median time to tumor progression of 20.3 weeks compared with 16.6 weeks for megestrol acetate. There was no statistically signiÞcant difference in the number of women having a clinical response. Both responders and nonresponders to tamoxifen beneÞted from exemestane treatment. In an adjuvant, double-blind clinical trial, the Intergroup Exemestane Study (IES), women who had already received two to three years of tamoxifen were randomized to one of two groups (Coombes RC, 2004). Either they completed the Þve-year course of tamoxifen or they switched to the aromatase inhibitor exemestane to complete a total of Þve years of hormonal treatment. Of the 4,742 patients enrolled, 2,362 were randomly assigned to switch to exemestane, and 2,380 continued to receive tamoxifen. The study demonstrated that the women who switched to exemestane had increasing disease-free survival compared with women who remained on tamoxifen. Overall survival was not signiÞcantly different in the two groups, with 93 deaths occurring in the exemestane group and 106 in the tamoxifen group. Severe toxic effects of exemestane were rare. Contralateral breast cancer occurred in 20 patients in the tamoxifen group and 9 in the exemestane group. This result was statistically signiÞcant. Fulvestrant, Single Agent Overview. Fulvestrant (AstraZeneca’s Faslodex) (Figure 18) is a low-toxicity, synthetic hormonal agent that overcomes tamoxifen resistance. In April 2002, the FDA approved fulvestrant for second-line treatment of patients with metastatic CaB whose disease had progressed despite prior antiestrogen therapy. It has now been approved in the UK and other European countries.

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H3C

OH

H O H

H

F

F

S HO

CF3 FIGURE 18. Structure of faslodex.

In addition, a European Phase III trial comparing fulvestrant and tamoxifen as Þrst-line therapy for patients with metastatic CaB has completed accrual, and AstraZeneca is planning trials in the adjuvant setting and in premenopausal women. In Japan, Phase III trials are ongoing in patients with metastatic CaB. Mechanism of Action. Fulvestrant is a selective estrogen receptor downregulator (SERD). SERDs act by binding to the cellular estrogen receptor in a competitive manner and downregulating estrogen receptors. Fulvestrant has no known estrogen-agonist effects and, because of its unique mechanism of action, shows no cross-resistance with SERMs. Clinical Performance. Two Phase III trials (one European and one North American) compared fulvestrant with anastrozole in ER-positive, metastatic, postmenopausal women in whom tamoxifen therapy had failed. The European trial compared 250 mg of fulvestrant (delivered as a single monthly intramuscular injection) with 1 mg/day anastrozole. The North American trial involved a third arm (125 mg fulvestrant), which was closed prematurely when a planned interim analysis showed that predeÞned efÞcacy criteria had not been met. In the North American trial involving 400 patients, the 250 mg fulvestrant group had a superior duration of response (19.3 months) compared with the anastrozole group (10.5 months)—a beneÞt that was not reproduced in the 451patient European trial (14.3 months and 14.0 months). Investigators suggested that this discrepancy may have resulted from differences in the frequency of follow-up (Carlson R, 2002). An update from the European trial reinforced the lack of a signiÞcant difference in survival rates between fulvestrant and anastrozole: time to death was 26.4 months for the fulvestrant group versus 24.2 months for the anastrozole group (Howell A, 2003). A combined analysis of the North American and European trials found a (nonsigniÞcant) trend toward improved efÞcacy with fulvestrant compared with anastrozole. At the annual meeting of ASCO in June 2003, researchers presented interim data from an ongoing study in Japan (Watanabe T, 2003). Thirty patients who had relapsed on antiestrogen therapy were treated with fulvestrant. A clinical beneÞt rate (complete response plus stable disease) of 60% led the authors to conclude that fulvestrant represents another treatment option for patients with metastatic CaB.

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Fulvestrant’s toxicity proÞle is comparable to that of tamoxifen and aromatase inhibitors. In the aforementioned Phase III trials, the incidence of side effects was similar in patients taking fulvestrant and anastrozole. Side effects of fulvestrant included hot ßashes (21%), vasodilatation (14%), and nausea (10%) (Jones S, 2001). Because fulvestrant has no estrogen-agonistic effects, clinicians do not expect the agent to cause endometrial cancer or to have a favorable effect on bone density and serum lipid levels. Goserelin, Single Agent Overview. Gonadotrophin-releasing hormone (GnRH) agonists—in particular goserelin (AstraZeneca’s Zoladex)—enable temporary suspension of estrogen production from the ovaries and, when used in combination with tamoxifen, result in total estrogen blockade. In terms of clinical beneÞt, this hormonal approach to the treatment of premenopausal ER-positive CaB has proven to be equivalent to chemotherapeutic regimens. Indeed, some physicians speculate that some chemotherapeutic drugs may exert their effect in part by ovarian ablation. For patients who require chemotherapy but are made amenorrheic by chemotherapeutic treatment, physicians may consider goserelin treatment unnecessary and prescribe tamoxifen alone. Mechanism of Action. Goserelin acetate is a GnRH analogue. GnRH analogues act by downregulating pituitary GnRH receptors, thereby suppressing the release of luteinizing hormone (LH) and follicle-stimulating hormone (FSH). These actions, in turn, reduce the main source of estradiol, which is produced in the ovaries, in premenopausal women. In HR-positive tumors, estrogen has been established as a promoter of cell growth and replication. Clinical Performance. Clinical trials have shown that goserelin, when used either alone or in combination with tamoxifen as an adjuvant systemic therapy in premenopausal women with ER-positive CaB, produces recurrence-free survival rates equivalent to those of chemotherapy (Robertson JF, 2003). These data demonstrate that goserelin offers an effective alternative to CMF chemotherapy for adjuvant therapy of premenopausal patients with ER-positive, node-positive early CaB. Nonpharmacological Approaches The treatment of CaB is multimodal: for the majority of patients, surgery and radiotherapy are used alongside chemotherapy and hormonal treatments. Surgery. Common surgical options for CaB are mastectomy (removal of the breast and sometimes the axillary lymph nodes) and breast-conserving surgery (BCS) (removal of the tumor and immediately surrounding tissue only, resulting in narrow margins of resection). Surgeons have also started to perform sentinel

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node biopsy during BCS to determine whether full axillary lymph node resection is necessary. More recently, the use of intraoperative ultrasound to localize nonpalpable tumors has resulted in a reduced re-excision rate due to positive margins in the excised tissue. In early-stage tumors, mastectomy and BCS have equivalent effects on survival, particularly if BCS is followed by radiotherapy. Surgical treatment for small early-stage CaB is likely to be BCS rather than mastectomy. In patients with large tumors at diagnosis, BCS may be possible if chemotherapy or hormonal therapy is given prior to surgery to shrink the tumor (neoadjuvant therapy). A minority of women at high risk of developing CaB may undergo elective bilateral radical mastectomy and, in some cases, bilateral oophrectomy. For total ovarian blockade, premenopausal women with CaB may undergo surgical oophrectomy rather than receive goserelin. Radiotherapy. CaB radiotherapy generally involves postoperative irradiation of the tumor site, chest wall, and draining lymph node regions. Its goal is the reduction of local or regional recurrence. Though toxic and timeconsuming—usually Þve days a week for six to eight weeks—postoperative radiation has been shown to reduce the risk of local recurrence. In a metaanalysis of 66 trials involving some 28,500 women, investigators found that radiation lowered the rate of local recurrence after surgery by a factor of three (EBCTCG, 1995). The effectiveness of radiation for increasing overall survival, regardless of recurrence, remains controversial. In a review of 18 randomized trials of radiation’s effect on survival, investigators found that overall survival was approximately 17% better in radiation-treated patients than in patients not receiving radiotherapy (Whelan TJ, 2000). However, in a meta-analysis of 10- and 20-year results from 40 randomized trials of radiation therapy, investigators found that although CaB mortality was signiÞcantly reduced, mortality from other causes—particularly vascular causes—increased (EBCTCG, 2000). Overall survival was statistically equivalent: 37% survival with radiation versus 36% for controls. Patient nodal status, age, and other factors strongly affected survival rates. The potential importance of these predictive factors appears particularly relevant in light of a follow-up meta-analysis, which found that radiation signiÞcantly improved overall survival of CaB patients by as much as 20% when molecular techniques were employed (Van de Steene J, 2000). EMERGING THERAPIES In the breast cancer (CaB) pharmaceutical market, the barriers to entry for new therapeutic agents are high, particularly for hormonal and cytotoxic therapies. Several classes of very well tolerated hormonal agents, administered as once-daily pills, can keep metastatic disease in check for two years or more when used in succession. When hormonal therapies and Þrst-line chemotherapy fail, several of the better-tolerated cytotoxic agents (capecitabine, vinorelbine, gemcitabine) can

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delay progression for many more months. For any new agent to gain approval in the metastatic setting, it must exhibit greater efÞcacy and/or a superior side-effect proÞle to existing agents or be relegated to third- or fourth-line use. In addition, efÞcacy is increasingly measured in terms of duration of disease control; the concept of clinical response (stable disease plus tumor response) is an important new endpoint, supplementing tumor response, disease-free survival, and overall survival. In the adjuvant market, the company developing the drug must invest in a trial recruiting more than 1,000 patients (the number required to achieve statistical signiÞcance) and then wait Þve to eight years (the amount of time required to show survival beneÞt). These conditions make entry into the CaB market time-consuming and expensive. The adjuvant market is a potentially lucrative one because of extended treatment periods and long survival times and the large number of patients involved. However, any new hormonal or cytotoxic therapy must demonstrate superiority to current therapies in both efÞcacy and toxicity proÞle. The arrival of several aromatase inhibitors (AIs) into the adjuvant market has further increased the barrier to entry for new hormonal agents. Currently, little late-stage development activity for new hormonal treatments is ongoing. One agent, TAS-108 (SRI and Tahio), is reportedly in Phase II studies in the United States and Phase I studies in Japan, with particular interest in tamoxifen-resistant disease. TAS-108 is a steroidal aromatase inhibitor under development by SRI and Taiho, for the potential Þrst-line treatment of metastatic breast cancer and secondline treatment of tamoxifen-resistant breast cancer. Data from a phase I study, showed that TAS-108 was well tolerated at doses ranging from 40 to 160 mg/day with no maximum tolerated dose achieved (Buzdar A, 2005). Toxicities associated with TAS-108 include hot ßashes, headache, and nausea and vomiting. There was evidence of antitumor activity, with stable disease noted in eight of the 16 patients enrolled, despite the fact that they were heavily pre-treated with the majority having received three or more lines of chemotherapy (63%), and three or more lines of endocrine therapy (75%). One patient had stable disease for 33 weeks and a second patient for 60 weeks. The remaining eight patients suffered progressive disease. A phase II study is ongoing in the U.S., although no data are yet available, and Phase III studies are being planned with the drug. Because biologic and other highly targeted therapies are likely to be used together with existing therapies, the barriers to market entry are lower for these agents. Clinician concerns relating to this group of drugs pertain to their burdensome cost and the potential for unusual or delayed toxicities. However, the enormous success of trastuzumab (Genentech/Roche’s Herceptin), in the treatment of metastatic HER2-positive disease and with several large ongoing trials in the adjuvant setting, has fueled enthusiasm for a targeted approach to treatment. Evaluating targeted agents in earlier disease is essential: treatment of lower tumor burden is more likely to prove successful, but research has also demonstrated that some pathways (particularly those involved in angiogenesis) may become redundant in advanced CaB.

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The complexities of hormone receptor activation and epidermal growth factor receptor (EGFR) pathways are yet to be fully elucidated. Data suggest that breast cancer growth is regulated by coordinated actions of the estrogen receptor (ER) and various growth factor receptor signaling pathways (Osborne CK, 2005) in what is termed “molecular crosstalk.” In tumors with active growth factor receptor signaling (e.g., HER2 gene ampliÞcation), tamoxifen may lose its estrogen antagonist activity and acquire more agonist-like activity, resulting in tumor growth stimulation. Indeed, clinical data support this concept of tamoxifen resistance, and many clinicians now replace tamoxifen with AIs for hormone-receptor-positive/HER2-positive patients. A signiÞcant number of novel agents in clinical development have the potential to improve treatment outcomes in the future. Herein are discussed agents that show the most promise for entry into the future CaB market. Table 8 summarizes drug therapies in development for CaB. Epothilones Overview. In the wake of taxanes’ success, epothilones, a novel class of anticancer drugs whose mechanism of action is similar to that of taxanes, are under investigation. Epothilones have several potential advantages over taxanes. Because they are soluble in water, they eliminate the need for cremophor. Cremophor is present in paclitaxel formulations to improve taxane solubility and is in part responsible for the allergic response that can occur during administration. Without the need for cremophor, epothilones are easier to administer (although still intravenous) and less likely to trigger an allergic response. Most important, they appear to be effective against cancer cells that are resistant to existing chemotherapies. Given these advantages, epothilones may challenge the place of taxanes in neoadjuvant and adjuvant therapy and in the treatment of metastatic disease. Many companies are investing signiÞcantly in epothilones and have epothilones in clinical development: Bristol-Myers Squibb’s ixabepilone is the leading agent in this class for CaB treatment, so we discuss this agent in detail. Another one, epothilone D (KOS-862), is undergoing Phase II investigation for CaB by Roche and Kosan in the United States and Europe. If epothilones are shown to be signiÞcantly more effective than taxanes, they may replace the latter class, but only if the side-effect proÞle is deemed acceptable. If epothilones exhibit efÞcacy similar to that of taxanes, have a superior side-effect proÞle, and circumvent taxane resistance, they are most likely to be used when clinicians are running out of other options. Mechanism of Action. Comprising a family of 16-member ring macrolides, epothilones were originally isolated in 1991 by Holfe and Reichenbach from the fermentation broth of the soil-derived myxobacterium Sorangium cellulosum. Epothilones work via a mechanism similar to that of taxanes: they stabilize spindle microtubules formed during mitosis, thereby preventing the

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TABLE 8. Emerging Therapies in Development for Breast Cancer Development Phase

Marketing Company

Epothilones Ixabepilone United States Europe Japan

III II —

Bristol-Myers Squibb Bristol-Myers Squibb —

Modified taxanes SA-109881 United States Europe Japan

III III —

Sanofi-Aventis Sanofi-Aventis —

Other microtubule modulators Vinflunine United States Europe Japan

— II —

— Pierre Fabre (France) —

Antimetabolites Pemetrexed (Alimta) United States Europe Japan

II II —

Eli Lilly Eli Lilly —

Cell-cycle inhibitors Temsirolimus (CCI-779) United States Europe Japan

S — —

Wyeth — —

Compound

Vascular endothelial growth factor inhibitors Bevacizumab (Avastin) United States III Europe — Japan — Epidermal growth factor receptor inhibitors Lapatinib United States III Europe — Japan — Pertuzumab (Omnitarg) United States II Europe — Japan — Erlotinib (Tarceva) United States II Europe — Japan —

Genentech (US)/Roche (EU) — —

GlaxoSmithKline — — Genentech/Roche — — OSI/Genentech/Roche — —

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TABLE 8. (continued) Development Phase

Marketing Company

Gene therapy and antisense Advexin United States Europe Japan

II — —

Introgen — —

Vaccines Theratope United States Europe Japan

S S —

Biomira Biomira —

COX-2 inhibitors Celecoxib (Celebrex) United States Europe Japan

S S —

Pfizer Pfizer —

Chemotherapy potentiators Tesmilifene United States Europe Japan

III III —

YM BioSciences YM BioSciences —

Compound

S = Suspended.

microtubules’ disassembly and consequently blocking cell division. Because their chemical structure differs from that of taxanes, epothilones can circumvent taxane resistance. Ixabepilone. Ixabepilone is a semisynthetic analogue of epothilone B being developed by Bristol-Myers Squibb. Multiple trials are evaluating ixabepilone in breast cancer treatment, including two ongoing Phase III trials in the United States. The primary end point is time to progression. The second Phase III study, study CA163-048, is evaluating ixabepilone in combination with capecitabine versus capecitabine alone in 1,200 patients. Both trials are evaluating ixabepilone in patients who have received at least one previous line of chemotherapy (including a taxane and an anthracycline). A U.S. Phase II study of ixabepilone and trastuzumab (Herceptin) in patients with HER2-positive metastatic CaB is also ongoing. In June 2004, Phase II data on ixabepilone were presented at the 40th American Society of Clinical Oncology (ASCO) meeting (Low JA, 2004). A total of 42 patients with metastatic or locally advanced CaB were administered 6 mg/m2 /day IV ixabepilone on days 1 to 5, every three weeks. Prior taxane-treated patients received a median of two prior chemotherapy regimens. Grade 3 and 4 toxicities per patient were neutropenia (32%), febrile neutropenia (10%), thrombocytopenia

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(5%), fatigue (15%), diarrhea (12%), nausea/vomiting (5%), constipation (2%), myalgia/arthralgia (2%), and sensory neuropathy (grade 3: 2%). Of the 33 patients who had prior taxane treatment, 6 (18%) achieved a partial response, 12 (40%) experienced stable disease, and a further 12 had disease progression. Of the 9 taxane-naive patients, 4 had a partial response and a further 4 had stable disease. In an earlier Phase II study, seven patients with taxane-refractory metastatic CaB were treated with ixabepilone at 50 mg/m2 in a one-hour infusion every 21 days (O’Shaughnessy J, 2002). Two subjects achieved partial responses and three achieved stable disease. Grade 3 toxicities—experienced in each case by a single patient—included fatigue, sensory neuropathy, proctitis, stomatitis, and thrombocytopenia. Two patients developed grade 3 neutropenia. In another Phase II trial involving 19 taxane-naive patients with metastatic disease, 10 patients achieved a partial response and 8 achieved stable disease; the overall disease control rate (including stabilization) was 95% (Roche H, 2002). (Although this study was small, this disease control rate compares favorably with the rate [74%] achieved with docetaxel alone.) In this group of patients, grade 3 toxicities included myalgia (Þve patients), sensory neuropathy (four), fatigue (three), dyspnea (three), diarrhea (one), and febrile neutropenia (one). Seven patients experienced grade 3 or 4 neutropenia. In another Phase II study, researchers changed ixabepilone’s dosing schedule to reduce neurotoxicity: patients received a three-hour (rather than a one-hour) infusion every 21 days (Thomas E, 2003). The researchers also reduced the dose from 50 mg/m2 to 40 mg/m2 to address the incidence and severity of mucositis and abdominal pain reported in other studies. In June 2003, at the ASCO meeting, researchers presented preliminary efÞcacy data from this study, which involved anthracycline-pretreated patients. Of the 44 evaluable patients, they determined 15 partial responses and 23 cases of stable disease. The incidence of grade 3 neuropathy fell from 37% to 16%. Grade 3 neutropenia was observed in 24% of the patients. Clinical investigations of ixabepilone are also underway in patients previously treated with anthracyclines. Novel and Modified Taxanes Overview. Paclitaxel has been a highly successful agent for the treatment of a variety of solid tumors. As a result, much interest has been shown in developing new formulations and analogues to address problems with its administration. The goal is to develop more-soluble, cremophor-free formulations and analogues that enable clinicians to administer higher doses of paclitaxel for greater efÞcacy while reducing the associated toxicity. These agents may also have an improved pharmacokinetic proÞle and different mechanism of intracellular uptake. The commercial potential of these agents has recently been demonstrated with the launch of American Pharmaceutical Partners’ nanoparticle formulation of paclitaxel, ABI-007 (Abraxane). With improved therapeutic indices, modiÞed taxanes could encourage longer term maintenance use and avert toxicity concerns with available taxanes, that is, unless their cost is prohibitive.

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Mechanism of Action. Taxanes suppress microtubule dynamics and disassembly, thereby stopping mitosis at the metaphase/anaphase transition and inducing cell death. Formulation. Companies are investigating a variety of approaches to modifying taxanes. These include the following: •

• • • • •

New paclitaxel (Bristol-Myers Squibb’s Taxol, generics) analogues (BristolMyers Squibb’s BMS-184476, in Phase II evaluation for CaB and other cancer indications); Oral formulations (Ivax’s Paxoral); Liposomal formulations (PÞzer/NeoPharm’s LEP); Polyglutamated formulations (Cell Therapeutics’ CT-2103; Xyotax); Polyethylene glycol–conjugated (pegylated) formulations (Enzon’s MacroMed); and Microsphere formulations (Guilford Pharmaceuticals’ Paclimer).

Many other agents are in clinical trials for CaB. We focus our discussion on the most promising agent, SanoÞ-Aventis’s SA-109881. SA-109881. SanoÞ-Aventis is developing SA-109881, an orally and intravenously active taxoid. A Phase III trial is ongoing in the United States and Europe. As of February 2004, prior to its merger with SanoÞ-Synth´elabo, Aventis was also planning to implement a Phase II screening program during 2004 to identify the tumors most sensitive to SA-109881. The Phase III trial is evaluating SA-109881 versus capecitabine as a Þrst-line treatment of metastatic CaB patients. SA-109881 will be given intravenously once every three weeks. A Phase II U.S. study is evaluating SA-109881 in chemotherapy-refractory metastatic CaB patients. Encouraging data from a Phase II trial in patients with metastatic CaB were disclosed at the 39th ASCO meeting in 2003 (Dieras VC, 2003). Patients previously treated by chemotherapy were stratiÞed into taxoid-responsive and taxoidresistant arms. At the time data were disclosed, 42 patients (21 in each arm) of the 58 enrolled had been evaluated for safety and efÞcacy. The median time between the last administration of the previous taxoids and the Þrst SA-109881 infusion was 13.6 months and 1.4 months in arms 1 and 2, respectively. A total of 175 cycles of SA-109881 were administered, with a median of 6 cycles and 2 cycles in arms 1 and 2, respectively. The safety proÞle was similar in both arms. Neutropenia was the main hematologic toxicity (59%; grade 3/4). Grade 2 toxicities were as follows: diarrhea (9%), neurosensory disorders (13%), fatigue (17%), myalgia (6%), arthralgia (6%), nausea (7%). Thus far, no cumulative toxicity or ßuid retention—a problematic side effect associated with established taxanes—have been reported. Preliminary efÞcacy results show partial response rates of 48% and 29% in the Þrst and second arms, respectively.

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These Phase II data are encouraging, especially for the Þrst-arm, given that docetaxel achieves approximately a 50% response rate as a Þrst-line treatment. If the ongoing Phase III trial results are positive, SA-109881 has potential to become a second-line treatment. Other Microtubule Modulators Overview. The success of the taxanes in CaB treatment has spurred R&D interest in many other microtubule inhibitors that do not fall within the other classes outlined here. These agents, which include Kosan Biosciences’ KOS-862, are as yet unproven in CaB because data are lacking. The only exception is Pierre Fabre/Bristol-Myers Squibb’s vinßunine (Javlor), an agent covered in detail in this section. Mechanism of Action. These agents work in a similar way to taxanes; they promote microtubule polymerization and inhibit tubulin depolymerization, arresting mitotic cell division at the metaphase/anaphase transition and inducing cell death. Vinca alkaloids inhibit microtubule polymerization at high drug concentrations. Vinflunine. Vinßunine (Pierre Fabre’s Javlor) (Figure 19) is a semisynthetic vinca alkaloid derived from vinorelbine. It is in Phase II clinical trials for the treatment of CaB in Europe and in other, non-major-market countries. Phase II data on vinßunine in metastatic CaB patients progressing after prior chemotherapy with anthracyclines and taxanes were presented at the 2004 ASCO meeting (Fumoleau P, 2004). Of the 60 patients enrolled, 45 were evaluable for efÞcacy, and 52 were evaluable for safety. Dose reduction was required in 11.4% of cycles. The relative dose intensity was 96.5%. Partial response (PR) was obtained in 16 out of 45 patients (35.6%), and stable disease (SD) in 15 patients (33.3%). By an intent-to-treat analysis, median progression-free survival was 4.2 months. From 16 out of 45 patients with a progression-free interval less than three months after taxane exposure, 7 patients (43.8%) achieved a PR. Grade 3/4

N

CF2CH3

N N H CH3O2C CH3O

CH3 N

OCOCH3 HO CO2CH3 CH3

FIGURE 19. Structure of vinflunine.

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related toxicities were neutropenia (63.5% patients), neutropenic infection (5.8% patients), constipation (19.2% patients), abdominal pain (13.5% patients), nausea (9.6% patients), and vomiting (7.7% patients). No grade 3/4 related peripheral neuropathy was observed. Although vinßunine has shown some promising results in the aforementioned Phase II trial, Phase III data are needed. The cytotoxic CaB market is already well established, so any therapy entering this market must show a signiÞcant advantage over standard treatments. Antimetabolites Overview. Although cytotoxic anticancer drugs are collectively the largest and most-established chemotherapy group, the agents in development have relatively poor clinical and commercial potential. A possible exception is pemetrexed (Eli Lilly’s Alimta). Mechanism of Action. Antimetabolites are incorporated into new nuclear material or combine irreversibly with vital cellular enzymes, such as those on the folate pathway, thereby preventing normal cellular division. The folate pathway is essential, by its provision of cofactors, to the de novo synthesis of DNA precursors (purines); compounds that inhibit this pathway interfere with DNA synthesis, thus causing tumor cell death. Previous generations of antifolates principally targeted a single pathway. For example, the antimetabolite/antifolate methotrexate inhibits dihydrofolate reductase, a key enzyme in the folate pathway, and 5-ßuorouracil (5-FU), combined with folic acid and raltetrexed (AstraZeneca’s Tomudex), inhibits thymidylate synthase, which is essential for de novo thymidine synthesis. Pemetrexed. Eli Lilly is developing a multitargeted antifolate/antimetabolite compound called pemetrexed (Alimta) (Figure 20). The compound exerts its antitumor activity by inhibiting multiple key enzymes involved in DNA synthesis: thymidylate synthase, dihydrofolate reductase, and glycinamide ribonucleotide (GAR) formyltransferase. The company hopes pemetrexed’s multitargeted action will overcome the resistance acquired through the overexpression of any single enzyme. Pemetrexed was approved initially for malignant pleural mesothelioma in the United States in February 2004 and has since been approved in the United States and Europe as a second-line therapy for non-small-cell lung cancer (NSCLC). Pemetrexed is in Phase II trials for CaB in the United States and Europe. Phase II data on pemetrexed in combination with gemcitabine (Eli Lilly’s Gemzar) were disclosed at the 2004 ASCO meeting (Ma CX, 2004). Eligible patients must have been treated with both an anthracycline and a taxane, in either the adjuvant or metastatic setting, and with no more than one chemotherapy regimen for metastatic disease (unless a taxane and anthracycline). Fifty-nine patients were enrolled, with a median of 0–1 prior adjuvant chemotherapy regimens (0 = 8%; 1 = 92%) and 0–2 prior metastatic chemotherapy regimens

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FIGURE 20. Structure of pemetrexed.

(0 = 39%; 1 = 59%; 2 = 2%). Twenty-eight patients (48%) had also received prior hormonal therapy. Patients received a median of Þve cycles with a median follow-up of 426 days. Considering grade 3 and 4 toxicities, 48 (81%) patients had neutropenia (grade 3 = 11, grade 4 = 37), although only 7 (12%) had febrile neutropenia. Fifteen patients (25%) had thrombocytopenia (grade 3 = 13, grade 4 = 2). Grade 3 or 4 nonhematologic toxicities included 10 (17%) patients with dyspnea, 10 (17%) with fatigue, 4 (7%) with rash, 3 (5%) with nausea, and 3 (5%) with anorexia. Overall, 14 (24%) patients achieved a partial response with a median duration of at least Þve months. Nine (15%) had stable disease for six months or more, with a median of 334 days. Median time to progression was 3.8 months. Median survival was 10.3 months, and the one-year survival rate was 49%. Results of a Phase II study of single-agent pemetrexed in the neoadjuvant setting in patients with locally advanced disease were disclosed at the 2002 ASCO meeting (Gomez HL, 2002). Data were available on 59 of 61 patients: 20 patients achieved a partial response, for a preliminary overall response rate of 34%; 32 patients had stable disease; 6 patients had disease progression; and 1 patient was lost to follow-up. Based on these results, pemetrexed would not be able to compete with an anthracycline followed by a taxane regimen—a combination that elicits better response rates. A Phase II study evaluating the efÞcacy of single-agent pemetrexed as thirdline therapy in patients with metastatic CaB showed promising results in this poorprognosis group (Mennel RG, 2001). Fifty-eight patients received 500 mg/m2 pemetrexed by intravenous infusion for ten minutes every 21 days. Seventy-two percent of the patients had visceral metastases. No dose reductions or omissions were necessary in a total of 169 cycles administered; the median number of cycles was two (range = 1–18). Analysis of 42 evaluable patients revealed four partial responses with a median duration of 5.9 months. Thirty-eight percent of patients experienced stable disease with a median duration of 4.5 months. Of the patients who received vitamin supplements (folic acid and B12 ) to prevent toxicity, 10%, 30%, and 14% experienced grade 3 or 4 leukopenia, neutropenia, and elevated transaminases, respectively. A second Phase II trial involved 38 patients with locally recurrent or metastatic CaB who were treated with a higher dose of pemetrexed (600 mg/m2 every 21 days) (Miles DW, 2001). Thirty-three patients had had prior chemotherapy:

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16 adjuvant, 12 metastatic, and 5 in both settings. This trial achieved a higher response rate but at the expense of greater toxicity. The overall response rate was 28%, including one complete response. The median duration of response was 8 months, and median survival was 13 months. Of the patients, 47% suffered grade 3 or 4 neutropenia, and 16% suffered grade 3 or 4 thrombocytopenia. Nonhematologic toxicities (grades 2 or 3) included elevated transaminases (92%), vomiting (34%), and mucositis (32%). Pemetrexed could ultimately replace methotrexate in many neoadjuvant and adjuvant regimens in earlier disease. Given the drug’s signiÞcant toxicity and lack of advantage over existing agents, the agent may Þrst be approved as thirdor fourth-line therapy for advanced CaB. Cell-Cycle Inhibitors Overview. Researchers have identiÞed novel components of cell-cycle regulation as targets for chemotherapeutic intervention. Within this class are agents at all stages of development, including product failures. One failure was RO-317453, a novel oral cell-cycle inhibitor with activity against a wide range of human tumor cell lines and xenografts, including breast cancer and drug-resistant models. RO317453 causes cells to arrest in the pro-metaphase, disrupting mitotic spindle formation and apoptosis. In June 2003, at ASCO’s annual meeting, researchers presented data showing that RO-317453 was well tolerated but had limited clinical activity in patients with metastatic CaB who had failed on either taxane or anthracycline therapy (Trigo Perez JM, 2003). Roche discontinued work with this agent in September 2002. Roscovitine (Cyclacel’s CYC-202) is a cyclin-dependent kinase (CDK) inhibitor. In January 2003, Cyclacel initiated a Phase IIa clinical trial of roscovitine in combination with capecitabine in patients with advanced CaB. The study is evaluating the compound’s safety, pharmacokinetic proÞle, antitumor activity, and effect on biomarkers. However preclinical data show this compound to have low potency and unfavorable bioavailability. The agent that is the most advanced in development and that may propel this class into more clinical trials is Wyeth’s temsirolimus. We discuss this agent in more detail further on. Mechanism of Action. Progression through the cell cycle depends on numerous signaling pathways and checkpoints (e.g., the cyclin families of proteins, the CDKs). The deregulation of these key regulators of the cell cycle is observed in most cancer cells. The inhibition of these enzymes can result in cell-cycle arrest and, ultimately, cell death, usually by apoptosis. Temsirolimus. Wyeth’s temsirolimus (CCI-779) is an ester analogue of rapamycin with improved aqueous solubility and pharmacokinetic properties. Temsirolimus selectively inhibits the mammalian target of rapamycin (known as mTOR), a serine/threonine kinase, by binding to the cytosolic (FK506 binding)

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protein (FKBP), which subsequently inhibits mTOR. The inhibition of mTOR, a downstream mediator in the PI3K/Akt signaling pathway, blocks a number of signal transduction pathways that suppress translation of several key proteins regulating the cell cycle. These effects lead to a cell-cycle block at the G1 phase, preventing progression to the S phase. In breast cancer, the PI3K/Akt signaling pathway can be activated by membrane receptors, including the HER (or ERBB) family of growth factor receptors, the insulin-like growth factor receptor (IGF-R), and the estrogen receptor (ER), revealing the potential for synergy with other agents. In March 2002, based on promising clinical data, the FDA designated temsirolimus for fast-track development for second-line treatment of renal cell carcinoma (RCC). Temsirolimus is undergoing a U.S. Phase III trial for metastatic CaB. The U.S. multicenter, randomized, double-blind, placebo-controlled Phase III trial under way is examining 30 mg temsirolimus administered in combination with letrozole versus letrozole alone. The study is recruiting postmenopausal women with locally advanced or metastatic CaB. The study is estimated to last for 34 months, and all subjects will have the option of participating in the long-term follow-up phase of the trial that involves follow-up every three months until disease progression; the primary endpoint is overall progressionfree survival. Phase II data were presented at ASCO 2004 (Baselga J, 2004). The trial consisted of three arms of approximately 30 evaluable patients per arm: letrozole alone, letrozole with temsirolimus 25 mg daily (daily arm), or letrozole with temsirolimus 75 mg daily for Þve days every two weeks (intermittent arm). All patients received 2.5 mg letrozole daily. At the time of composing this reference, 55 patients had been enrolled. Initially, 6 patients had been enrolled on each of these two “high-dose” (HD) schedules, but 3 patients in each arm had toxicity that resulted in dose delay/reduction or discontinuation. The protocol was amended, and doses were reduced to “low-dose” (LD) schedules: 10 mg temsirolimus daily and 30 mg temsirolimus intermittently. The most frequently occurring grade 3 and 4 temsirolimus-related toxicities were stomatitis for the HD schedules (2/6 patients, 2/6 patients) and diarrhea for the LD schedules (0 patients, 1/7 patients). No grade 3 or 4 toxicities were reported in the letrozole alone arm. Of 55 patients, 7 had been on study for more than 40 weeks. Preliminary tumor responses are available for 19 evaluable patients. Temsirolimus patients (n = 13) had 1 CR (HD schedule), 3 PRs (HD schedule), 9 SDs (6 on HD schedule, 3 on LD schedule; SD at 24 weeks or beyond for 4 patients on HD schedule). Letrozole-alone patients (n = 6) had 2 PRs and 4 SDs (SD at 24 weeks or beyond for 1 patient). Phase II data in patients with stage III/IV CaB unresponsive to taxane- or anthracycline-based chemotherapy have been released (Chan S, 2003). Temsirolimus, at 75 mg or 250 mg, was administered weekly as a 30-minute IV infusion to 34 patients. Researchers noted two objective responses in measurable

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liver metastases in the Þrst 16 evaluable patients. Two patients in the study suffered grade 3 or 4 events, and at least 30% reported grade 1 or 2 events, which often resolved without discontinuing temsirolimus. In future, agents such as temsirolimus could be approved in combination with hormonal treatments. Currently, hormonal agents are not combined with chemotherapy or targeted agents because such combinations have appeared detrimental in terms of efÞcacy and toxicity. Vascular Endothelial Growth Factor Inhibitors Overview. Angiogenesis, the formation of new blood vessels, plays a major role in many normal physiological processes and in several pathological conditions, including solid tumor growth and metastasis. Investigators have detected vascular endothelial growth factor (VEGF) expression in tumor samples from 90% of CaB patients that involve four or more nodes (n = 124) (Gown A, 2001). VEGF expression was graded 3 or 4 in 36% of patients. Further, Þve-year overall survival was signiÞcantly lower among patients with high VEGF expression: 44% versus 67%. VEGF inhibition is an area of considerable commercial interest in the pharmaceutical industry. Agents within this class in early-phase development for CaB include AstraZeneca’s ZD-6474, EntreMed’s 2-methoxyestradiol (2-ME2), and Bayer’s sorafenib (BAY-43-9006). These agents are not included in our discussion because the data are still insufÞcient. Bevacizumab is covered in detail because it is the only VEGF inhibitor that has reached Phase III trials for CaB. Mechanism of Action. VEGF is a multifunctional cytokine and potent permeability factor that is secreted in response to hypoxia (reduced oxygen). The biological effects of VEGF are mediated by the binding of VEGF to one of three endothelial surface receptors: VEGF-R1 (ßt-1), VEGF-R2 (ßk-1/kdr), or VEGFR3; binding to the coreceptor neurophilin enhances signaling. VEGF has a major angiogenesis-promoting effect. Researchers hope that blocking this receptor will, in the best-case scenario, kill the solid tumor that depends on new blood vessel formulation for its survival or at least slow tumor growth by preventing new vessel formation and thus limiting the supply of blood. Bevacizumab. Bevacizumab (Genentech’s/Roche/Chugai’s Avastin) is a humanized monoclonal antibody (MAb) that prevents VEGF from binding to its receptors. Its lead indication is colorectal cancer, for which bevacizumab gained U.S. and EU marketing approval in February 2004 and January 2005, respectively. Bevacizumab is also in development for numerous other cancers; Phase III trials are ongoing in non-small-cell lung, renal cell, ovarian, pancreatic, and breast cancer. Numerous Phase II and III trials for bevacizumab in CaB are underway, including combinations with docetaxel with or without capecitabine in locally advanced or metastatic CaB, or with erlotinib in metastatic CaB.

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The trial that led to bevacizumab’s U.S. approval for colorectal cancer involved more than 900 patients with metastatic CRC. Researchers found that the adding bevacizumab to a standard-care regimen (5-FU/leucovorin/irinotecan) produced a statistically signiÞcant prolongation of time to disease progression (6.2 versus 10.6 months) and increased survival time by about Þve months (15.6 versus 20.3 months) (Hurwitz H, 2003).These Þndings exceeded the expectations of the study design. Trials have also shown that bevacizumab is active in previously treated patients with metastatic CaB (Hillan K, 2003). A Phase III, 462-patient trial of bevacizumab in patients with refractory metastatic CaB compared a combination of bevacizumab and capecitabine with capecitabine alone. Although no improvement in progression-free survival (the primary endpoint) was observed, a statistically signiÞcant increase in the secondary endpoint—objective response rate—was revealed (9.1% versus 19.8% by independent review; 19.1% versus 30.2% by investigator assessment). The adverse event proÞle in this study was similar to the observed events in Phase II studies and included venous thrombosis, hypertension, and asymptomatic proteinuria. No increase in serious bleeding occurred in the bevacizumab arm of the study. An ongoing Phase III trial is investigating the efÞcacy of bevacizumab combined with paclitaxel as Þrst-line treatment for locally recurrent or metastatic CaB. In April 2005, Genentech/Roche reported that a preliminary efÞcacy analysis of the study data was positive (Genentech/Roche, press release, April 2005). At that point, 722 patients were evaluated. No safety data were disclosed, although the companies reported that “adverse events appeared to be similar to those observed in previous clinical trial” and that bevacizumab-related events “included neuropathy, hypertension, and proteinuria. Serious bleeding and blood clots were rare in this study.” Preliminary Phase II data of bevacizumab in combination with erlotinib were disclosed at the ASCO meeting in 2004 (Dickler M, 2004). Erlotinib (150 mg orally/day) and bevacizumab (15 mg/kg IV every 3 weeks) was given to patients with metastatic CaB following one to two prior chemotherapy regimens. The primary endpoint was response rate; secondary end points included safety and time to tumor progression. Of the 13 patients enrolled at the time, 9 were evaluable for response. One patient had a conÞrmed PR (11%), 2 had SD at nine weeks (22%), 5 had PD (56%), and 1 (11%) withdrew from the study after one cycle because of an allergic drug eruption. Two patients (15%) experienced grade 4 toxicities: pulmonary embolus 1 (8%) and neutropenia 1 (8%). Toxicity potentially related to erlotinib and/or bevacizumab included rash (grade 2, 46%), diarrhea (grade 2, 23%), hypertension (grade 3, 15%), and nausea/vomiting (grade 3,: 8%). Phase II data of bevacizumab in combination with docetaxel were also presented at ASCO 2004, but the data were preliminary and efÞcacy data were not available (Overmoyer B, 2004). In a Phase II, monotherapy, dose-escalation study, 75 patients with previously treated metastatic CaB (median = 2 regimens) were treated with bevacizumab—3 mg/kg (18 patients), 10 mg/kg (41 patients), and 20 mg/kg (16

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patients)—once every two weeks (Cobleigh MA, 2001). At 22 weeks, 17% of the patients were responding or had stable disease; after 12 months, three patients were progression-free. Bevacizumab was well tolerated. No bleeding was reported in this trial, though 7% of patients developed thrombosis, 22% developed mild hypertension, and 15% developed proteinuria. It remains to be seen whether the mortality risk associated with these complications will deter clinicians from prescribing them to CaB patients who already have a good treatment outcome from older therapies. Epidermal Growth Factor Receptor Inhibitors Overview. The ERBB tyrosine kinase family includes four related transmembrane receptors: erbB1, also known as HER1 or EGFR; erbB2 (HER2); erbB3 (HER3); and erbB4 (HER4). Each receptor contains an extracellular binding region and an intracellular kinase. The HER3 kinase is defective; thus, signaling through HER3 requires heterodimerization with another EGFR family member. HER2 has no known natural ligand and, thus, also typically requires heterodimerization. However, at high concentrations, HER2 homodimers may signal even in the absence of ligand. Ligand binding results in homo- or heterodimer formation with activation of the associated receptor tyrosine kinase. Receptor activation triggers multiple intracellular pathways, including the mitogen-activated protein kinase (MAPK) and AKT/PI3K, that result in increased proliferation, resistance to apoptosis, and increased angiogenesis. Overexpression of HER2 proteins, which occurs in 22–28% of breast tumors, is associated with poor prognosis in CaB patients. Estimates of the prevalence of EGFR overexpression in breast tumors vary widely; the American Cancer Society estimates prevalence at 14–91%. Agents that target HER2 include the MAb trastuzumab and the EGFR tyrosine kinase inhibitors (described further on in this chapter). Vaccine approaches targeting HER2 are also the subject of investigation, as described later. Many EGFR inhibitors, especially EGFR1, have not shown promise in CaB treatment. Nevertheless, this class is of immense scientiÞc and commercial interest because of the success of trastuzumab and other agents in other cancer indications. Other agents being investigated for CaB include Takeda’s small-molecule inhibitor of HER2 tyrosine kinase, TAK-165. One more agent of considerable, yet controversial, interest is AstraZeneca’s EGFR1 inhibitor, geÞtinib (Iressa). The controversy surrounds postmarketing efÞcacy data in NSCLC that have failed to show improved survival. GeÞtinib is still in development for CaB. Mechanism of Action. EGFR is part of the ERBB family of receptors, which consists of an extracellular ligand-binding domain, a transmembrane domain, and an intracellular tyrosine kinase domain. Binding to the receptor results in tyrosine kinase activity and receptor autophosphylation which then initiate signal transduction cascades implicated in cell proliferation. Drug intervention, either at the EGFR or the intracellular tyrosine kinase, blocks cell signaling pathways, thereby preventing proliferation and subsequent tumor growth.

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Lapatinib. GlaxoSmithKline’s lapatinib is an orally administered EGFR tyrosine kinase inhibitor that has the added beneÞt of blocking ErbB −2/HER2 tyrosine kinase. Because of lapatanib’s dual-acting properties, the agent might be used in a wider patient population. Lapatinib is undergoing two Phase III trials for CaB treatment in the United States. One trial is comparing capecitabine with or without lapatinib in refractory locally advanced or metastatic CaB. A total of 372 patients (186 per treatment arm) will be accrued for this study within 17 months. The primary endpoint is time to disease progression. The second Phase III trial is examining paclitaxel with or without lapatinib as a Þrst-line therapy for patients with advanced or metastatic CaB. The primary end point is time to disease progression. A total of 570 patients are planned for enrollment. Preliminary Phase II data were disclosed at the 2004 ASCO meeting (Blackwell KL, 2004). The primary objectives of this study were to assess the safety and efÞcacy of monotherapy Lapatinib at 1,500 mg daily in women with trastuzumabrefractory metastatic CaB. Eligible patients had ErbB2 overexpressing CaB with disease progression following either one or two prior trastuzumab-containing regimens. Preliminary efÞcacy results regarding the initial 36 patients include 3 PRs (8+ weeks, 16+ weeks, and 44 weeks) and 5 SDs (range = 12 − 36 + weeks). For 15 patients with preliminary safety data, adverse events considered by investigators to be drug-related were all grade 1–2 except for one grade 3 rash. For these 15 patients, drug-related adverse events observed in more than one patient were anorexia, nausea, rash, vomiting, diarrhea, and weight loss. Updated data were presented in December 2004 at the 27th Annual San Antonio Breast Cancer Symposium (Blackwell KL, 2004). At that time, 48 women had been enrolled. A total of 4 women responded to therapy, including 1 woman who achieved a complete response; 7 women were progression-free at 24 weeks. Preliminary Phase II data were also disclosed at the 2003 ASCO meeting but were available for only three patients (Kaplan EH, 2003). Patients with disease progression after one or two prior treatments with trastuzumab alone or in a combination chemotherapy regimen were administered lapatinib (1,250 mg daily). Inclusion criteria were stage IIIb or IV CaB, conÞrmation of erbB-2 overexpression prior to Þrst-line trastuzumab, Karnofsky performance status greater than 70, and normal left ventricular ejection fraction (LVEF). Preliminary side-effect data included reports of rash and diarrhea. No cardiac toxicity was reported. Pertuzumab. Genentech/Roche/Chugai’s next-generation HER2-directed MAb, pertuzumab (Omnitarg), inhibits HER2 dimerization. It is in Phase II clinical trials for a range of solid cancers, including CaB. A Phase III study is planned, although not yet recruiting, that will be a multicenter, open-label extension study available only to patients who have completed 17 cycles (approximately one year) of treatment with single-agent pertuzumab in a Genentech-sponsored Phase II cancer study. Eligible patients must have shown no signs of unacceptable pertuzumabrelated toxicity in the previous trial. Genentech claims that pertuzumab is the Þrst in a new class of targeted potential therapeutic agents known as HER dimerization inhibitors (HDIs). Such agents

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may have application in non-HER2-overexpressing cancers because binding of the antibody to the dimerization domain of HER2 directly inhibits the ability of HER2 (the most common pairing partner) to dimerize with other HER receptor proteins. Inhibiting receptor dimerization prevents the activation of HER signaling pathways. Only Phase I data have been reported so far. Twenty-one patients with locally advanced, recurrent, or metastatic solid tumors who had progressed during or after standard therapy were recruited to a dose-escalating Phase I study. Three patients had CaB. Nineteen patients received at least two cycles of pertuzumab at doses ranging from 0.5 mg/kg to 15 mg/kg. Pertuzumab was generally well tolerated, and a maximum tolerated dose was not reached in the study. The most common adverse events were asthenia (62%), vomiting (52%), nausea (48%), rash (43%), diarrhea (43%), and anemia (33%). The majority of cases were grade 1 or 2 in severity. However, after cycle 2, one patient suffered a myocardial infarction resulting in grade 4 left ventricular failure. Because of the cardiotoxicities associated with trastuzumab, the authors concluded that this may be a class effect, and great care would be needed throughout the clinical development of this agent. Two partial responses were seen in this heavily pretreated population—one in ovarian cancer and one in pancreatic cancer. Interestingly, neither were in patients whose tumors overexpressed HER2 (as measured by ßuorescent in-situ hybridization [FISH]). One unconÞrmed disease regression was noted in a patient with prostate cancer. Stable disease (2.5–5.5 months) was observed in a further six patients. Pharmacokinetic analysis showed that, in line with other antibody therapies, three-times-weekly administration of pertuzumab would be optimal. Pertuzumab’s novel mechanism of action will add insight to this complex area of molecular cell biology. Were it to demonstrate efÞcacy in an HER2-negative CaB population (approximately 75% of patients), it would have wide application in this indication. Erlotinib. Erlotinib (OSI-774/CP-358774/Tarceva), another EGFR tyrosine kinase inhibitor, is under development by OSI Pharmaceuticals in alliance with Genentech and Roche. This orally active quinazoline derivative is undergoing Phase II trials in the United States. Phase II trials being carried out on erlotinib include combinations with trastuzumab as a Þrst-line therapy in metastatic CaB associated with HER2/neu overexpression and with bevacizumab-treated metastatic CaB patients previously treated with one or two lines of therapy. Data disclosed at the ASCO meeting in 2003 revealed that an erlotinib/docetaxel/capecitabine combination was well tolerated (Bruno R, 2003). Nine of the 23 patients treated to date had conÞrmed partial responses. A further 4 patients had unconÞrmed partial responses, and there were 2 unconÞrmed complete responses. Preliminary Phase II data of erlotinib in combination with bevacizumab were disclosed at the ASCO meeting in 2004 (Dickler M, 2004). Erlotinib (150 mg orally/day) and bevacizumab (15 mg/kg IV every 3 weeks) was given to patients

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with metastatic CaB following one or two prior chemotherapy regimens. The primary endpoint was response rate; secondary end points included safety and time to tumor progression. Of the 13 patients enrolled at the time, 9 were evaluable for response. One patient had a conÞrmed PR (11%), 2 had SD at 9 weeks (22%), 5 had PD (56%), and 1 patient (11%) came off study after one cycle because of an allergic drug eruption. Two patients (15%) experienced grade 4 toxicities: pulmonary embolus (8%) and neutropenia (8%). Toxicity potentially related to erlotinib and/or bevacizumab included rash (grade 2, 46%), diarrhea (grade 2, 23%), hypertension (grade 3, 15%), and nausea/vomiting (grade 3, 8%). Gene Therapy and Antisense Overview. Researchers are keenly interested in the theoretical rationale for using gene therapy and antisense approaches to treat cancer, but these approaches have been studied for years without any signiÞcant progress. Despite anecdotal reports of therapeutic responses in some patients, there is still no unequivocal proof of clinical efÞcacy. The primary obstacle appears to be the transcription rates of the gene in vivo and the gene’s delivery to tumor sites. Mechanism of Action. The primary approach to gene therapy is to either replace or activate aberrant tumor-suppressor genes or to inactivate oncogenes by inserting the gene or antisense DNA using a vector (typically an attenuated virus such as an adenovirus). Advexin. Introgen’s Advexin is an adenoviral p53 gene therapy for the treatment of multiple tumors. In about 50% of cases of aggressive CaB, the p53 gene is mutated so that protein is no longer available in sufÞcient amounts to control cell growth. Loss of p53 is associated with more aggressive and resistant tumors, as well as early metastasis and reduced survival rates. Advexin is undergoing Phase II trials in the United States as a treatment for CaB. At ASCO in 2003, researchers presented preliminary Phase II data from nine patients with locally advanced CaB (Cristofanilli M, 2003). Given by intratumoral injection, Advexin was shown to be safely combined with a two-drug standard chemotherapy regimen, doxorubicin and docetaxel. Ninety percent of the patients had major responses to the therapy. Additional data from the trial were presented at the 2004 San Antonio Breast Cancer Symposium (Cristofanilli M, 2004). The addition of Advexin to the doxorubicin/docetaxel combination achieved an average of nearly 80% reduction in tumor size and an average of nearly 70% reduction in lymph node tumor size in all 12 patients with locally advanced breast tumors. Overall disease-free survival at 25 months was 83%, and overall survival at 25 months was 92%. Patients who received Advexin and chemotherapy achieved a greater reduction in tumor size than did those who were treated with chemotherapy alone. Despite this apparent success, for gene therapy to be considered a therapeutic option, randomized trials must be conducted. They will need to compare regimens

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that include gene therapy with standard chemotherapy regimens to determine whether the gene therapy actually contributes to the regimen’s efÞcacy. Vaccines Overview. Although the concept of tumor vaccines is a viable one, these therapies remain unproven in the treatment of cancer. Even if research proves that vaccines possess activity similar to that of cytotoxic agents, their uptake may be hindered by their high costs and reimbursement problems. Biomira/Merck KGaA’s Theratope (STn-KLH) is the most clinically advanced vaccine for CaB. Other vaccines include Pharmexa’s HER2 protein vaccine, AutoVac (PX-104.1), which is undergoing Phase II trials for metastatic CaB. Mechanism of Action. Cancer vaccines are designed to stimulate the immune system to launch a response against the speciÞc target contained by the vaccine. In general, research has shown that the most effective antitumor immune responses are achieved by stimulating T cells, which can recognize and kill tumor cells directly. Theratope. Biomira’s Theratope underwent a Phase III international trial in collaboration with Merck KGaA and Chiron, but both of these latter companies are no longer involved. Merck KGaA decided to discontinue its involvement with theratope in 2004 because additional trials were likely to be required to support registration. Biomira is seeking a collaborative partner prior to further development of theratope. Theratope comprises a synthetic Sialyl-Tn (STn) antigen linked to keyhole limpet hemocyanin (KLH) protein carrier; it is administered with Corixa’s DetoxB-adjuvant. In 2000, the FDA granted theratope fast-track status as an adjunct to Þrst-line combination therapy for metastatic CaB. The working hypothesis for this vaccine is that it stimulates an immune response to the tumor-associated STn epitope, thereby producing an anticancer effect. The STn epitope, expressed as part of the mucin-1 (MUC-1) glycoprotein, is associated with most types of adenocarcinoma. Although STn is expressed in some normal tissues, scientists believe its expression to be relatively tumorspeciÞc. A randomized Phase III trial of theratope followed up 1,030 women with metastatic CaB at 120 sites in the United States and Europe who had no evidence of disease progression following Þrst-line chemotherapy. Patients received intravenous low-dose cyclophosphamide on day 3, followed by four subcutaneous injections of either theratope or a control vaccine at 0, 2, 5, and 9 weeks. The control vaccine differed from theratope in not having the active STn cancer-associated antigen—it had the protein carrier. The women also received immunomodulatory low-dose cyclophosphamide to inhibit the suppressor T-cell response induced by shed cancer mucins. Data released in June 2003 and at the 2004 ASCO meeting showed that theratope had not met its primary

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end points: time to disease progression and survival (Biomira, press release, June 2003; Ibrahim NK, 2004). However, the data did show that one subset of patients—women on hormonal treatment followed by chemotherapy—appeared to have a favorable trend toward improvement in survival. Further analyses of these data are under way. Data on subset analysis of this Phase III trial were presented at the ASCO meeting in 2004 (Ibrahim NK, 2004). Of a planned accrual of 950 patients, 1,030 were enrolled and 1,028 randomized to receive either adjuvant chemotherapy plus Theratope (n = 523) or placebo (n = 505). Median duration of follow-up was 22.2 months. Median overall survival (MOS) was signiÞcantly greater in theratope-treated patients who received concurrent hormone therapy and who had high immune responses to naturally occurring clustered STn versus low immune responses (41.1 versus 25.4 months). MOS for patients in the hormonal subset without stratiÞcation into low versus high immunoresponse to naturally occurring clustered STn (n = 350) showed a statistically signiÞcant difference between the two treatment arms: women in the theratope arm (n = 180) survived a median of 36.5 months, while those in the control vaccine arm (n = 170) survived a median 30.7 months. Survival for women not receiving hormonal therapy did not differ signiÞcantly between the two treatment arms. Cyclooxygenase-2 Inhibitors Overview. Cyclooxygenase-2 (COX-2) is involved in carcinogenesis, tumor growth, and metastasis, and it is often overexpressed in premalignant and malignant lesions. Indeed, studies in CaB patients have shown that COX-2 expression is associated with a substantially worse clinical outcome (Ragaz J, 2003). Preclinical studies provided solid evidence that inhibiting this enzyme with selective COX-2 inhibitors could prevent carcinogenesis, slow the growth of established tumors, and enhance tumor response to radiation and chemotherapeutic agents without appreciably affecting normal tissue response to radiotherapy. PÞzer’s SC-236 is under preclinical development and has been studied in vitro in HER2 /neu-transfected breast tumor cell lines, alone and in combination with other agents. Researchers concluded from this work that a tyrosine kinase inhibitor that blocked HER2 /neu and EGFR in combination with a COX-2 inhibitor may elicit a better response in patients with CaB than either agent alone. Investigators have studied the effect of celecoxib (PÞzer’s Celebrex) on breast tumors in preclinical studies (Abou-Issa HM, 2000). Both Cornell Research Foundation and Collagenex have been granted patents protecting their methods for treating cancer with COX-2 inhibitors. Recent controversy has surrounded the COX-2 inhibitor class, and they have been associated with increased cardiovascular events. Indeed, the trial that initially identiÞed this problem was conducted in (colorectal) cancer patients. Cancer patients are particularly prone to thromboembolic events. This situation has led to massive uncertainty about the future of COX-2 inhibitors. We include them in our analysis for completeness, as clinical trials seem to still be ongoing.

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O

H2N S O

N

N CF3

H3C FIGURE 21. Structure of celecoxib (R1 = NH2 , R2 = CH3 ).

Mechanism of Action. The antitumor and chemopreventive mechanisms of COX-2 inhibitors have not been fully elucidated. Researchers theorize that these agents may exert their effect via tumor necrosis factor (TNF) and nuclear factor kappa beta (NFκβ) inhibition. Celecoxib. PÞzer’s celecoxib (Celebrex) (Figure 21) is registered in all seven markets for the treatment of rheumatoid arthritis and osteoarthritis, and it has been studied extensively as a chemopreventive agent in individuals with familial adenomatous polyposis (FAP). Celecoxib is undergoing various clinical trials for CaB, including Phase III trials in the United States and Spain, investigating it both as an adjunct to chemotherapy regimens for the treatment of advanced disease and as a neoadjuvant treatment. One ongoing U.S. and Spanish Phase III study is randomizing postmenopausal women at increased risk of developing CaB to receive exemestane with or without celecoxib. More than 5,000 patients are planned for project accrual. Another ongoing U.S. Phase III trial with a projected accrual of almost 7,000 CaB patients will compare the event-free survival of postmenopausal women with receptorpositive primary breast cancer when treated with exemestane versus anastrozole as adjuvant therapy with or without celecoxib. In one ongoing Phase II trial, celecoxib is being given with letrozole in a neoadjuvant setting to 20 patients; it is being compared with letrozole or exemestane (PÞzer’s Aromasin) alone (Toi M, 2003). The combination treatment has been well tolerated; however, despite one complete response, no signiÞcant difference between the groups was observed at this early stage. Phase II data were presented at the 2004 ASCO meeting (Chow LW, 2004). Three cycles of 5-FU, epirubicin, and cyclophosphamide (FEC), with or without celecoxib (400 mg bid), were given at three-week intervals to women with histologically proven, locally advanced CaB. A total of 31 patients with a median age of 45.6 years were enrolled (15 FEC, 16 FEC plus celecoxib). Response rates were as follows: FEC, 62.5% FEC plus celecoxib, 81.3% (complete response 18.8%, partial response 62.5%). In the FEC plus celecoxib arm, mean tumor size decreased from 4.25 cm to 2.04 cm. The regimens were well tolerated, and no clinical cardiac toxicity was detected. Among tumors that showed clinical

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response (n = 9) to the FEC/celecoxib combination, a signiÞcantly higher level of COX-2 gene expression was observed versus nonresponders (n = 6). In another Phase II trial, celecoxib is being studied in combination with exemestane in patients with advanced CaB; 30 patients have been enrolled so far. According to data reported at ASCO 2003, 9 of the 13 evaluable patients (69%) remained on therapy without disease progression at six months (Canney P, 2003). Of the 19 patients evaluable for response, 6 (31%) had a partial response, 8 (42%) had stable disease, and 5 (26%) had progressive disease, giving a clinical beneÞt (partial response plus stable disease) in 73% of patients. Four patients experienced gastrointestinal toxicity, three experienced hot ßashes/sweats, and one patient reported headaches. The combination of celecoxib and exemestane shows promising early activity and acceptable toxicity. This trial will continue until 53 patients are enrolled. Chemotherapy Potentiators Overview. Potentiators of chemotherapy are agents intended to increase the efÞcacy of existing cytotoxic anticancer agents via a number of different mechanisms. Agents that inhibit the DNA repair enzymes that reduce the efÞcacy of DNA-damaging agents are under extensive preclinical study as potentiating agents. We restrict our discussions in this class to YM BioSciences’ tesmilifene because it is the most advanced in clinical development. Mechanism of Action. According to the theory of how intracellular histamine receptor ligands exert their cell-proliferative action, intracellular histamine mediates DNA and protein synthesis. This action may be caused by the downregulation of growth-inhibitory prostaglandins. Increased expression of intracellular histamine receptors may also be present in tumor cells. Antagonism of the intracellular action of histamine at intracellular histamine receptors by selective antihistamines may disrupt growth of malignant cells. Tesmilifene. YM BioSciences/Shin Poong’s small-molecule antihistamine agent tesmilifene is undergoing Phase III clinical trials in the United States and Europe as a treatment for CaB. A Phase III trial involving 305 patients with metastatic CaB has been completed (Reyno L, 2004). The trial compared the addition of tesmilifene to doxorubicin with anthracycline monotherapy. The Þnal analysis was conducted as planned after 256 progression events (median follow-up = 20.5 months). There was no signiÞcant difference between arms in response rate, response duration, or the primary end point, progression-free survival. The study was closed to additional accrual and the tesmilifene arm was discontinued. The researchers did Þnd that patients receiving the tesmilifene/doxorubicin combination had improved median survival of 23.6 months, versus 15.6 months for those receiving the anthracycline alone. Tesmilifene/doxorubicin was associated with more gastrointestinal and CNS toxicity than doxorubicin alone. No consistent inßuence on quality of life was detected.

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Despite these negative data, a registrational Phase III trial with tesmilifene in 700 patients with metastatic CaB or recurrent local disease was initiated in the United States and Western Europe in March 2004. The trial will compare the overall survival times of patients receiving tesmilifene in combination with epirubicin (PÞzer’s Ellence, generics)/cyclophosphamide with the survival period of patients taking epirubicin/cyclophosphamide alone. The primary end point of this pivotal trial is survival.

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Cervical Cancer

ETIOLOGY AND PATHOPHYSIOLOGY Background Cervical cancer (CaC) afßicts approximately 44,000 women each year in the major markets under study (United States, France, Germany, Italy, Spain, United Kingdom, and Japan), making it the fourth most common female malignancy. In many developing countries in sub-Saharan Africa, Central and South America, and south and southeast Asia, CaC is the most common lethal female cancer, and its incidence in these countries continues to increase. Developing countries now account for approximately 80% of CaC cases. The increase in incidence is primarily the result of lack of awareness of the disease and its outcome as well as lack of national screening programs for the detection of early, precancerous cervical changes, and HPV infection. In Western countries, screening programs have been credited with reducing CaC incidence by 74% (Peto J, 2004). Diagnosis of CaC is made following cytological analysis of Pap smears (described further on) and conÞrmed by biopsy. Occasionally, an incidental diagnosis of CaC may be made during a hysterectomy. For our purposes, the deÞnition of CaC refers to invasive carcinoma only. Invasive CaC is conÞrmed when cervical cancer cells are found deep within the tissue of the cervix rather than on the surface of the cervix only (known as cervical cancer in situ). A detailed diagnosis of CaC is made according to the deÞnitions of the 1994 International Federation of Gynecology and Obstetrics (FIGO) clinical staging system, detailed in Table 1, Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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297

TABLE 1. Carcinoma of the Uterine Cervix: FIGO Clinical Staging System Stage 0 Stage I Stage 1a

Stage Ia1 Stage Ia2 Stage Ib Stage Ib1 Stage Ib2 Stage II Stage IIa Stage IIb Stage III

Stage IIIa Stage IIIb Stage IV

Stage IVa Stage IVb

Carcinoma in situ, cervical intraepithelial neoplasia (CIN) grade III. The carcinoma is strictly confined to the cervix (extension to the corpus would be disregarded). Invasive carcinoma that can be diagnosed only by microscopy. All macroscopically visible lesions—even with superficial invasion—are allotted to stage IB carcinomas. Invasion is limited to a measured stromal invasion with a maximal depth of 5.0 mm and a horizontal extension no wider than 7.0 mm. (Depth of invasion should be no greater than 5.0 mm taken from the base of the epithelium of the original tissue superficial or glandular. The involvement of vascular spaces—venous or lymphatic—should not change the stage designation.) Measured stromal invasion no greater than 3.0 mm in depth and extension no wider than 7.0 mm. Measured stromal invasion greater than 3.0 mm and no greater than 5.0 mm with an extension no wider than 7.0 mm. Clinically visible lesions limited to the cervix uteri or preclinical cancers greater than stage IA. Clinically visible lesions no greater than 4.0 cm. Clinically visible lesions greater than 4.0 cm. Cervical carcinoma invades beyond the uterus, but not to the pelvic wall or to the lower third of the vagina. No obvious parametrial involvement. Obvious parametrial involvement. The carcinoma has extended to the pelvic wall. On rectal examination, there is no cancer-free space between the tumor and the pelvic wall. The tumor involves the lower third of the vagina. All cases with hydronephrosis or nonfunctioning kidney are included, unless they are due to other causes. Tumor involves lower third of the vagina, with no extension to the pelvic wall. Extension to the pelvic wall and/or hydronephrosis or nonfunctioning kidney. The carcinoma has extended beyond the true pelvis, or has involved the mucosa of the bladder or rectum (biopsy-proven). (A bullous edema, as such, does not permit a case to be designated as stage IV.) Spread to adjacent organs. Spread to distant organs.

FIGO = International Federation of Gynecology and Obstetrics. Source: Benedet JL, et al. Carcinoma of the cervix uteri. Journal of Epidemiology and Biostatistics. 2001;6(1):5–44.

which are based on criteria relating to tumor diameter and depth of invasion; invasion outside of the uterus; involvement of the pelvic wall, parametrium, bladder, kidney, and rectum; and spread to distant organs. Anatomy The uterine cervix is the lower, narrow tip of the uterus that extends into and joins the vagina, as shown in Figure 1. It is conical in shape and projects through the anterior wall of the vagina, which divides it into an upper portion, called the endocervix, and a lower, vaginal portion called the ectocervix. The endocervix is separated in the front from the bladder by Þbrous tissue called the parametrium, within which lie the uterine arteries. On the rounded extremity of the ectocervix

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Fallopian tube

Secrum

Ovary Uterus Bladder Cervix Vagina

Rectum Anus

FIGURE 1. Female reproductive system showing position of the uterine cervix.

is a small circular aperture called the external oriÞce of the uterus. Externally, the oriÞce is bounded by two lips, called the anterior and posterior lips. The cervix is made up of epithelium and underlying stroma. The stroma contains both smooth muscle and Þbrous tissues. The ectocervix is lined with squamous epithelium, which comprises several layers: basal, parabasal, intermediate, and superÞcial. The basal layer is a single row of cells on a thin basement membrane; active mitosis occurs in this layer. The superÞcial layer varies in thickness, depending on the degree of estrogen stimulation. The response of the epithelium to sex hormones is identical to that of the vagina. The endocervix is covered by mucin-secreting, simple, columnar epithelium, which lines the surface. The border of the endocervix and ectocervix is located between the stratiÞed squamous epithelium of the ectocervix and the columnar epithelium of the endocervix and is called the squamocolumnar junction. During sexual development, under the inßuence of hormones, the endocervix progresses onto the ectocervix and lies within the vagina. Under the inßuence of vaginal secretions, the endocervix undergoes transformation from columnar to squamous epithelium. The majority of cervical tumors arise at the transformation zone. Etiology Risk Factors. All sexually active women are at risk for CaC. The most important risk factor for the development of CaC is infection with human papilloma virus, or HPV, which is transmitted through genital-to-genital sexual contact. Researchers estimate that as much as 99% of CaC cases are related to persistent HPV infection. However, other, secondary factors play a role because most cases of HPV infection have no clinical manifestation and regress spontaneously. Even the majority of cases of HPV-induced cellular neoplasia or abnormalities do not progress to invasive cancer; approximately one-third of these cases will become CaC. No apparent underlying genetic or hereditary basis exists for the development of CaC. Infection with Human Papillomavirus. Papillomaviruses are small, doublestranded DNA viruses encased in a protein capsid that contain a circular genome consisting of approximately 7,900 base pairs. The HPV-16 genome (Figure 2)

ETIOLOGY AND PATHOPHYSIOLOGY

Regulatory noncoding region LCR

299

Transforming proteins E6

E7

Major capsid protein L1 E1 Replication HPV 16

Major capsid protein L2 E2 Replication and transcription E5

FIGURE 2. HPV-16 viral genome structure.

TABLE 2. Prominent Proteins of the Human Papillomavirus Genome and Their Functions Protein E1 E2 E6 E7 L1 L2

Function/Comments DNA-dependent ATPase. Allows unwinding of the viral genome and acts as an elongation factor for DNA replication. Regulates viral DNA transcription and replication. Involved in transformation of the host cell by binding p53 tumor suppressor protein. Transforming protein, binds to retinoblastoma protein pRB/p107. Major capsid protein. Can form virus-like particles. Minor capsid protein. Possible DNA packaging protein.

has three regions: the upstream regulatory region (URR), the early (E) region, and the late (L) region. The E region is divided into seven areas that code for different proteins important at various stages during the life cycle and crucial for viral replication (see Table 2). E6 and E7 produce the proteins that cause malignant cellular transformation and oncogenesis and therefore represent targets for therapeutic intervention. The HPV E6 gene encodes a small 16–19 kDa basic protein, E6, which is expressed in the nuclear matrix. The E6 protein encoded by high-risk HPV types interacts with the host p53 tumor suppressor protein and disrupts p53’s normal function by causing its degradation. Absence of functional p53 protein prevents the activation of p53-mediated apoptosis and makes the cell highly susceptible to DNA damage. To associate with p53, the E6 protein requires a cellular protein called E6-associated protein (E6-AP). Like other oncogenic virus proteins, such as SV40 large T antigen and adenovirus E1A, E6 proteins of high-risk HPV abrogate the ability of wild-type p53 to activate transcription. However, the mechanism of the E6 protein’s action is different from that of SV40 large T antigen and the adenovirus E1A protein. The E6 protein causes degradation of p53, whereas the other oncogenic proteins interact with p53 and disrupt its function

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without degrading the protein. Research has shown that E6-dependent degradation of p53 occurs through the cellular ubiquitin proteolysis pathway (Werness BA, 1990). The HPV E7 gene is expressed together with the E6 gene as a part of early transcription. The E7 protein has transforming and transcriptional regulatory functions (Barbosa MS, 1989). It controls the cell cycle by interacting with the tumor suppressor retinoblastoma (Rb) protein P105 and other Rb-associated proteins, such as P107 and P130, displacing the E2F transcription factor (Chellappan S, 1992). This action results in the unregulated activation of the cell cycle in the infected cell. The E6 and E7 proteins are invariably expressed in HPV-associated tumors (zur Hausen H, 2002). The absolute requirement for continued expression of E6 and E7 genes to maintain the HPV transformed (or cancerous) phenotype makes these proteins extremely attractive targets for immunotherapy of cervical tumors. Several strategies are being used to induce host immunologic responses against these proteins, including (1) recombinant vaccinia vectors that express E6 and E7 genes, (2) soluble recombinant proteins with adjuvant, and (3) synthetic peptides that represent the appropriate epitopes. Because both transforming genes are expressed from the same messenger RNA (mRNA), this transcript has become a target for gene inhibition therapies. Natural infection of humans by HPV does not appear to provoke strong antibody responses against E6 and E7 proteins; antibody responses against E6 or E7 proteins from HPV-16 are most prevalent in patients with advanced CaC but appear to have little prognostic or therapeutic value (Frazer IH, 1996). About 100 types of HPV have been described, which have been numbered in the order of their discovery. An HPV type is considered new when at least 10% of the gene sequences in the HPV regions E6, E7, and L1 (Figure 2) differ from any previously known type. Each type of HPV has an afÞnity for particular types of epithelium. Oncogenic potential varies among subtypes, and this variation is reßected in the E6 and E7 proteins. Low-risk HPVs have a much smaller capacity than high-risk HPVs to induce malignant transformation, owing to functional differences between their E6 and E7 proteins. Approximately 30 types of HPV are known to infect the anogenital tracts of both sexes; of these, types 16, 18, 31, and 45 have the highest risk of causing CaC (Hwang TS, 2003; Munoz N, 2003). The E6 and E7 proteins of HPV types 16 and 18 are attractive targets for the development of both preventive and therapeutic vaccines and antisense therapies. Sexual Activity Under Age 18 Years. Becoming sexually active at an early age is associated with an increased risk of CaC (Zarama Marquez FA, 2003). This risk is the result of increased fragility of developing cervical cells in younger women, making the cells more liable to damage during sexual intercourse. This damage may make them more susceptible to HPV infection. Teenagers who smoke in addition to having early sexual intercourse further increase their risk of developing CaC (Zarama Marquez FA, 2003; Berrington de Gonzalez A, 2004).

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301

Multiple Sexual Partners. An increase in the incidence of HPV infection and CaC has been found in women who have multiple sexual partners (Zarama Marquez FA, 2003; Berrington de Gonzalez A, 2004), primarily because it increases the likelihood of sexual interactions with a male carrier of HPV. Therefore, the incidence of HPV in the male population is a factor that affects the incidence of HPV infection in women and, ultimately, the incidence of CaC. In addition, the incidence of sexually transmitted diseases (STDs) generally is higher in women who have multiple sexual partners, and cervical neoplasia may develop following a sexually transmitted infection. Teenagers are particularly at risk for STDs. Women who have only one sexual partner may still be at increased risk if their partners have many sexual contacts (Castellsague X, 2003). Cigarette Smoking. Women who smoke cigarettes have a higher risk of CaC than nonsmokers (Minkoff H, 2004; Eifel PJ, 2002; Berrington de Gonzalez A, 2004). Although certain chemicals from cigarette smoke have been found in cervical tissue, it remains uncertain whether cigarette smoke exerts a direct carcinogenic effect. Also important may be the fact that the immune system is often impaired in smokers, making these women more susceptible to infection by HPV. Immunosuppression. As with all infectious diseases, people who are immunocompromised are at higher risk of infection. Groups at high risk for HPV infection, and therefore CaC, include women who carry the HIV virus; the correlation between HIV infection and development of CaC is so strong that infected women are advised to undergo a Pap smear every six months. HIV-infected women who develop CaC tend to exhibit very aggressive disease (Ahdieh-Grant L, 2004). Socioeconomic and Ethnic Factors. The incidence of CaC is higher in economically disadvantaged women, a population that often includes ethnic groups such as African-Americans and Hispanics (Bazargan M, 2004; Singh GK, 2004). This increase in incidence results from the lack of awareness, the lack of health insurance, and the consequent low uptake of Pap screening. Other factors that discourage women from undergoing Pap screening include cultural inßuences and the stigma of HPV as an STD. Pathophysiology Following infection by HPV, preinvasive cancer may develop over a period of months to years. As much as two-thirds of cases of severe neoplasia (CIN3, carcinoma in situ) will progress to invasive CaC if left untreated (Paraskevaidis E, 2002). Progression is slow. It can take 3–30 years from HPV infection to the development of invasive cancer; on average, it takes ten years (Table 3). The degree of neoplasia is determined by the extent of mitotic activity and proliferation index of immature cells and abnormal cell nuclei. If these aberrations are detected only in the lower third of the epithelium, the lesion is designated cervical intraepithelial neoplasia grade 1 (CIN1). Lesions that also involve the

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TABLE 3. Characteristics of Progression from Cervical Intraepithelial Neoplasia to Cervical Cancer Grade of Neoplasia CIN 1–2 CIN 3 Microinvasion

Median Age

Median Time to Progression

Cases That Progress to Cervical Cancer (%)

22 28 41

7 years to carcinoma in situ 7–10 years to microinvasion 2 years to invasive carcinoma

50 66 100

CIN = Cervical intraepithelial neoplasia.

middle third or the upper third are termed CIN2 and CIN3, respectively. The designation adenocarcinoma in situ is used when normal endocervical epithelial cells are replaced by irregular columnar cells with increased mitosis but there is no obvious stromal invasion. More than 90% of invasive CaC cases are squamous-cell carcinomas, which originate in the cell lining of the lower cervix; another 5% are adenocarcinomas, which originate from glandular-type cells in the endocervix. Researchers have identiÞed several subtypes of adenocarcinoma and estimate the distribution of these subtypes as follows: endocervical (60%), endometrioid (10%), clear-cell carcinomas (10%), and adenosquamous carcinoma (20%). The adenocarcinomas tend to be more difÞcult to diagnose and are more aggressive in nature, but they are managed primarily in the same way as CaC of squamous-cell origin. The remaining 5% of invasive CaC cases are rare and include small-cell carcinomas and cervical sarcomas; both have a poor prognosis. CaC is staged in all major markets according to the FIGO guidelines, established in 1994 and subject to regular review (Benedet JL, 2003) (Table 1). This staging system has also been adopted by the National Comprehensive Cancer Network (NCCN). The network published its version of FIGO guidelines in 2003 (www.nccn.org). The American Joint Committee for Cancer (AJCC) has developed a TNM classiÞcation system for CaC that deÞnes stage groupings. Between 80% and 90% of cervical carcinomas are of squamous origin, and approximately 95% of these carcinomas originate in the transformation zone (see “Anatomy,” above). For this reason, the whole transformation zone should be sampled in a Pap smear to enable accurate interpretation. The cells taken during a Pap smear include cells from the epithelium of the ectocervix and the endocervix. The proportion of squamous-cell carcinomas to cervical adenomas is declining, resulting in an apparent increase in adenomas. This effect is attributed to the increased sensitivity of Pap smears for detecting squamous-cell carcinomas. Squamous carcinomas can be classiÞed as large-cell keratinizing, large-cell nonkeratinizing, or small-cell. Most experts believe that small-cell carcinomas have a poorer prognosis than the other types. Rarely, anaplastic small-cell carcinomas are diagnosed; these carcinomas resemble oat-cell carcinoma of the lung and frequently have neuroendocrine features. Anaplastic small-cell carcinomas of the cervix are extremely aggressive and have a poor prognosis—less than 50% survival, even for patients diagnosed with stage I disease.

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TABLE 4. Survival by Stage of Cervical Cancer FIGO Stage Stage IA Stage IB Stage IIA Stage IIB Stage III Stage IVa Stage IVb

Five-Year Survival >95% 80–90% 75–80% 60% 40–65% >20% 5–20%

FIGO = International Federation of Gynecology and Obstetrics.

Approximately 80% of adenocarcinomas of the cervix originate in the endocervical glandular epithelium. The remaining 20% of tumors are endometreoid, clear cell, intestinal cell, or of mixed origin; these tumors are sometimes difÞcult to differentiate from cancers of ovarian or endometrial origin. CaC may invade the lymphatic system, spreading initially to local lymph nodes within the pelvis and ultimately to the aortic lymph nodes and lymph nodes in the groin. Local spread of the cancer to the outer vagina and vulva is common. Distant metastases may also arise, disseminated via the bloodstream; distant metastases typically manifest in the lungs, liver, and brain. In CaC, survival rates correlate with tumor stage. The prognosis for patients with early disease is good when they are treated with surgery and/or radiotherapy. For bulky tumors and disease that has spread to the lymph nodes, cure rates are lower, despite treatment with surgery and/or radiochemotherapy (Table 4). The outlook for patients with metastatic disease is bleak, and although treatment may result in a small survival beneÞt, it is largely palliative. The FIGO staging system incorporates additional prognostic factors. Survival correlates with disease characteristics such as the following: • • • •

Tumor diameter in early disease. Tumor bulk in advanced disease. Unilateral or bilateral involvement of the pelvic wall. Lymph node involvement (size and number).

Other prognostic factors include posthysterectomy invasion of the lymph-vascular space, which worsens prognosis and histologic tumor type (Eifel PJ, 1994; Logsdon MD, 1999). One analysis demonstrated that the risk of death associated with adenocarcinoma is almost twice the risk associated with squamous-cell carcinoma (Eifel PJ, 1995). CURRENT THERAPIES Surgery is the mainstay of treatment for early-stage, localized cervical cancer (CaC). Radiotherapy, in the form of external beam radiation (EBT), is employed

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at all stages. The utility of cytotoxic chemotherapy in patients with locally advanced, metastatic, or recurrent disease has been investigated in clinical trials with a variety of study designs. These designs include neoadjuvant chemotherapy, administered prior to surgical resection or radiation; concomitant chemotherapy and radiation, in which both modalities are administered together; adjuvant chemotherapy, in which surgery or radiation is followed by chemotherapy; and palliative chemotherapy for patients with widely disseminated tumor not suitable for palliative radiation. Cisplatin (Bristol-Myers Squibb’s [BMS’s] Platinol-AQ, Nippon Kayaku’s Randa, generics) is the agent of choice for pharmaceutical treatment of CaC. Its single-agent activity has produced response rates of 18–31% in multiple trials conducted in more than 900 patients. In advanced disease a study adding topotecan to the standard regimen of single agent cisplatin administered every 3 weeks resulted in a highly signiÞcant prolongation of survival (9.4 versus 6.5 months). In addition, this combination was found to be well tolerated and not associated with a decrease in quality of life. Interestingly, the response rate (13%) to cisplatin alone in this trial was surprisingly low and was most likely related to the new era of upfront chemo-radiation treatments. Long HJ, Bundy BN, Grendys EC et al. Randomized phase III trial of cisplatin with or without topotecan in carcinoma of the uterine cervix: a Gynecologic Oncology Group study. Journal of Clinical Oncology 2005;23:4626–4633. It has been used for palliative treatment of metastatic, locally advanced, and recurrent disease for several decades. Clinicians use either single-agent cisplatin or cisplatin in combination with other agents; combination therapy increases response rates (and usually toxicity) but does not greatly affect survival. Increasingly, cisplatin is used in addition to radiotherapy and in bulky nonmetastatic disease. Cisplatin-containing regimens are now used in all but very early-stage disease, either as an adjuvant to surgery or as Þrst-line treatment in more extensive, inoperable disease contained within the pelvis (stages IIB–IVA). Five out of six trials demonstrate a survival advantage for cisplatin-based, concurrent chemoradiotherapy compared with non-cisplatin-based chemoradiotherapy or radiotherapy alone (Rose PG, 1999). The use of systemic cisplatin in addition to radiotherapy is thought to reduce the likelihood of distant as well as local relapse. The data from trials of concurrent cisplatin and radiotherapy (Whitney C, 1999; Morris M, 1999; Rose PG, 1999; Keys HM, 1999; Peters WA, 2000; Thomas GM, 1999) prompted the National Cancer Institute (NCI) to release a clinical announcement supporting the concurrent use of cisplatin-based chemotherapy with radiotherapy for high-risk, early-stage, and locally advanced CaC (Loizzi V, 2003). Women with CaC for whom treatment with radiotherapy is being considered should be offered concurrent cisplatin with their course of radiotherapy. These patients include the following: • • •

Patients with locally advanced cervical cancer. Patients with bulky, clinical stage IB (greater than 4 cm) CaC. Patients who are treated with radiotherapy.

CURRENT THERAPIES

• •

305

Patients with high-risk, early-stage CaC (node-positive or margin-positive). Patients who will be treated with radiotherapy following hysterectomy.

Neoadjuvant treatment that incorporates cisplatin is used increasingly to raise the possibility of performing curative surgery. However, numerous trials have failed to show a beneÞt from using cisplatin prior to radiotherapy in bulky stage IB and II disease—indeed, some trials have shown a reduction in survival using this regimen. Table 5 lists the most commonly used regimens for the treatment of CaC. Cisplatin Single-Agent Regimen Overview. Cisplatin (BMS’s Platinol-AQ, Nippon Kayaku’s Randa, generics) is the most-established single agent for the treatment of metastatic CaC (mCaC), producing response rates of up to 25%. U.S. national guidelines recommend its use in conjunction with radiotherapy for CaC stages Ib through IVa. This combined treatment modality is also used extensively in the treatment of earlier stages of CaC. In mCaC, response rates are signiÞcantly higher when cisplatin is used in combination with other cytotoxic agents: up to 46%. However, improved survival in mCaC patients also has been reported using a combination of cisplatin with topotecan (Long H, 2004). In an attempt to determine an optimal cisplatin combination, four combinations are being investigated in a randomized Phase III trial. Clinicians interviewed note that single-agent cisplatin is not included in this trial because of the evidence showing better response rates with combinations. Cisplatin is also becoming widely established as a radiosensitizer and in some cases is replacing both radiotherapy alone and surgery. No randomized trials have compared different cisplatin-based regimens, but a widely accepted schedule is 40 mg/m2 (maximum of 70 mg/week) intravenous (IV) given weekly with concurrent radiotherapy at 1.8–2.0 Gy daily, Þve days a week for six weeks. At this modest dose level, cisplatin is considered to be a radiosensitizer, although some clinicians claim it will still have a cytotoxic effect on both local and occult distant disease. Typically, when cisplatin is used in the absence of radiotherapy, a standard dose of 100 mg/m2 IV is given every three weeks. Mechanism of Action. Cisplatin (Figure 3) is a platinum agent. Platinum agents generate highly reactive, charged platinum complexes that bind covalently to DNA, leading to intrastrand, interstrand, and less importantly, protein crosslinking of DNA. These cross-links inhibit transcription and/or DNA replication

FIGURE 3. Structure of cisplatin.

306

TABLE 5. Current Regimens Used for Cervical Cancer Regimen

Regimen Components

Availability

Dose mg/m2

Cisplatin single agent

Cisplatin (BMS’s Platinol-AQ, Nippon Kayaku’s Randa, generics)

US, F, G, I, S, UK, J

IV weekly (with 40 radiotherapy) 100 mg/m2 IV every 3 weeks

Cisplatin plus 5-fluorouracil (5-FU)

Cisplatin (BMS’s Platinol-AQ, Nippon Kayaku’s Randa, generics)

US, F, G, I, S, UK, J

Cisplatin 70 mg/m2 IV day 1

5-fluorouracil (generics)

US, F, G, I, S, UK, J

5-FU 1,000 mg/m2 /day IV continuous infusion days 1–4

Cisplatin plus 5-fluorouracil (5-FU) plus hydroxyurea

Cisplatin (BMS’s Platinol-AQ, Nippon Kayaku’s Randa, generics)

US, F, G, I, S, UK, J

5-fluorouracil (generics)

US, F, G, I, S, UK, J

Hydroxyurea (BMS’s Hydrea/Litalir, generics)

US, F, G, UK, J

Two cycles with radiotherapy and 2 cycles following completion of radiotherapy Cisplatin 50 mg/m2 IV days 1, 29

5-FU 1,000 mg/m2 /day IV continuous infusion days 1–4 and 29–32 Hydroxyurea 2,000 mg/m2 oral, twice weekly 2 hours before radiotherapy, weeks 1–6

Common Toxicities Leukopenia Nausea and vomiting Hearing loss (100 mg/m2) Neurotoxicity (100 mg/m2 ) Leukopenia Granulocytopenia Nausea and vomiting Diarrhea

Leukopenia Gastrointestinal toxicity Nausea and vomiting

TABLE 5. (continued) Regimen Cisplatin plus topotecan

Cisplatin plus ifosfamide

Carboplatin plus ifosfamide

Cisplatin plus paclitaxel

Regimen Components

Availability

Dose mg/m2

Common Toxicities

307

Cisplatin (BMS’s Platinol-AQ, Nippon Kayaku’s Randa, generics)

US, F, G, I, S, UK, J

Cisplatin 50

IV day 1

Topotecan (GSK’s Hycamtin)

US, F, G, I, S, UK, J

Cisplatin (BMS’s Platinol-AQ, Nippon Kayaku’s Randa, generics)

US, F, G, I, S, UK, J

Topotecan 0.75 mg/m2 days 1–3 Repeat cycle every 3 weeks Cisplatin 50 mg/m2 IV day 1

Ifosfamide (BMS’s Ifex, Baxter’s Mitoxana/Holoxan, Shionogi’s Ifomide, generics)

US, F, G, I, S, UK, J

Carboplatin (BMS’s Paraplatin, generics)

US, F, G, I, S, UK, J

Ifosfamide (BMS’s Ifex, Baxter’s Mitoxana/Holoxan, Shionogi’s Ifomide, generics)

US, F, G, I, S, UK, J

Cisplatin (BMS’s Platinol-AQ, Nippon Kayaku’s Randa, generics)

US, F, G, I, S, UK, J

Paclitaxel (BMS’s Taxol, generics)

US, F, G, I, S, UK, J

Ifosfamide 1,000 mg/m2 IV continuous infusion over 24 hours day 1 (with mesna 6,000 mg/m2 ) Repeat cycle every 3 weeks Carboplatin IV AUC = 4, day 1

Ifosfamide 1,600 mg/m2 IV continuous infusion over 24 hours day 1 (with mesna 6,000 mg/m2 ) Repeat cycle every 3 weeks Cisplatin 75 mg/m2 IV day 2

Paclitaxel 135 mg/m2 IV day 1 Repeat cycle every 3 weeks

Neutropenia Anemia Nausea and vomiting

Neutropenia Leukopenia Thrombocytopenia Nausea and vomiting Neurotoxicity

Neutropenia Leukopenia Thrombocytopenia Nausea and vomiting

Neutropenia Leukopenia Thrombocytopenia Nausea and vomiting Diarrhea

308 TABLE 5. (continued) Regimen Cisplatin plus gemcitabine

Paclitaxel plus ifosfamide plus cisplatin (TIP regimen)

Regimen Components

Availability

Dose mg/m2

Common Toxicities

Cisplatin (BMS’s Platinol-AQ, Nippon Kayaku’s Randa, generics)

US, F, G, I, S, UK, J

Cisplatin 75

Gemcitabine (Eli Lilly’s Gemzar)

US, F, G, I, S, UK, J

Paclitaxel (BMS’s Taxol, generics)

US, F, G, I, S, UK, J

Gemcitabine 1,250 mg/m2 IV days 1 and 8 Repeat cycle every 3 weeks Paclitaxel 175 mg/m2 IV day 1

Ifosfamide (BMS’s Ifex, Baxter’s Mitoxana/Holoxan, Shionogi’s Ifomide, generics) Cisplatin (BMS’s Platinol-AQ, Nippon Kayaku’s Randa, generics)

US, F, G, I, S, UK, J

Ifosfamide 5,000 mg/m2 IV day 1

US, F, G, I, S, UK, J

Cisplatin 75 mg/m2 IV day 1 Repeat cycle every 3 weeks

US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan. AUC = Area under the curve; IV = Intravenous; PO = By mouth. BMS = Bristol-Myers Squibb; GSK = GlaxoSmithKline.

IV day 2

Neutropenia Leukopenia Gastrointestinal toxicity Anemia Granulocytopenia

Neutropenia Leukopenia Thrombocytopenia Nausea and vomiting Diarrhea

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309

mechanisms. The principal sites of reaction are the N7 atoms of guanine and adenine. The formation of intrastrand cross-links between cisplatin and neighboring guanines has been shown to correlate with clinical response. When used in conjunction with radiotherapy, cisplatin may also act as a radiosensitizer. Possible mechanisms of radiosensitization include inhibition of repair of sublethal radiation damage and synchronizing tumor cells into a radiosensitive phase of the cell cycle. Clinical Performance. Two pivotal, randomized, Phase III clinical studies investigated single-agent cisplatin with concurrent radiotherapy. The Þrst, the Gynecologic Oncology Group’s GOG120 study (Rose P, 1999), recruited 176 patients with stage IIB, stage III, or stage IVA disease, without para-aortic lymph-node involvement, to receive treatment with a control regimen or one of two investigational treatments. The control arm (177 patients) was treated with hydroxyurea and concurrent radiotherapy. The Þrst investigational arm recruited 174 patients who received 40 mg/m2 cisplatin weekly and concurrent EBT, at 1.8–2.0 Gy daily, Þve days a week for six weeks. The second investigational arm included 173 patients who received cisplatin, 5-ßuorouracil (5-FU), and hydroxyurea with concurrent radiotherapy. The results showed that the relative risk of recurrence for patients treated with cisplatin/EBT was 0.57 (95% conÞdence intervals: 0.42–0.78), and the relative risk of death was 0.61 (95% conÞdence intervals: 0.44–0.85) compared with the hydroxyurea/radiotherapy treatment. Progression-free survival at two years was 67% versus 47%. The frequency of adverse events was similar in the cisplatin/EBT arm and in the control arm; the most important toxicity was hematologic. In the three-drug combination arm, the clinical beneÞt achieved was similar to that of the cisplatin/EBT arm, but both hematologic and gastrointestinal toxicities occurred signiÞcantly more frequently (Rose PG, 2003). The authors concluded that single-agent cisplatin should be added to radiotherapy for this patient population and should replace the older agent, hydroxyurea, in this setting. The second study, GOG123, recruited patients from 1992 to 1997 with stage IB bulky (greater than 4 cm) disease and compared radiotherapy with cisplatin (183 patients) or without cisplatin (186 patients), followed by hysterectomy (Keys HM, 1999). Patients received 40 mg/m2 cisplatin weekly for a maximum of six doses. Relative risks of progression and death when treatment included cisplatin were 0.51 (95% conÞdence intervals: 0.34–0.75) and 0.54 (95% conÞdence intervals: 0.34–0.86), respectively. The three-year survival rates were 83% for radiotherapy/cisplatin treatment and 74% for radiotherapy alone. In the combined therapy arm, patients had a higher rate of hematologic toxicity (21% versus 2%) and gastrointestinal toxicity (14% versus 5%). These data conÞrm the beneÞt of concurrent cisplatin and radiotherapy for the treatment of early-stage, bulky disease. Overall, the risk of death from CaC fell 30–50% with concurrent, cisplatinbased chemoradiotherapy compared with radiotherapy alone. Generally, the relatively low dose of cisplatin used with radiotherapy has an acceptable toxicity

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proÞle. However, clinical beneÞt from cisplatin treatment for mCaC patients is suboptimal, and new agents should be tested in this setting. Cisplatin plus 5-Fluorouracil Regimen Overview. The combination of cisplatin (BMS’s Platinol-AQ, Nippon Kayaku’s Randa, generics) and 5-ßuorouracil (5-FU, generics) has been widely used for decades to treat advanced CaC and, more recently, earlier-stage disease. Preclinical data have shown that adding 5-FU to cisplatin and radiotherapy has a synergistic effect, thought to result from the impairment of DNA repair that renders lethal the radiation-induced tumor cell damage that is usually sublethal. These data provide the rationale for clinical investigation of this combination. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

•

Cisplatin (Figure 3) is a platinum agent. Platinum agents generate highly reactive charged platinum complexes that bind covalently to DNA, leading to intrastrand, interstrand, and less importantly, protein cross-linking of DNA. These cross-links inhibit the transcription and/or DNA replication mechanisms. The principal sites of reaction are the N7 atoms of guanine and adenine. The formation of intrastrand cross-links between cisplatin and neighboring guanines has been shown to correlate with clinical response. When used in conjunction with radiotherapy, cisplatin may also act as a radiosensitizer. Possible mechanisms of radiosensitization include inhibition of repair of sublethal radiation damage and synchronizing tumor cells into a radiosensitive phase of the cell cycle. 5-FU (Figure 4) is a ßuoropyrimidine antifolate. Antifolates exert their cytotoxic effect via several mechanisms. One of the most important targets for 5-FU is thymidylate synthase (TS), a key enzyme in de novo thymidine synthesis. This pyrimidine, which requires folate cofactors, is an essential requirement for normal DNA synthesis. By inhibiting TS, 5-FU inhibits DNA synthesis and causes cytotoxicity. In addition, because 5-FU is a structural analogue of uracil, it is misincorporated into RNA in place of uracil and interferes with normal RNA function, an action that triggers cell death. When used in conjunction with radiotherapy, 5-FU may also act as a radiosensitizer by synchronizing tumor cells into a radiosensitive phase of the cell cycle.

FIGURE 4. Structure of 5-fluorouracil (5-FU).

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Clinical Performance. A Phase III study, initiated in 1986 by the Gynecologic Oncology Group, randomized patients with para-aortic, lymph-node-negative, stage IIB, stage III, or stage IVA CaC to receive either cisplatin with 5-FU or single-agent hydroxyurea, both administered with concurrent radiotherapy (Whitney C, 1999). Patients (n = 177) were randomized to cisplatin/5-FU and 190 received treatment with hydroxyurea. All parameters measured demonstrated a beneÞt of cisplatin/5-FU over hydroxyurea. The relative risk of progression for patients treated with cisplatin/5-FU was 0.79 (90% conÞdence limits: 0.61–0.99) compared with the risk for the hydroxyurea-treated patients. The relative risk of mortality was 0.74 (90% conÞdence limits: 0.58–0.95) for patients treated with cisplatin/5-FU, compared with hydroxyurea-treated patients. Hematologic toxicity was more common and much more severe in the hydroxyurea-treated patients, but other toxicities were equivalent in the two arms. A second randomized Phase II study recruited 403 patients who had either large (greater than 5 cm diameter) stage IB or IIA CaC, or stage IIB, stage III, or stage IVA CaC to receive either radiotherapy alone or radiotherapy plus 5-FU and cisplatin (Morris M, 1999). Five-year survival was 73% for the chemoradiotherapy group versus 58% for radiotherapy alone (p = 0.004), and disease-free survival was 67% versus 40% (p > 0.001). The incidence of hematologic toxicity was higher in the group receiving chemotherapy; however, the data afÞrm that the survival beneÞt gained warrants the use of concurrent chemoradiotherapy. Between 1991 and 1996, 268 patients with early-stage (IA2, IB, or IIA) CaC were randomized in a clinical trial to receive either cisplatin/5-FU with concurrent radiotherapy or radiotherapy alone (Peters W, 2000). Twenty-Þve patients were ineligible, leaving 127 in the chemoradiotherapy arm and 116 in the radiotherapy arm. Because data from other studies were reported while this trial was ongoing, an unscheduled interim analysis was performed to determine any beneÞt for including chemotherapy in treatment. The four-year, progressionfree rates were 80% for chemoradiotherapy and 63% for radiotherapy. The incidence of grade 4 toxicities, primarily hematologic, was higher among the chemoradiotherapy-treated patients: 17% versus 3.5%. No chemotherapy-related deaths were reported. Cisplatin plus 5-Fluorouracil plus Hydroxyurea Regimen Overview. Data do not support a role for single-agent 5-FU (generics) in CaC as they do for cisplatin (BMS’s Platinol-AQ, Nippon Kayaku’s Randa, generics) and hydroxyurea (BMS’s Hydrea/Litalir, generics). However, 5-FU has been studied in combination with other agents in an attempt to improve clinical beneÞt. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

Cisplatin is a platinum agent. Platinum agents generate highly reactive charged platinum complexes that bind covalently to DNA, leading to

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•

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intrastrand, interstrand, and less importantly, protein cross-linking of DNA. These cross-links inhibit the transcription and/or DNA replication mechanisms. The principal sites of reaction are the N7 atoms of guanine and adenine. The formation of intrastrand cross-links between cisplatin and neighboring guanines has been shown to correlate with clinical response. When used in conjunction with radiotherapy, cisplatin may also act as a radiosensitizer. Possible mechanisms of radiosensitization include inhibition of repair of sublethal radiation damage and synchronizing tumor cells into a radiosensitive phase of the cell cycle. 5-FU is a ßuoropyrimidine antifolate. Antifolates exert their cytotoxic effect via a number of mechanisms. One of the most important targets for 5-FU is TS, a key enzyme in de novo thymidine synthesis. This pyrimidine, which requires folate cofactors, is an essential requirement for normal DNA synthesis. By inhibiting TS, 5-FU inhibits DNA synthesis and causes cytotoxicity. In addition, because 5-FU is a structural analogue of uracil, it is misincorporated into RNA in place of uracil and interferes with normal RNA function, an action that also triggers cell death. When used in conjunction with radiotherapy, 5-FU may act as a radiosensitizer by synchronizing tumor cells into a radiosensitive phase of the cell cycle. Hydroxyurea inhibits the enzyme ribonucleotide reductase, thereby blocking deoxyribonucleotide formation and DNA synthesis. Ribonucleotide reductase enzyme is closely related to proliferative status in cancer cells. It is involved in the de novo synthesis of all the precursors used in DNA synthesis. It converts ribonucleotide diphosphates to deoxyribonucleotides. Hydroxyurea is a drug that is speciÞc to the cell-cycle phase; it exerts its action during the S phase.

Clinical Performance. In a randomized Phase III study (GOG120) that recruited women with stage IIB, III, or IVA CaC from 1992 to 1997, 173 patients were treated with cisplatin/5-FU/hydroxyurea with concurrent EBT (Rose P, 1999). The control arm of 177 patients received hydroxyurea and radiotherapy; a third group received cisplatin and radiotherapy. For the arm receiving the triplet chemotherapy, the relative risks of disease progression and death were reduced 0.55 (95% conÞdence intervals: 0.40–0.75) and 0.58 (95% conÞdence intervals: 0.41–0.81), respectively, compared with the control arm. The clinical beneÞt, equivalent to that of single-agent cisplatin, was offset in part by the high incidence of adverse effects, which was double the incidence observed in the control arm and the cisplatin arm. The study concluded that although the triplet regimen had greater efÞcacy compared with hydroxyurea alone, no additional beneÞt was gained by adding 5-FU and hydroxyurea to cisplatin. Cisplatin plus Topotecan Regimen Overview. Cisplatin (BMS’s Platinol-AQ, Nippon Kayaku’s Randa, generics) has been investigated in combination with numerous other agents in an attempt to

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improve clinical efÞcacy. The addition of topotecan (GlaxoSmithKline’s [GSK’s] Hycamtin) has resulted in some encouraging Þndings (topotecan is used routinely for the treatment of ovarian cancer). Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

•

Cisplatin (Figure 3) is a platinum agent. Platinum agents generate highly reactive charged platinum complexes that bind covalently to DNA, leading to intrastrand, interstrand, and protein cross-linking of DNA, resulting in the inhibition of transcription and/or DNA replication mechanisms. Topotecan (Figure 5) is a semisynthetic, water-soluble derivative of camptothecin, which is a cytotoxic alkaloid extracted from plants that acts as an inhibitor of topoisomerase I (the same mechanism of action as irinotecan). Topoisomerase I is the enzyme that produces reversible, single-strand breaks in DNA during DNA replication. These single-strand breaks relieve torsional strain and allow DNA replication to proceed. Topotecan binds to the topoisomerase I-DNA complex and prevents the religation of the DNA strand, resulting in double-strand DNA breakage and cell death. Unlike irinotecan, topotecan is not a pro-drug. As a result, topotecan has different antitumor activities and toxicities than irinotecan. Topotecan is a radiation-sensitizing agent and is speciÞc to the S phase of the cell cycle.

Clinical Performance. A recently reported study presented Phase III data from a three-arm, randomized study of patients with stage IV recurrent or persistent CaC who had recovered from the effects of prior surgery, radiotherapy, or chemoradiotherapy (Long H, 2004). Between 1999 and 2002, 356 evaluable patients were randomized to receive cisplatin alone (145 patients); cisplatin plus topotecan (148 patients); or methotrexate, vinblastine, doxorubicin, and cisplatin (MVAC) (63 patients; this arm was stopped early owing to excessive toxicity).

CH3 N CH3 HO

O N N O HO H3C

O

FIGURE 5. Structure of topotecan.

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The response rate in the cisplatin/topotecan arm was 26%—double the response in the single-agent cisplatin arm (p = 0.0004). Median progression-free survival and overall survival were superior in the combination arm: respectively, 4.6 months versus 2.9 months, and 9.2 months versus 7.0 months. The cisplatin/topotecan combination is unique in producing a survival advantage—not just an improved response rate—compared with cisplatin, and a 24% reduction in the risk of death. However, this improvement came at the cost of increased toxicity associated with the inclusion of topotecan into the treatment—namely, neutropenia (70% versus 1%) and anemia (32% versus 23%). Despite the additional toxicities, some investigators advocate cisplatin plus topotecan as the treatment of choice in this patient population. A randomized Phase III study in mCaC is under way to compare cisplatin combined with either topotecan or one of three other cytotoxic agents. Cisplatin plus Ifosfamide Regimen Overview. Both cisplatin (BMS’s Platinol-AQ, Nippon Kayaku’s Randa, generics) and ifosfamide (BMS’s Ifex, Baxter’s Mitoxana/Holoxan, Shionogi’s Ifomide, generics) have single-agent activity in CaC, prompting investigation whether combining the two agents improves clinical activity. Ifosfamide has been studied as a single agent and in combination with other drugs in different studies. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

•

Cisplatin (Figure 3) is a platinum agent. Platinum agents generate highly reactive charged platinum complexes that bind covalently to DNA, leading to intrastrand, interstrand, and protein cross-linking of DNA, resulting in inhibition of transcription and/or DNA replication mechanisms. Ifosfamide (Figure 6) is an alkylating agent pro-drug and a structural analogue of cyclophosphamide. After being metabolized in the liver to its active form, ifosfamide binds DNA and transfers alkyl groups, causing the formation of DNA cross-links and strand breaks. As a result, this agent prevents DNA replication and causes tumor cell death. The action of ifosfamide is nonspeciÞc to the cell cycle. Ifosfamide is administered almost exclusively with mesna, a compound that inactivates the alkylating activity and helps prevent the cystitis associated with this drug.

Clinical Performance. A randomized study compared single-agent cisplatin, given at 50 mg/m2 , with the combination of cisplatin at the same dose level and ifosfamide at 5g/m2 , administered every three weeks up to six courses, in 454 advanced CaC patients (Omura GA, 1997). The combination regimen achieved a higher response rate (31% versus 18%) and longer response duration (4.6 months versus 3.2 months) than cisplatin alone. However, the combination did not improve overall survival and was associated with more-toxic events.

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FIGURE 6. Structure of ifosfamide.

Another study reported in the same year evaluated cisplatin/ifosfamide as neoadjuvant chemotherapy in patients with stage IIB CaC (de Jonge E, 1997). Sixty-eight patients with previously untreated stage IIB CaC were given two cycles of chemotherapy: cisplatin at 20 mg/m2 and ifosfamide at 1.2 g/m2 on days 1–5, repeated every three weeks. Responders were subsequently randomized to either surgery or radiotherapy. A clinical response was documented in 80% of evaluable patients (95% conÞdence intervals: 69–91%). The response rate among the intent-to-treat population was 64.7%. Twenty-one patients were randomized to surgery and 23 patients to radiotherapy. One patient had grade 4 thrombocytopenia, four had grade 3 anemia, one patient had leukopenia, one had nausea, and a third suffered vomiting. The authors concluded that the combination of cisplatin and ifosfamide has acceptable toxicity and produces a clinical response rate of 80% in previously untreated patients with stage IIB CaC. Carboplatin plus Ifosfamide Regimen Overview. In some cancers, such as ovarian and non-small-cell lung cancer, carboplatin (BMS’s Paraplatin, generics) has replaced cisplatin because it has been shown to have equivalent efÞcacy but with an improved side-effect proÞle (Paccagnella A, 2004; du Bois A, 2003). To date, the role of carboplatin as a radiosensitizer has not been fully elucidated, though studies have been performed with carboplatin as either a neoadjuvant treatment for CaC or for treatment of mCaC. Carboplatin has been studied in combination with ifosfamide (BMS’s Ifex, Baxter’s Mitoxana/Holoxan, Shionogi’s Ifomide, generics). Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

•

Carboplatin (Figure 7) is a platinum agent. Platinum agents generate highly reactive charged platinum complexes that bind covalently to DNA, leading to intrastrand, interstrand, and protein cross-linking of DNA, resulting in inhibition of transcription and/or DNA replication mechanisms. Ifosfamide (Figure 6) is an alkylating agent pro-drug and a structural analogue of cyclophosphamide. After being metabolized in the liver to its active form, ifosfamide binds DNA and transfers alkyl groups, causing the formation of DNA cross-links and strand breaks. As a result, this agent prevents

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FIGURE 7. Structure of carboplatin.

DNA replication and causes tumor cell death. The action of ifosfamide is cell-cycle nonspeciÞc. Ifosfamide is used almost exclusively with mesna, a compound that inactivates the alkylating activity and helps prevent the cystitis associated with this drug. Clinical Performance. Investigators in Germany reported an analysis of patients with CaC who, following hysterectomies, were found to possess one or more “high-risk features” for disease recurrence. These patients were subsequently randomized to receive adjuvant chemotherapy (carboplatin [area under the curve (AUC) of 4] plus ifosfamide [1.6 g/m2 /day x three days] x four cycles) followed by EBT radiotherapy, with or without erythropoietin (epoetin alfa, Janssen-Cilag’s Eprex) (10,000 IU three times/week with oral iron). A total of 256 patients were enrolled in the trial, which was well-balanced for known prognostic factors in this malignancy. The investigators found that the requirement for transfusion was substantially reduced in the epoetin-treated patient population (10% versus 32%, p = 0.0004). At a median follow-up of approximately two years, the one-year and two-year relapse-free survivals (primary study endpoint) were 91% versus 86% and 81% versus 70%, respectively, both in favor of the epoetin-treated patient population (p = 0.074). Further follow-up of this study will provide important data on the signiÞcance of anemia and epoetin treatment in the risk of relapse and ultimate survival in locally advanced cervical cancer. Another study performed in Germany (Kuehnle H, 1992) investigated carboplatin and ifosfamide in combination as a neoadjuvant treatment in 34 patients with CaC. After chemotherapy, patients underwent surgery or radiotherapy according to feasibility. Nineteen patients achieved remission; three achieved complete remission. The most common toxicities were myelosuppression, grade 4 leukopenia (28%), and thrombocytopenia (13%); these effects led the authors to conclude that this combination was not recommended for standard practice in this setting. Cisplatin plus Paclitaxel Regimen Overview. Paclitaxel (BMS’s Taxol, generics) is an important anticancer agent with proven activity in a number of cancers, in particular non-small-cell lung cancer, breast cancer, and ovarian cancer. It has also been studied, in combination with cisplatin (BMS’s Platinol-AQ, Nippon Kayaku’s Randa, generics) for the

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treatment of CaC. The combination of cisplatin and paclitaxel is in widespread use for the treatment of mCaC. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

•

Cisplatin (Figure 3) is a platinum agent. Platinum agents generate highly reactive charged platinum complexes that bind covalently to DNA, leading to intrastrand, interstrand, and protein cross-linking of DNA, resulting in inhibition of transcription and/or DNA replication mechanisms. Paclitaxel (Figure 8) is a taxane. Taxanes stabilize the mitotic spindle microtubules formed during cell division, thus preventing depolymerization. This stability inhibits the dynamic reorganization of the microtubules, resulting in a mitotic block at interphase with a consequential inhibition of cell proliferation and eventual cell death.

Clinical Performance. A Phase II study has investigated the efÞcacy of paclitaxel in combination with cisplatin as Þrst-line treatment in 47 patients with advanced or recurrent CaC (Rose P, 1999). Patients were recruited to the study between June and October 1996. Forty-one patients were assessable for response; 5 had a complete response and 14 had a partial response (overall response rate of 46.3%; 95% conÞdence intervals: 30.7–62.6%). The median progression-free survival was 5.4 months, and the median survival was at least 10 months. Interestingly, response was more frequent in disease outside of Þelds that had received prior radiotherapy, a Þnding that suggests untreated disease is more chemosensitive. Another small Phase II study (Piver MS, 1999) investigated the same combination of agents. Of 20 patients, 2 had complete responses and 7 had partial responses, and again, the response rate was improved in patients with recurrence outside radiotherapy-treated Þelds. Again also, the treatment was well tolerated, the most signiÞcant toxicity being grade 3/4 neutropenia in 55% of patients. A GOG Phase III randomized study in 280 patients reported statistically signiÞcant, superior response rates for paclitaxel and cisplatin in combination compared with cisplatin alone: 36% versus 19% (Moore DH, 2001). The median

FIGURE 8. Structure of paclitaxel.

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progression-free interval was also superior: 4.8 months versus 2.8 months. However, these responses did not translate into improved survival: 9.7 months versus 8.8 months is not signiÞcant. The incidence and degree of hematologic toxicity were higher in the combination arm and, although reversible, were deemed unacceptable by the authors in light of the modest improvement in efÞcacy. Cisplatin/paclitaxel is being investigated in an ongoing, four-arm, randomized Phase III study to determine whether response rate or survival is superior for this combination. Cisplatin plus Gemcitabine Regimen Overview. Cisplatin (BMS’s Platinol-AQ, Nippon Kayaku’s Randa, generics) is well established as the most active agent for the adjuvant treatment of CaC and for mCaC. In an attempt to improve on its duration of response and, ultimately, the survival of patients, cisplatin has been combined with new-generation cytotoxic agents such as gemcitabine (Eli Lilly/Spaly Bioquimica’s Gemzar). A meta-analysis (Mutch DG, 2003) concluded that although single-agent gemcitabine was generally inferior to cisplatin, when it was used concurrently with cisplatin and/or radiotherapy, objective response rates were high and survival was prolonged. The drug also showed promise when used with cisplatin as neoadjuvant therapy. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

•

Cisplatin (Figure 3) is a platinum agent. Platinum agents generate highly reactive charged platinum complexes that bind covalently to DNA, leading to intrastrand, interstrand, and protein cross-linking of DNA, resulting in the inhibition of transcription and/or DNA replication mechanisms. Gemcitabine (Figure 9) is an antimetabolite that shares characteristics common to all nucleoside analogues, including mediated transport by membrane transporters, activation by intracellular metabolic steps that retain the nucleotide residues in the cell, and the formation of active phosphate derivatives. Gemcitabine exhibits cell-phase speciÞcity, primarily killing cells undergoing DNA synthesis (S phase) and blocking the progression of cells through the G1/S-phase boundary. It is metabolized intracellularly by nucleoside kinases to the active diphosphate and triphosphate nucleosides.

The cytotoxic effect of gemcitabine is attributed to a combination of two actions of the diphosphate and the triphosphate nucleosides, which leads to inhibition of DNA synthesis. First, gemcitabine diphosphate inhibits ribonucleotide reductase, which is responsible for catalyzing the reactions that generate the deoxynucleoside triphosphates for DNA synthesis. The inhibition of this enzyme by the diphosphate nucleoside reduces the concentrations of deoxynucleotides, including 2
-deoxycytidine 5
-triphosphate (dCTP). Second, gemcitabine triphosphate competes with dCTP for incorporation into DNA. The reduction in the intracellular concentration of dCTP (by the action of the diphosphate) enhances the

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NH2 N HO

O O

OH

N F F

FIGURE 9. Structure of gemcitabine.

incorporation of gemcitabine triphosphate into DNA (self-potentiation). After the gemcitabine nucleotide is incorporated into DNA, only one more nucleotide is added to the growing DNA strands. After this addition, further DNA synthesis is inhibited. DNA polymerase epsilon is unable to remove the gemcitabine nucleotide and repair the growing DNA strands (masked chain termination). Clinical Performance. Data supporting the use of cisplatin/gemcitabine combinations come from Phase I and Phase II clinical trials. Researchers performed a meta-analysis of 15 Phase I and Phase II trials (Mutch DG, 2003) and found gemcitabine to be beneÞcial when given in combination with cisplatin. For example, one Phase II study (Burnett AF, 2000), in which 19 patients recruited between July 1997 and January 1999 received a median of Þve cycles of gemcitabine plus cisplatin therapy, produced a response rate of 41%. There was one complete response, lasting 14 months, and six partial responses. In the patients not previously radiated, the response rate was 57%; in the radiated patients, the response rate was 30%, with all responses occurring in the radiation Þeld. Ongoing Phase III studies have been designed to conÞrm the beneÞt of adding gemcitabine to cisplatin and to compare the combination with the other cisplatin combinations. Between December 1998 and June 2003, 57 patients with recurrent, bulky, stage II (21 patients), stage III (25 patients), or stage IVA (11 patients) squamouscell CaC were treated with six courses of gemcitabine (1,250 mg/m2 ) on days one and eight, in combination with cisplatin (70 mg/m2 ) on day one, every three weeks (Mutch DG, 2003). Forty-six patients were evaluable for response. The overall response rate was 50%: 15% complete responses, 35% partial responses, and 7% stable disease. The remaining 20 patients (43%) progressed during treatment. One- and two-year survival rates were 44% and 26%, respectively. Studies are also under way to investigate the effect of gemcitabine-containing doublets. A presentation at the recent American Society of Clinical Oncology (ASCO) meeting in New Orleans (Porras A, 2004) described a study that used gemcitabine (1,000 mg/m2 ) in combination with irinotecan (100 mg/m2 ) on days 1 and 8, repeating the cycle every 21 days. The overall response rate in this Phase I study with ten patients who had locally recurrent and/or metastatic disease was 33% (complete response rate 22%, partial response rate 11%). Hematologic toxicity, particularly anemia, was common, and one patient died of grade 4 diarrhea.

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The Gynecologic Oncology Group of the NCI is recruiting patients to a fourarm, multicenter, Phase III study in stage IVB CaC, as detailed on the group’s Web site (www.cancer.gov/clinicaltrials). The study is randomizing women. As of February 2006, this trial is still recruiting (aim for 600 patient in 4 years—150 in each of the 4 arms) to receive cisplatin in combination with one of these agents: gemcitabine, paclitaxel, topotecan, or vinorelbine. Physicians eagerly await the outcome of this study because it should deÞnitively answer the question about the optimal Þrst-line treatment for patients with mCaC. If the gemcitabine combination proves to be superior to the comparators, it may be established as the treatment of choice for mCaC and launch for this indication. Paclitaxel plus Ifosfamide plus Cisplatin (TIP) Regimen Overview. Cisplatin (BMS’s Platinol-AQ, Nippon Kayaku’s Randa, generics) and ifosfamide (BMS’s Ifex, Baxter’s Mitoxana/Holoxan, Shionogi’s Ifomide, generics) are regarded as active drugs in the treatment of CaC. Paclitaxel (BMS’s Taxol, generics) (Figure 8) also showed activity in a randomized trial, thus prompting investigation of combinations of all three agents. The TIP regimen (also known as the Zanetta regimen) is used, primarily in Europe, as a neoadjuvant treatment prior to hysterectomy. Studies have also been performed to investigate this regimen for the treatment of mCaC. The typical dose levels used in the TIP regimen are paclitaxel IV 175 mg/m2 , ifosfamide IV 5 g/m2 , and cisplatin IV 75 mg/m2 , repeated at three-week intervals. Mechanism of Action. •

•

•

Paclitaxel (Figure 8) is a taxane. Taxanes stabilize the mitotic spindle microtubules, formed during cell division, thus preventing depolymerization. This stability inhibits the dynamic reorganization of the microtubules, resulting in a mitotic block at interphase with a consequential inhibition of cell proliferation and eventual cell death. Cisplatin (Figure 3) is a platinum agent. Platinum agents generate highly reactive, charged platinum complexes that bind covalently to DNA, leading to intrastrand, interstrand, and protein cross-linking of DNA, resulting in inhibition of transcription and/or DNA replication mechanisms. Ifosfamide (Figure 6) is an alkylating agent pro-drug and a structural analogue of cyclophosphamide. After being metabolized in the liver to its active form, ifosfamide binds DNA and transfers alkyl groups, causing the formation of DNA cross-links and strand breaks. As a result, this agent prevents DNA replication and causes tumor cell death. The action of ifosfamide is cell-cycle nonspeciÞc. Ifosfamide is used almost exclusively with mesna, a compound that inactivates the alkylating activity and helps prevent the cystitis associated with this drug.

Clinical Performance. In a randomized trial, researchers compared two neoadjuvant regimens involving 182 patients with locally advanced squamous-cell

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CaC (Buda A, 2003). Patients randomly received three cycles of ifosfamide and cisplatin (IP) or ifosfamide, cisplatin, and paclitaxel (TIP). The TIP regimen yielded a slightly higher clinical response rate (89%) than the IP regimen (78%). Optimal response (residual tumor less than 3 mm) resulted signiÞcantly more often with TIP (43%) than with IP (22%). With a short follow-up, survival also tended to be better after TIP, though TIP caused grade 4 neutropenia in 37% of patients compared with only 16% of patients on the IP regimen. Four toxic deaths were observed, two in each group. Of the 182 patients enrolled, 164 completed chemotherapy and 178 underwent surgery. The aim of a second, Phase II trial was to assess the efÞcacy of TIP for persistent or recurrent squamous-cell CaC (Zanetta G, 1999). Forty-Þve women were treated with the TIP regimen; 31 had received prior radiation. Complete response was observed in 15 patients and partial response in another 15, yielding an objective response rate of 67% (95% conÞdence interval: 51–81%). Ten complete responders underwent subsequent surgery. The median survival for nonresponders was 6 months; for partial responders, more than 9 months; and for complete responders, more than 13 months. The most signiÞcant side effect was myelotoxicity; 91% of patients experienced grade 3 or 4 myelotoxicity. One woman had life-threatening toxic effects. The authors concluded that the TIP combination is highly effective for salvage treatment, particularly in unradiated patients (Zanetta G, 2000). For radiated women, the response rate is higher than response rates observed with other regimens, but further investigation is needed. Nonpharmacological Approaches Surgery is the mainstay treatment of localized, early-stage CaC—either simple extrafascial hysterectomy (the uterus and a small rim of vaginal cuff are removed from a plane outside the peritoneum) or colonization of the cervix for tumors less than 3 mm. Early invasive tumors larger than 3 mm are commonly treated by radical hysterectomy (the uterus and upper two-thirds of the vagina are removed). Early-stage disease with small tumors may be managed exclusively by radiotherapy (EBT and brachytherapy) rather than surgery; this choice of treatment may depend on the requirement to preserve fertility. With survival rates of up to 90%, radiotherapy and surgery are about equally effective as single options for treating very small CaCs in their earliest stages. Surgery is not routinely an option for locally advanced disease, which is treated by radiotherapy, usually with concurrent chemotherapy. Select patients with recurrent disease who have received maximum radiotherapy may be managed by extensive pelvic surgery for palliation, although this procedure is associated with urinary function complications. Studies have been conducted to investigate the potential therapeutic beneÞt of concurrent hyperthermia with radiotherapy and/or chemotherapy for the treatment of CaC. A Dutch Phase III trial demonstrated a survival and local control beneÞt gained from the addition of hyperthermia to chemoradiotherapy (Jones EL, 2003). The authors evaluated response and toxicity in patients with locally advanced CaC. All patients achieved clinical complete response and durable local control;

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the three-year survival rate improved from 27% to 51% with the addition of hyperthermia. The triple-modality treatment was well tolerated. Further trials are warranted to ascertain whether hyperthermia should be incorporated into standard treatment for CaC. EMERGING THERAPIES For several decades, the mainstay of drug treatment for cervical cancer (CaC) has been cisplatin. However, to achieve a major improvement in survival of patients with advanced CaC and to reduce the recurrence for stage I, II, and III patients, a new approach to treatment is required. Given that virtually all cases of CaC are caused by persistent infection with human papillomavirus (HPV), this virus has become an attractive target for new therapies. Indeed, vaccination against high-risk, CaC-related HPV subtypes has been under investigation for several years. These vaccines can be used in three settings: • • •

As population-wide preventive vaccines administered to females before they become sexually active. For women at high risk of developing CaC due to HPV infection with high-risk subtypes and/or cervical neoplasia or carcinoma in situ. For women who have been diagnosed with CaC, possibly as an adjuvant to surgery and likely in combination with standard chemotherapeutic agents.

Only the last of these uses is covered herein because it is the only one related to the direct treatment of CaC. Other approaches being tested to improve the outcome for CaC patients include new biological agents that target vascular endothelial growth factor (VEGF) and epidermal growth factor receptor (EGFR). In February 2004, the Þrst VEGFtargeted antibody, bevacizumab (Roche/Genentech’s Avastin), was approved for the treatment of metastatic colorectal cancer. GeÞtinib (AstraZeneca’s Iressa) and cetuximab (ImClone/Bristol-Myers Squibb [BMS]/Merck KGaA’s Erbitux), both EGFR-targeted agents, have been approved for non-small-cell lung cancer (NSCLC) and colorectal cancer, respectively. These developments have fueled interest in the use of targeted agents to treat CaC. Although CaC is relatively chemo-insensitive, new-generation cytotoxic agents are being tested in an attempt to improve patient outcome, primarily in CaC with distant metastases and often in combination with cisplatin. However, success has been elusive. For example, development of the protein kinase C inhibitor bryostatin (GPC-Biotech) was discontinued in March 2004 owing to lack of efÞcacy and a poor toxicity proÞle. This naturally occurring agent, isolated from the marine bryozoan Bugula neritina, had undergone Phase II testing, in combination with cisplatin, for advanced CaC. Based on data from the study, the authors concluded that the addition of bryostatin confers no beneÞt (Nezhat F, 2004). A report of a Phase I study of the natural product angiogenesis inhibitor TNP-470 (Abbott/Takeda’s Fumigillin) detailed a complete response in a patient

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previously treated with radiotherapy for stage IIIa squamous-cell carcinoma (Kudelka AP, 1998). On presentation with multiple pulmonary tumors but no evidence of local disease, she was treated with TNP-470 and remained in remission at eight months following completion of therapy. Another three patients in this trial experienced stable disease, but 14 patients progressed. A Phase II study failed to conÞrm the activity of TNP-470 in CaC. Because no further details have been released, we assume there is no further development of this agent at this time. Early clinical trials with agents such as docetaxel (Aventis’s Taxotere), an established treatment for other solid cancers, have been reported in the literature. A study of single-agent docetaxel in patients with metastatic CaC (mCaC) yielded a response rate of 19%. In the neoadjuvant setting, docetaxel produced a response rate of 34% in CaC patients, but only 17% became suitable candidates for surgery (Vallejo CT, 2003). Our research revealed no use of docetaxel in clinical practice in any of the markets under study at this time. A Phase I/II study is recruiting in the United States to examine the combination of docetaxel with carboplatin for the treatment of recurrent stage IVb CaC, based on promising Phase I data from a German study (Rein DT, 2002). Because slow progression is characteristic of CaC, retinoids have been investigated in an attempt to redifferentiate abnormal or neoplastic cervical cells. Retinoids may be used in either the preventive or therapeutic setting (Alvarez R, 2003). Also, retinoid receptors may interact with EGFRs. A study investigating the retinoid cis-retinoic acid (Ligand Pharmaceutical’s Aliretinoin) and interferon-alpha 2a (Roche’s Roferon-A) showed an overall response rate of 50% (12% complete response rate) in 32 previously untreated patients who had locally advanced, squamous-cell CaC. After a median response duration of three months, 9 of the 16 responders eventually progressed (Wilailak S, 2003; Braud AC, 2002). Table 6 summarizes the key emerging therapies in development for CaC. Vaccines Overview. Although the concept of a therapeutic tumor vaccine is viable, these therapies remain largely unproven in the treatment of cancer. Recent Phase III data subgroup analysis shows that Dendreon’s Provenge, a vaccine for the treatment of metastatic prostate cancer, has clinical activity in goodperformance status patients with less-aggressive tumor characteristics. Together with an improvement in survival (approximately eight months), this factor may stimulate renewed interest in initiating and recruiting patients for CaC therapeutic vaccine trials, particularly because CaC has an established viral cause. Provenge, shown to activate T cells, has fast-track status with the FDA and could be the Þrst therapeutic cancer vaccine to reach the market. Vaccines offer three major advantages over current treatments: • • •

They are generally well tolerated. The anticipated autoimmunity has not materialized as a clinical problem. They have convenient dosing regimens.

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TABLE 6. Emerging Therapies in Development for Cervical Cancer Development Phase

Marketing Company

— D —

— Xenova/Cancer Research Technology —

— II —

— Transgene —

Platinum agents Oxaliplatin United States Europe Japan

— II —

— Sanofi-Aventis —

Antimetabolites Capecitabine United States Europe Japan

II II —

Roche

II II —

AstraZeneca AstraZeneca —

III II —

Genentech/OSI/Roche Genentech/OSI/Roche —

II II —

Merck/KGaA/Takeda Merck/KGaA/Takeda —

Bioreductive agents Tirapazamine United States Europe Japan

II II I

Sanofi-Aventis Sanofi-Aventis Sanofi-Aventis

Efaproxiral United States Europe Japan

II — —

Allos Therapeutics — —

Compound Vaccines TA-HPV and TA-CIN United States Europe Japan MVA-HPV-IL-2 United States Europe Japan

EGFR-targeted agents Gefitinib United States Europe Japan Erlotinib United States Europe Japan EMD-72000 United States Europe Japan

—

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If vaccines are shown to be efÞcacious at any stage of CaC, they will be rapidly adopted and widely used. One approach that has attracted attention is the development of preventive vaccines targeted at high-risk subtypes of HPV. Such vaccines could be used to immunize girls prior to becoming sexually active; Phase III clinical trials are underway. The attraction of preventive vaccines would be greatest in the developing world, where the incidence of CaC is high and screening is not economically viable. Success in this arena could also lead to the replacement of Pap smear tests in countries where they are currently established, and preventive vaccination might have better penetration and compliance than Pap tests. Additionally, HPV-directed vaccines could be used for the treatment of cervical intraepithelial neoplasia (CIN) and so reduce the likelihood of progression to invasive CaC. In short, development of a successful vaccine could dramatically affect the incidence of CaC. Most vaccines in development for CaC target one or both of the two oncogenic proteins, E6 and E7, of the highly oncogenic subtypes of HPV (types 16 and 18), which are highly associated with the disease. Such vaccines have been extensively studied in the preventive setting and are designed to induce an antibody-mediated response (Adams M, 2001). Other multivalent vaccines have been developed that target as many as four HPV subtypes in an attempt to maximize protection against the most common cancer-causing agents. Given the time scale of CaC development following HPV infection, it will be some decades before a preventive vaccine will affect the incidence of CaC, even if widespread vaccination were undertaken. Endpoints of preventive studies may be either presence of HPV infection (detected by DNA analysis) or cervical neoplasia; either will enable more rapid assessment of vaccine efÞcacy than the endpoint of invasive CaC development. Longer-term follow-up is required to assess the effect of vaccination on the development of CaC and to determine its efÞcacy as a cancer preventive agent. A vaccine derived from HPV type 16 has been shown to reduce the incidence of HPV type 16 infection and its associated CIN (Koutsky L, 2002). In a doubleblind Phase III study performed in the United States, 2,392 sexually active women aged 16–23, with no history of an abnormal Pap smear test result, were given three doses of either the HPV type 16 vaccine or placebo. Participants were dosed with vaccine on day 0, at 2 months, and at 6 months; they were tested for HPV type 16 infection at enrollment, at 1 month after the third vaccination, and every 6 months subsequently. Median follow-up was 17.4 months after completion of the vaccination regimen. Of the 41 cases of new HPV type 16 infection identiÞed in the study, including 9 cases of HPV-related CIN, all occurred in women in the placebo group, thus demonstrating the vaccine’s 100% effectiveness in this study. In May 2004, GlaxoSmithKline (GSK) announced the launch of a doubleblind, placebo-controlled, Phase III clinical trial, the Papilloma Trial to Prevent Cervical Cancer in Young Adults (PATRICIA), which aims to recruit 13,000 women aged 15–25 years. The vaccine, developed in collaboration with MedImmune (MEDI517, Cevarix), targets HPV types 16 and 18 and contains a novel

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proprietary adjuvant, AS04, which contains 3’-deacylated monophosphoryl lipid A and alum to enhance the immune response. Another approach to vaccine therapy is to vaccinate women who have CIN or carcinoma in situ. In these indications, HPV infection is prevalent, and clinical trials have shown that vaccines can reverse neoplasia. If proved effective in this setting, vaccines could eliminate the need for surgery and would enjoy rapid uptake—even conservative surgery such as conization can lead to impaired fertility and the ability to carry a baby to full term. Therapeutic vaccines must induce cell-mediated immunity because antibodies cannot eliminate the virus once it has become intracellular. Transgene recently reported data from a Phase II clinical study of its vaccine, MVA-HPV-IL-2, in high-grade CIN (grades 2 and 3). MVA-HPV-IL-2 is a recombinant virus that expresses HPV type 16 proteins E6 and E7 and interleukin-2 (IL-2). Results showed clinical and histological responses in 5 of 15 patients. The company is planning a new trial to test the highest vaccine dose in this patient population. Data for MVA-HPV-IL-2 in the treatment of CaC are discussed further on. Xenova has been developing two vaccines directed against HPV: TA-HPV and TA-CIN. Preclinical studies have shown that TA-CIN can induce a cell-mediated response, in the absence of an adjuvant; this formulation has been extended into clinical study. A Phase I, double-blind, randomized, placebo-controlled, and doseescalating safety and immunogenicity trial that enrolled 40 women was completed in July 2000. TA-CIN was found to be very well tolerated; no serious adverse events were reported during the trial. It was also conÞrmed to be immunogenic (Knutson KL, 2001). We describe clinical studies of TA-HPV and TA-CIN for the treatment of CaC further on. Mechanism of Action. Cancer vaccines are designed to stimulate the immune system to launch a response against the speciÞc target contained by the vaccine. Vaccines for CaC have targeted highly oncogenic viral proteins expressed by HPV. Some vaccines also express cytokines, such as IL-2, that act as immunostimulants in order to augment the host response to vaccination. TA-HPV and TA-CIN. Xenova, in conjunction with the European Organization for the Research and Treatment of Cancer (EORTC), was codeveloping TA-HPV and TA-CIN for the prevention and treatment of CaC, however this has now been discontinued. TA-HPV vaccine targeted E6 and E7 proteins of HPV types 16 and 18. TA-CIN targeted these proteins as well as the L2 component of HPV types 16 and 18. MVA-HPV-IL-2. Transgene’s MVA-HPV-IL-2 is a recombinant virus vaccine that expresses the HPV type 16 proteins E6, E7, and IL-2. This vaccine is in Phase II development for CaC in Europe. No development is underway in the United States or Japan for this indication. The rationale behind this agent is to induce a speciÞc T-lymphocyte-mediated response against the E6 and E7 HPV

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proteins and a nonspeciÞc IL-2 immune-stimulatory response. MVA-HPV-IL-2 is also under investigation for the treatment of CIN and vulva intraepithelial neoplasia. A Phase II clinical study conducted in Mexico that investigated MVA-HPVIL-2 for the treatment of CaC in 27 patients treated with a high dose (5 × 106 units) has been reported (www.transgene.fr). Two patients had stable disease at a minimum of six months’ follow-up (primary efÞcacy criteria). However, the clinical activity observed did not reach the efÞcacy threshold outlined in the protocol, which was amended to allow patients to receive chemotherapy following disease progression during the study. Two partial responses (seven months) and two stabilizations, out of Þve patients, have been reported. Based on the results of this study, the company concluded that the efÞcacy demonstrated was not sufÞcient to justify continuing the development of MVA-HPV-IL-2 as a monotherapy. Studies of MVA-HPV-IL-2 in combination with chemotherapy have not been identiÞed. Investigation of MVA-HPV-IL-2 is continuing for the treatment of the precancerous indications, CIN, and vulva intraepithelial neoplasia. Platinum Agents Overview. Cisplatin is the backbone of chemotherapy for the treatment of CaC. In some situations, however, despite the lack of studies showing equivalence, carboplatin is used in place of cisplatin because of the former’s improved toxicity proÞle. Research is ongoing to identify novel platinum agents with improved activity against CaC. To date, nedaplatin (Shionogi’s Aqupla), approved in Japan, has demonstrated clinical beneÞt, and oxaliplatin, approved in the United States and Europe for the treatment of colorectal cancer, is being investigated. Mechanism of Action. Platinum agents generate highly reactive, charged platinum complexes that bind covalently to DNA, leading to intrastrand, interstrand, and, less importantly, protein cross-linking of DNA. These cross-links result in inhibition of transcription and/or DNA replication mechanisms. The principal sites of reaction are the N7 atoms of guanine and adenine. The formation of intrastrand cross-links between platinum and neighboring guanines has been shown to correlate with clinical response. When used in conjunction with radiotherapy, platinum agents may also act as radiosensitizers. Possible mechanisms of radiosensitization include inhibition of repair of sublethal radiation damage and synchronization of tumor cells into a radiosensitive phase of the cell cycle. Oxaliplatin. Oxaliplatin (SanoÞ-Aventis’s Eloxatin) (Figure 10), a platinumbased chemotherapeutic agent with a 1,2-diaminocyclohexane (DACH) carrier ligand, has been approved in all the major markets for the treatment of metastatic colorectal cancer. It recently received approval in the United States as an addition to 5-ßuorouracil (5-FU) for adjuvant treatment after surgery for colorectal cancer. Oxaliplatin has shown in vitro and in vivo efÞcacy against many tumor cell lines, including some that are resistant to cisplatin and carboplatin, a discovery that suggests it has potential utility in cisplatin-refractory CaC. The retention of

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NH2

O

O

O

O

Pt NH2

FIGURE 10. Structure of oxaliplatin.

the bulky DACH ring by activated oxaliplatin is thought to result in the formation of platinum-DNA adducts, which appear to be very effective at blocking DNA function (Raymond E,1998). A Phase II study was conducted to determine the efÞcacy of oxaliplatin therapy in patients with previously treated squamous-cell carcinoma of the cervix (Fracasso PM, 2003). Patients who had received a maximum of one previous chemotherapeutic treatment were treated with oxaliplatin at 130 mg/m2 every 21 days. Twenty-three of the 24 patients entered in the study had received prior cisplatin or carboplatin. One complete response lasting 2.2 months and one partial response lasting 3.2 months were achieved. Another nine patients had stable disease. The most frequently reported drug-related toxicities were anemia, nausea and vomiting, and neurotoxicity. Three patients had a grade 3, infusion-related allergic response that was resolved mainly by increasing oxaliplatin infusion time. Oxaliplatin has only modest activity against cisplatin-resistant or carboplatin-resistant disease. Antimetabolites Overview. First launched in 1959, the antimetabolite 5-FU has proven, though modest, efÞcacy against CaC. It is used concurrently with cisplatin or immediately after radiotherapy, and it is used for the treatment of disseminated disease. The evidence of 5-FU activity provides the rationale for testing new-generation antimetabolites such as capecitabine, a tumor-activated pro-drug of 5-FU with a better side-effect proÞle. Mechanism of Action. Antimetabolites are structural analogues of naturally occurring compounds. Antimetabolites interfere with the production of nucleic acids. They work through a variety of mechanisms, including competition for binding sites on enzymes and incorporation into nucleic acids. The three categories of antimetabolites, based on their sites of action, are as follows: antifolates, purine analogues, and pyrimidine analogues. Capecitabine. Capecitabine (Roche’s Xeloda) (Figure 11) is an orally active pro-drug of the ßuoropyrimidine 5-FU approved for the treatment of metastatic breast cancer and colorectal cancer; it is under extensive investigation in other cancer indications, including adjuvant use. Capecitabine is selectively tumoractivated to its cytotoxic moiety, ßuorouracil, by thymidine phosphorylase (TP). Within normal and tumor cells, ßuorouracil is further metabolized to two active metabolites, 5-ßuoro-2-deoxyuridine monophosphate (FdUMP) and 5ßuorouridine triphosphate (FUTP). FdUMP inhibits DNA synthesis by reducing

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O CH3 HN

O F

N O H3C

HO

O

N

OH

FIGURE 11. Structure of capecitabine.

normal thymidine production; FUTP inhibits RNA and protein synthesis by competing with uridine triphosphate. The active moiety of capecitabine, ßuorouracil, is speciÞc to the cell-cycle phase (S phase). In one case study, a patient with mCaC who was treated with one course of capecitabine evinced a response that resulted in extended survival (Hindenburg AA, 2003). However, the patient experienced severe toxicity, including grade 4 hand and foot syndrome, a side effect commonly associated with capecitabine. An interesting study by a group of Japanese investigators has shown that TP levels in cervical tumors are elevated following radiotherapy, a Þnding that suggests CaC may be amenable to treatment with capecitabine (Oguri H, 2004). A multicenter Phase II study in the United States is recruiting patients with persistent or recurrent adenocarcinoma of the cervix to receive capecitabine as a single agent following failure of Þrst-line therapy (www.cancer.gov). Patients receive oral capecitabine twice daily on days 1–14. Courses repeat every 21 days in the absence of disease progression or unacceptable toxicity. No results have been reported at the time of writing this report. EGFR-Targeted Agents Overview. Cervical intraepithelial dysplasia and invasive cancers have been shown to possess increased levels of EGFR; 90% of CaC tumors reportedly overexpress the receptor (Boiko I, 1998). In addition, levels of expression correlate with the stage of cancer and degree of HPV infection (Kersemaekers AM, 1999). Interestingly, evidence is accumulating for a nicotine-induced upregulation of EGFR in CaC cell lines, with a resultant increase in cell proliferation (Mathur RS, 2000[a]). These observations suggest that EGFR is a valid therapeutic target for CaC therapies as well as a potential marker for the disease (Mathur RS, 2000[b]). A vast amount of research and development is ongoing in the area of EGFR inhibition. Two main approaches are being investigated: the speciÞc inhibition of EGFR-associated tyrosine kinase and monoclonal antibodies (MAbs) directed at the external domain of EGFR itself. To date, geÞtinib (AstraZeneca’s Iressa), a

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small-molecule EGFR tyrosine kinase inhibitor, has been approved for the thirdline treatment of NSCLC, and cetuximab (ImClone/Merck KGaA/Bristol-Myers Squibb’s Erbitux), an EGFR-directed MAb, has been approved for second-line use, in combination with chemotherapy, for colorectal cancer. Also, recent data have shown that cetuximab provides a signiÞcant survival beneÞt when administered with radiotherapy for the treatment of head and neck cancer, a squamouscell malignancy. This information may be relevant for the treatment of CaC, particularly CaC of squamous-cell origin, which is also treated by radiotherapy. Mechanism of Action. EGFR is a 170-kilodalton transmembrane glycoprotein encoded by the (c-erbB-1 ) proto-oncogene. EGFR comprises three domains with distinct functions: extracellular ligand-binding domain (binds to different EGFrelated growth factors); transmembrane domain (anchors the receptor in the cell membrane); and cytoplasmic tyrosine kinase domain (activates the intracellular response cascade). Therapeutic intervention of the EGFR is either by MAbs that competitively bind to the extracellular receptor or by small-molecule inhibitors of the receptor-associated tyrosine kinase. Gefitinib. GeÞtinib (AstraZeneca’s Iressa) is a small-molecule inhibitor of EGFR tyrosine kinase. It launched in the United States and Japan as a singleagent, third-line treatment for NSCLC and is in Phase II development for CaC in the United States and Europe. GeÞtinib is a competitive inhibitor of the ATPbinding site and blocks ligand-induced EGFR activation, inhibiting EGFR-driven cell proliferation. Data from a Phase II trial involving patients with advanced or metastatic squamous-cell CaC were presented at the American Society of Clinical Oncology (ASCO) meeting in 2003 (Viens P, 2003). Twenty-three patients received 500 mg of geÞtinib each day for a median of 1.8 months. Six patients had stable disease that lasted 25–114 days. Twelve patients had progressive disease; the median time to disease progression was 47 days. A Phase II study, underway in the United States, is recruiting CaC patients (and patients with epithelial ovarian cancer) to receive single-agent geÞtinib and to determine any reduction in phosphorylation of EGFR (or other molecules downstream on the same signaling pathway) and whether this reduction correlates with clinical response. At this time, it has not been possible to identify additional clinical data for geÞtinib in CaC; it does not appear to be a focus of AstraZeneca’s development strategy. The relationship between EGFR expression and response to geÞtinib took an interesting turn with the publication of two papers showing that geÞtinib responders express a mutant form of the EGFR tyrosine kinase (Paez JG, 2004; Sordella R, 2004). However, early data suggest that CaC samples do not express the mutation. Erlotinib. Erlotinib (OSI Pharmaceuticals’ Tarceva), a small-molecule EGFR tyrosine kinase inhibitor with a mechanism of action similar to that of geÞtinib,

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is under development by OSI in alliance with Genentech and Roche. This orally active quinazoline derivative is in Phase III trials in the United States and Europe and in Phase I trials in Japan with Roche subsidiary Chugai for NSCLC cancer. A Phase II, multicenter study of erlotinib is recruiting patients with persistent or recurrent squamous-cell CaC. Patients will receive oral erlotinib once daily until evidence of disease progression or unacceptable toxicity. The drug has been approved in Europe. EMD-72000. EMD-72000 (Merck KGaA/Takeda’s matuzumab) is a humanized IgG1 antibody directed at the EGFR. EMD-72000 competes for binding at the receptor with the EGFR’s natural ligands, EGF and tumor growth factor-alpha. By February 2004, a Phase II study named EMD 72000-023 had enrolled 41 CaC patients in Germany, the United Kingdom, and the Netherlands. The study is still ongoing at the time of writing. Interestingly, cetuximab (ImClone/BristolMyers Squibb/Merck’s Erbitux) has been approved for the treatment of refractory metastatic colorectal cancer; its approval was based on improved response rate but, as yet, no proven survival beneÞt. This precedent may increase the likelihood of EMD-72000’s approval for CaC, an indication with a signiÞcant unmet treatment need. Bioreductive Agents Overview. Bioreductive, or hypoxia-selective, cytotoxins represent a promising new class of drugs that may have single-agent activity or, most likely, will be combined with other cytotoxic agents or with radiotherapy. Tirapazamine is a lead compound within this class and has been under investigation for solid tumors, including CaC, head and neck cancer, and NSCLC. Data showing lack of additional activity and increased toxicity when added to standard agents have lowered the chances for tirapazamine’s success. Mechanism of Action. Bioreductive agents are unique because they exhibit selective cytotoxicity toward hypoxic cells (Peters KB, 2001). This pharmacological strategy is designed to exploit the fact that tumor cells are typically less well oxygenated than healthy cells. This characteristic contributes to tumor progression and to radiotherapy and chemotherapy resistance in a variety of ways, such as activation of certain signal transduction pathways and gene regulatory mechanisms. In the human body, metabolism of the active drug turns it into a toxic free radical when the compound undergoes one-electron reduction by cytochrome P450 reductase. In hypoxic cells, the resulting free radical induces single-strand and double-strand breaks in DNA, whereas in well-oxygenated tissues, the free radical is further oxidized and is transformed into an inactive metabolite. Tirapazamine. Tirapazamine (SanoÞ-Aventis’s Tirazone) (Figure 12) is the lead agent in a new class of bioreductive drugs known as benzotriazine diN-oxides (Peters KB, 2002). It is in Phase II trials in the United States and

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FIGURE 12. Structure of tirapazamine.

Europe and in Phase I development in Japan for the treatment of CaC. Based on promising activity in a Phase II trial, a Phase III study is underway in Australia, investigating tirapazamine in combination with cisplatin and radiotherapy for the treatment of squamous-cell head and neck cancer. The Þnal data are expected in 2007. Cervical cancers are known to be hypoxic tumors because of the development of necrotic areas in even relatively small tumors. Radiation and chemotherapy failures have been attributed to tumor hypoxia because tumor oxygenation is critical for the free-radical formation that causes DNA damage. In preclinical models, tirapazamine enhances both radiation and chemotherapy effectiveness. Data from a Phase I/II clinical study in women with locally advanced CaC have been described (Craighead PS, 2000). Patients received standard radiotherapy and cisplatin treatment to which escalating doses of tirapazamine were added. The study identiÞed a tirapazamine dose level of 290 mg/m2 , given with a cisplatin dose on days 1, 15, and 29, plus a dose of tirapazamine at 220 mg/m2 given on days 8, 10, 12, 22, 24, and 26 as the maximum tolerated dose with acceptable toxicity. Radiotherapy was given daily on days 1–5, 8–12, 15–19, 22–26, and 29–33. At six months, 13 of 15 patients had complete pelvic control of disease. The authors concluded that the use of tirapazamine with concurrent cisplatin and pelvic radiotherapy has acceptable toxicity and should be considered for further Phase II testing. Results from an earlier Phase I study coordinated by the NCI in the United States were also presented (Aghajanian C, 1997). This study combined tirapazamine with cisplatin in 12 heavily pretreated patients with recurrent CaC. Tirapazamine was administered as an intravenous (IV) infusion over two hours, followed one hour later by a one-hour infusion of cisplatin at 75 mg/m2 . A maximally tolerated dose of 330 mg/m2 of tirapazamine was determined for the schedule described, and all observed responses were seen at this dose level. Despite the use of prophylactic antiemetic, the dose-limiting toxicity was nausea and vomiting. All 12 patients were evaluated for response. Two major responses were seen (17%). In addition, there were three minor responses (25%) and four patients achieved disease stabilization (33%). All major and minor responses were seen at the highest dose level tested: 330 mg/m2 . Details of tirapazamine in other Phase II or Phase III studies in CaC have not been identiÞed, and interest in its utility in NSCLC has waned. However, studies

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FIGURE 13. Structure of efaproxiral.

of tirapazamine in conjunction with radiotherapy are ongoing in head and neck cancer, and it is this radiosensitizer role that holds most interest for clinicians. Efaproxiral. In August 2002, Allos Therapeutics initiated a Phase I/II clinical trial of efaproxiral (Efaproxyn in the United States, Revaproxyn outside the United States) (Figure 13), a synthetic small molecule, for patients with locally advanced CaC who were receiving cisplatin-based chemoradiotherapy. The trial enrolled patients in the United States and Canada. Efaproxiral is an allosteric modiÞer of hemoglobin that enhances the diffusion of oxygen to hypoxic (oxygen-deprived) tumor tissues, thereby making radiation therapy more effective. Allos’s lead indication for efaproxiral is the treatment of brain metastases from breast cancer with whole brain irradiation; this breast cancer indication resulted from a subpopulation analysis of Phase III data that showed no signiÞcant improvement in survival for patients with brain metastases from other types of primary tumor. The FDA awarded efaproxiral orphan drug status for brain metastases of breast cancer and an approvable letter that states, “If the study shows effectiveness in this population (increased survival) using the pre-speciÞed analysis, and the study is otherwise satisfactory, we believe it would, together with the subset result in the earlier clinical study, support approval.” The ongoing Phase I/II clinical trial is an open-label, multicenter study of efaproxiral administered to patients with stage IIIa or IIIb CaC who are receiving a course of weekly cisplatin with both external beam therapy (EBT) or brachytherapy. The goal of the Phase I part of the study is to assess the safety and tolerability of escalating doses of efaproxiral in this combination and to determine the maximum tolerated dose (MTD) of efaproxiral in patients with CaC. The objective of the Phase II part of the trial is to further evaluate the safety proÞle and to assess the efÞcacy of efaproxiral at the MTD in combination with cisplatin and radiation therapy, to be determined by the disease’s progression rate at two years. The trial is expected to take about two years to complete enrollment. Efaproxiral is also showing promise in other indications.

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Chronic Lymphocytic Leukemia (CLL)

ETIOLOGY AND PATHOPHYSIOLOGY Overview Chronic lymphocytic leukemia (CLL) is the most common leukemia in the Western world but is rare in Japan and other Asian countries. In 95% of chronic lymphocytic leukemias, the affected cells are B lymphocytes; in the remaining 5%, T lymphocytes are affected. CLL is further differentiated according to morphological, immunophenotypical, cytogenetic, and molecular characteristics, and the clinical course and treatment of these subtypes vary widely. Pathophysiology Mature blood cells (red cells, white cells, and platelets) are produced in the bone marrow from pluripotent hematopoietic stem cells. (Figure 1 shows the different lineages of blood cells and the stages involved in their maturation.) The blood cells mature and differentiate through a sequence of steps involving a series of complex—and incompletely understood—interactions with growth factors, cytokines, and other cells in the bone marrow. Once mature, the blood cells leave the bone marrow and enter the general circulation, where they have a limited life span. T and B lymphocytes (often known as T and B cells) are white blood cells, and they are vital constituents of the immune system. When the body is infected with a pathogen, T and B cells are mobilized to target and kill the pathogen. Although other immune cells are involved in this process, lymphocytes possess Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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Pluripotent stem cell in bone marrow

B stem cell

T stem cell

Pre-B cell

Prothymocyte

Dendritic stem cell

Dendritic cell

Myeloid stem cell

Precursor colony-forming cells

Neutrophil B lymphocyte T lymphoblast Megakaryocyte Basophil Erythrocyte Monocyte

Platelets

Eosinophil

B cell T cell

Macrophage

Plasma cell

FIGURE 1. The hematopoietic cascade: development of mature blood cells from pluripotent hematopoietic stem cells.

unique qualities that allow them to adapt during infection and become speciÞc to the invading pathogen. In addition, once an infection has been cleared from the body, most of these lymphocytes die, but a few “memory” lymphocytes remain. Upon future reinfection with the same pathogen, these cells are immediately activated, so each successive immune response is therefore quicker and more accurate than the one before. Cancer occurs when a series of genetic mutations, which are hereditary and/or environmental in nature, takes place in a single cell. The resulting cell proliferates without control and disrupts the normal functioning of the organ in which it originates. A lymphoid leukemia results when this process occurs in a T or B

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lymphocyte, and the expanding malignant cells disrupt the balance of normal blood cells in the bone marrow, blood, and lymphoid organs. Lymphoid leukemias can be acute or chronic and can arise from lymphocytes at different stages of development. CLL is a malignancy of small, morphologically mature but immunologically less mature lymphocytes that accumulate in the blood, bone marrow, lymph nodes, spleen, and liver. No single deÞned genetic mutation or abnormality is solely responsible for all CLL cases; instead, this disease is characterized by an array of different chromosomal deletions (described later). Investigators have also described a multitude of aberrations associated with apoptotic proteins, suggesting that in most cases, CLL cells accumulate as a result of an abnormally long life span rather than an accelerated rate of proliferation. The median age of onset of CLL is 65–68 years (Call TG, 1994), although approximately 20% of cases occur in people younger than age 55. Survival is dependent on the stage of disease and ranges from a median of 12 years in patients with the earliest stage to 2–5 years in those with advanced disease (Pangalis GA, 2002). CLL also tends to occur more in men than in women. Onset is usually insidious, and up to 70% of patients are asymptomatic. Diagnosis is often made following routine blood tests or investigations for an unrelated disease. In symptomatic patients, the most frequent clinical Þndings are fatigue, loss of appetite, weight loss, and enlarged lymph nodes. Table 1 lists laboratory features typical of CLL. CLL has no single pattern of progression—approximately one-third of patients never require treatment and die from causes unrelated to CLL; one-third have an initial indolent phase followed by progression; and the remaining third have aggressive disease at the outset requiring immediate therapy. CLL’s progressive symptoms are related to leukocyte inÞltration of the bone marrow, spleen, and lymphoid tissue. This inÞltration, together with disruptions in normal hematopoietic function, results in anemia, neutropenia, thrombocytopenia, and immunological dysfunction. The most important immunological dysfunction is the lack of immunoglobulins (hypogammaglobulinemia), a condition that occurs in up to 60% of patients with advanced disease. This condition leaves the patient more susceptible to infection, which is a primary cause of death and morbidity. Autoimmune disease, an immune response against the body’s own cells and tissues, occurs in 10–35% of untreated CLL patients. This condition is a natural complication of CLL but is also associated with purine analogue therapy (a common treatment for CLL); however, the exact frequency of treatment-induced autoimmune disease is unknown. The autoimmunity usually manifests as autoimmune hemolytic anemia. Immune thrombocytopenia, pure red-cell aplasia, and immune neutropenia occur less frequently. Autoimmunity is usually successfully treated with corticosteroids; steroid-refractory patients may have to undergo a splenectomy. In 3–10% of patients with CLL, the disease undergoes a transformation into a more aggressive condition distinct from CLL. The transformation is usually

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TABLE 1. Laboratory Features Characteristic of Chronic Lymphocytic Leukemia Diagnostic Modality

Features Characteristic of CLL

Blood count

• >5,000 lymphocytes/microliter of peripheral blood. • Anemia and thrombocytopenia present at the time of initial diagnosis in approximately 20% of patients; both are usually mild, but their presence denotes a poor prognosis. • Polyclonal increases in gamma globulins (present in approximately 15% of patients). • Hypogammaglobulinemia (8%). • Autoimmune thrombocytopenia (3%). • Pure red-cell aplasia (0.5%). • Agranulocytosis (0.5%).

Blood smear

• A large number of small, morphologically matureappearing lymphocytes are visible; the nucleus is large, a nucleolus usually not evident, and only a thin band of cytoplasm is evident (a small proportion of lymphocytes may be larger with a larger nucleus and a visible nucleolus). • ‘‘Smudge cells’’ (ruptured cells) may be visible; these are lymphocytes that appear flattened or smudged in the process of slide preparation. • When leukocyte counts are extremely high (in excess of 200,000/mL), whole blood viscosity may be abnormally high.

Bone marrow biopsy

• The proportion of mature-appearing lymphocytes in the bone marrow aspirate exceeds 30% of all nucleated cells. • Infiltrative patterns of lymphocytes that may be nodular (10%), interstitial (30%), or diffuse (35%). A mixture of infiltrative patterns is observed in 25% of patients. • The infiltration pattern is significant in determining prognosis; diffuse infiltration is associated with advanced disease and poorer prognosis, whereas nodular and interstitial patterns (nondiffuse) are associated with lessadvanced disease and better prognosis.

Cytogenetic/molecular analysis

• Cytogenetic analysis is undertaken in research settings only.

Immunophenotypical analysis

• Low levels of surface immunoglobulin, only a single light chain. • Expression of one or more B-cell-associated antigens —CD19, CD20, and CD23. • Coexpression of CD5.

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into large-cell lymphoma (also known as Richter’s syndrome), and the prognosis for these patients is poor—median survival is six months. Transformation into prolymphocytic leukemia occurs occasionally, and transformation into acute leukemia is rare. Classification. The World Health Organization (WHO) published a consensus classiÞcation system that categorizes lymphoid disorders based on morphology, immunophenotype, genetic features, and clinical characteristics (Harris NL, 1994). Under the WHO classiÞcation system, CLL and small lymphocytic lymphoma (the lymphoid form of CLL) are grouped into a single entity (CLL). Advances in monoclonal antibody and ßow cytometry technology have established immunophenotyping as a routine diagnostic test for CLL. Using these methods, CLL is easily distinguished from other B-cell neoplasms because the cells aberrantly express CD5, a T-cell marker. Staging. Three major staging systems exist for the classiÞcation of CLL. (These systems are described in Table 2.) The original Rai system, published in 1975, consists of stages 0-IV and is based on the presence of lymphadenopathy, organomegaly, and cytopenias (Rai K, 1975), demonstrating a correlation between Rai stage and survival. This system was later modiÞed from the Þve-tier TABLE 2. Common Staging Systems Used in the Treatment of Chronic Lymphocytic Leukemia System Rai staging system

Stage

Definition

0 I II III

Lymphocytosis only Lymphocytosis and lymphadenopathy Lymphocytosis, spleen or liver enlargement Lymphocytosis and anemia (hemoglobin <11 g/dL) Lymphocytosis and thrombocytopenia (platelet count <100,000 mL)

IV Modified Rai staging system

Binet staging system

Low risk of progression Intermediate risk of progression High risk of progression A

B C

Rai stage 0 Rai stage I or II Rai stage III or IV Lymphocytosis, with enlargement of <3 lymphoid areasa ; no anemia or thrombocytopenia Lymphocytosis, with enlargement <3 lymphoid areas Lymphocytosis and either anemia (hemoglobin <10 g/dL) or thrombocytopenia (platelet count <100,000/mL), or both

a. The following lymphoid areas are included: cervical, axillary, inguinal (whether unilateral or bilateral), spleen, and liver.

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system to a three-tier system that categorizes patients as having a low, intermediate, or high risk of disease progression. The Binet classiÞcation system (A, B, C) was devised based on a retrospective analysis of disease burden that draws a correlation between the number of nodal groups involved with disease and bone marrow failure (Binet JL, 1981). The National Cancer Institute (NCI)’s guidelines for CLL state that the major distinctions and beneÞts of the Binet system derive from its recognition that (1) a predominantly splenic form of the disease may have a better prognosis in the Binet system than in the Rai systems, (2) patients with comorbid anemia or thrombocytopenia have a similar prognosis, and (3) the presence of either of these two conditions can be grouped in the same stage rather than in separate stages (National Cancer Institute, 2003). Although both the Rai and Binet systems group patients according to their risk of progression, the early stages of disease do not correspond well between the two systems. Binet’s good prognosis group, A, includes twice as many patients as Rai’s stage 0 because it includes all Rai stage 0, two-thirds of Rai stage I, and one-third of Rai stage II (Dighiero G, 2002). The original and modiÞed Rai systems are used throughout the United States; all three systems are used in Europe, although physicians quote Binet more frequently. The overlap among staging systems has made the comparison of clinical trials using different staging systems difÞcult. Although the Rai and Binet staging systems each give general indications as to a patient’s prognosis, survival within each stage can vary signiÞcantly, particularly in those patients with Binet stage A and Rai stage 0. As many as 30% of these patients have “smoldering” CLL, which progresses slowly and never requires therapy; other patients have more progressive disease that will eventually require treatment and may be fatal. The median survival of patients with Rai stage 0 exceeds 12 years and may reach 20 years with a 10-year overall survival rate of 70–75%. Patients with Rai stage I and II have a median survival of 8–10 years and 5–8 years, respectively, whereas recent data show a median survival of 5 years and longer in Rai stage III and IV patients (Pangalis GA, 2002). Prognostic Factors. CLL is heterogeneous in both molecular pathology and clinical course, and many questions regarding the most appropriate management of this disease remain unanswered. Researchers continue to investigate both biological and clinical parameters in an attempt to reÞne the staging systems and accurately determine the prognostic outcome for patients with CLL and to individualize treatment. We discuss the most common markers further on, but many other markers, including interleukin (IL)-6, IL-10, tumor necrosis factor, intracellular BCL-2, vascular endothelial growth factor (VEGF), and other cytokines and enzymes, are being investigated for their potential prognostic signiÞcance. Lymphocyte Doubling Time. The lymphocyte doubling time (LDT) is calculated as the number of months it takes the absolute lymphocyte count to double in number. This factor has been conÞrmed as a prognostic indicator that is independent of stage in a variety of studies (Shanafelt T, 2003). After a 118-month

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follow-up, median survival was 61 months in patients with an LDT of less than 12 months, and median survival was not yet reached in patients with an LDT of more than 12 months (Montserrat E, 1986). According to published literature, this factor is subject to variation (Shanafelt T, 2003). Serum Beta-2 Microglobulin. The serum marker beta-2 microglobulin correlates with tumor burden and disease stage in patients with CLL. A retrospective study from the M.D. Anderson Cancer Center (Houston, TX) found this serum marker to be the strongest predictor of Þve-year survival in a multivariate analysis that controlled for age, sex, and performance status (Keating MJ, 1995). Similar results have been reported in other retrospective trials; however, a prospective trial did not Þnd beta-2 microglobulin to be a signiÞcant predictor of survival in a multivariate analysis that controlled for stage and LDT (Molica S, 1999). Additional studies are required to further classify the role of beta-2 microglobulin in prognosis. Immunoglobulin Mutation Status. The B-cell antigen receptor, or immunoglobulin (Ig), is a surface molecule that detects the presence of pathogenic antigens and activates an immune response. Its synthesis during B-cell development occurs via a process of gene rearrangement. Gene segments are selected and assembled in such a way that each B lymphocyte produces an Ig that is unique to that cell. When the B lymphocyte encounters an antigen, the Ig molecule is altered by a process known as somatic hypermutation, which makes the Ig more speciÞc for the antigen, thus enhancing the immune response. The presence of mutations in an Ig is evidence of its encounter with antigen and a marker of B-cell maturity. CLL was initially thought to be a malignancy of antigen-naive B cells with unmutated Ig. Researchers subsequently found that a subset of CLL patients had mutated Ig, suggesting the cell of origin was more mature and had encountered antigen. In 1999, two studies simultaneously reported the prognostic signiÞcance of Ig mutation status in CLL (Damle RN, 1999; Hamblin TJ, 1999). The studies demonstrated that patients with a mutated Ig status had signiÞcantly longer survival than those with unmutated Ig. Many studies have conÞrmed the prognostic signiÞcance of Ig mutation, and researchers have established correlations between Ig mutation status and the need for chemotherapy, the response to chemotherapy, and the risk of relapse after transplantation (Shanafelt T, 2003). Serum Thymidine Kinase. Thymidine kinase is an enzyme involved in the DNA synthesis salvage pathway, a process required for the synthesis of new DNA precursors in dividing cells. Present in dividing cells and absent in resting cells, it is a marker of proliferation. A recent study demonstrated the relationship between serum thymidine kinase level and mutation status: a level greater than 15 U/L was a strong predictor of unmutated Ig genes (Magnac C, 2003). Other studies have described a relationship between thymidine kinase level and LDT, lymphocyte count, and beta-2 microglobulin. This enzyme has also been shown to be a signiÞcant predictor of survival and response to treatment (Shanafelt

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T, 2003). Commercial assays are available for the measurement of this enzyme. Additional large-scale studies are required to determine its prognostic importance in multivariate analyses. CD38. CD38 is a cell-surface molecule expressed by mature B cells. Its expression on CLL cells has been shown to have prognostic signiÞcance in univariate and multivariate analyses, which have included clinical-stage, cytogenetic abnormalities and beta-2 microglobulin levels (Chevallier P, 2003; Del Poeta G, 2001; Ibrahim S, 2001). CD38 expression has been associated with unmutated immunoglobulin status, though results are discordant in approximately 30% of cases. Whether CD38 has prognostic signiÞcance in patients with known Ig mutation status is controversial (Hamblin TJ, 2002; Thunberg U, 2001). This marker may also vary during the course of disease. CD38 is reportedly a useful and easily measured prognostic marker, but it is limited by differences in expression over time for individual patients and is not a surrogate for Ig mutation status (Shanafelt T, 2003). ZAP70. ZAP70 is a signaling molecule involved in transducing intracellular signals from the T-cell receptor to the nucleus in T lymphocytes. This molecule was found to be aberrantly expressed in malignant CLL B cells that have unmutated Ig genes. The aberrant expression of ZAP70 in this subset of CLL cells was discovered by microarray gene chip expression proÞling and conÞrmed by protein analysis (Klein U, 2001; Rosenwald A, 2001). It was also found to be elevated in CLL cells that are CD38-positive (Durig J, 2003). A ßow cytometric assay that could identify ZAP70 expression is under investigation. Although promising, additional studies are needed to further deÞne the prognostic value of this marker in relation to other CLL markers. Cytogenetic Abnormalities. As many as 80% of CLL patients have identiÞable cytogenetic abnormalities, although these problems are more common in patients with advanced disease than in those with early-stage disease (Pangalis GA, 2002). No single speciÞc abnormality or gene has been consistently correlated with the pathogenesis of CLL, and any relationship between the genetic changes and the evolution of the disease has yet to be fully elucidated. The most common cytogenetic changes found in CLL are deletions in chromosomes 11, 13, and 17, and the presence of an additional chromosome 12. Deletions in chromosome 17 are associated with p53 oncogene inactivation (CaligarisCappio F, 1999). These chromosomal abnormalities have prognostic signiÞcance (Table 3) and affect treatment decisions in hospitals where the technology is available to perform the analysis. Etiology No etiologic factors have been clearly deÞned for CLL, although few studies have been performed. Most CLL cases occur sporadically, but approximately 1 in 20 patients has a familial form of the disease. The presence of familial cases clearly suggests that inherited genetic factors contribute to the development of

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TABLE 3. Common Cytogenetic Changes in Chronic Lymphocytic Leukemia and Their Significance

Marker (Mutation)

Frequency of Marker (%)

Deletions in chromosome 13

35–55

Additional chromosome 12

15–20

Deletions in chromosome 11

15–20

Defects in chromosome 17 affecting the p53 oncogene

15

Significance of Mutation and Additional Comments Associated with benign disease (stable or slowly progressing); patients with this marker survive as long as age-matched controls. Associated with atypical morphology, aggressive disease, and poor prognosis; patients tend to have unmutated immunoglobulin variable genes. Patients have more aggressive disease; patients who are carriers of the ATM genes found on chromosome 11 may be at greater risk of developing CLL. Associated with advanced disease, high proliferation rate, shortened survival, resistance to chemotherapy, and increased likelihood of Richter’s syndrome.

ATM = Ataxia-telangiectasia mutated.

the disease, although the speciÞc abnormal genes connected to CLL have not yet been identiÞed. The incidence of CLL in Þrst-degree relatives is three times greater than in the general population (3 in 10,000, compared with 1 in 10,000). CLL is less common in the Japanese and other Asian populations than in populations originating in the Western world, and the incidence does not increase in Asian expatriates. Environmental factors such as ionizing radiation, chemicals, and viruses do not seem to be associated with the pathogenesis of this disease. CURRENT THERAPIES Despite relatively good long-term survival rates, chronic lymphocytic leukemia (CLL) is considered incurable. For many years, Þrst-line therapy was dominated by the oral alkylating agent chlorambucil (GlaxoSmithKline’s Leukeran). The introduction of purine analogues as monotherapy or in combination with alkylating agents into current treatment strategies has vastly improved response rates, although no improvement in overall survival has yet been noted. Patients with CLL do not have a wide choice of therapy options. The overall population is elderly and has a low tolerance for toxic chemotherapy regimens. Once patients have failed both chlorambucil and ßudarabine (Schering AG and Berlex’s Fludara), the disease becomes extremely difÞcult to treat. The aim of current drug regimens is to obtain the highest possible rate and duration of remission while balancing the associated toxicity and infection rate in these patients. Table 4 describes the current regimens used to treat CLL.

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CHRONIC LYMPHOCYTIC LEUKEMIA (CLL)

TABLE 4. Current Regimens Used for Chronic Lymphocytic Leukemia Regimen Components Regimen

Agent

Chlorambu- Chlorambucil cil (single (GlaxoSmithagent) Kline’s Leukeran)

Availability

Dose

US, F, G, I, S, UK

Chlorambucil: 10 mg/m2 /d on days 1–7. Cycle repeated every 28 days.

Fludarabine Fludarabine US, F, G, I, (single (Berlex’s FluS, UK, J agent) dara, Schering’s Fludara/Fludar/ Bebeflur, generics)

Fludarabine (oral): 40 mg/m2 /d on days 1–5, (IV) 25 mg/m2 /d on days 1–5. Cycle repeated every 28 days.

Fludarabine/ Fludarabine US, F, G, I, cyclophos- (Berlex’s Fludara, S, UK, J phamide Schering’s (FC) Fludara/Fludar/ Bebeflur, generics)

Fludarabine (oral): 40 mg/m2 /d on days 1–3, (IV) 25 mg/m2 /d on days 1–3. Cycle repeated every 28 days.

Cyclophosphamide US, F, G, I, (Bristol-Myers S, UK, J Squibb’s Cytoxan/ Endoxan/ Endoxana, Baxter’s Endoxan/ Endoxana, Shionogi’s Endoxan, Pfizer’s Neosar/Cyclostin, generics) Fludarabine/ Fludarabine US, F, G, I, cyclophos- (Berlex’s Fludara, S, UK, J phamide/ Schering’s mitoxanFludara/Fludar/ trone Bebeflur, (FCM) generics)

Cyclophosphamide: 250 mg/m2 /d on days 1–3. Cycle repeated every 28 days.

Fludarabine (oral): 40 mg/m2 /d on days 1–3, (IV) 25 mg/m2 /d on days 1–3. Cycle repeated every 28 days.

Common Toxicities • Myelosuppression • Mild gastrointestinal disturbances • Increased risk of secondary malignancy • • • •

Myelosuppression Increased infection Neurotoxicity Malaise/fatigue

• Nausea/vomiting • Anorexia • Myelosuppression • Increased infection • Neurotoxicity (fludarabine) • Alopecia (more common with cyclophosphamide) • Malaise/fatigue • Nausea/vomiting • Anorexia • Hemorrhagic cystitis (cyclophosphamide) • Neutropenia

• Myelosuppression • Increased infection • Neurotoxicity (fludarabine) • Alopecia (more common with cyclophosphamide)

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TABLE 4. (continued) Regimen Components Regimen

CHOP

Agent

Availability

Cyclophosphamide US, F, G, I, (Bristol-Myers S, UK, J Squibb’s Cytoxan/ Endoxan/ Endoxana, Baxter’s Endoxan/ Endoxana, Shionogi’s Endoxan, Pfizer’s Cyclostin, generics) Mitoxantrone US, F, G, I, (Serono S, UK, J Lab/Wyeth/ Takeda’s Novantrone, Baxter’s Onkotrone, generics) Cyclophosphamide US, F, G, I, (Bristol-Myers S, UK, J Squibb’s Cytoxan/ Endoxan/ Endoxana, Baxter’s Endoxan/ Endoxana, Shionogi’s Endoxan, Pfizer’s Neosar/Cyclostin, generics) Doxorubicin US, F, G, I, (Pfizer’s S, UK, J Adriamycin/ Adriblastine, Bristol-Myers Squibb’s Rubex, Kyowa’s Adriacin, generics) Vincristine (Eli US, F, G, I, Lilly/EG S, UK, J Labo/LillyShionogi’s Oncovin, generics)

Dose Cyclophosphamide: 200 mg/m2 /d on days 1–3. Cycle repeated every 28 days.

Common Toxicities • • • •

Malaise/fatigue Nausea/vomiting Anorexia Hemorrhagic cystitis (cyclophosphamide

• Cardiac toxicity (mitoxantrone)

Mitoxantrone: 6 mg/m2 /d on day 1. Cycle repeated every 28 days.

Cyclophosphamide: 750 mg/m2 /d on day 1. Cycle repeated every 28 days.

• Myelosuppression • Increased infection • Alopecia • Malaise/fatigue • Nausea/vomiting • Anorexia • Hemorrhagic cystitis (cyclophosphamide)

Doxorubicin: 50 mg/m2 /d on day 1. Cycle repeated every 28 days.

Vincristine: 1.4 mg/m2 /d on day 1. Cycle repeated every 28 days.

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CHRONIC LYMPHOCYTIC LEUKEMIA (CLL)

TABLE 4. (continued) Regimen Components Regimen

Agent

Availability

Dose

Common Toxicities

Prednisonea

US, F, G, I, S Prednisone/ Prednisolone: 50 mg/m2 on days 1–5. Cycle repeated every 28 days. Prednisolone US, F, G, I, (generics) S, UK, J CVP Cyclophosphamide US, F, G, I, Cyclophosphamide: (Bristol-Myers S, UK, J 750 mg/m2 /d on day. Cycle Squibb’s repeated every Cytoxan/ 28 days. Endoxan/ Endoxana, Baxter’s Endoxan/ Endoxana, Shionogi’s Endoxan, Pfizer’s Neosar/Cyclostin, generics) Vincristine (Eli US, F, G, I, Vincristine: Lilly/EG S, UK, J 1 mg/m2 /d on day 1. Cycle Labo/Lillyrepeated every Shionogi’s 28 days. Oncovin, generics) US, F, G, I, S Prednisone/ Prednisonea (generics) Prednisolone: 50 mg/m2 on days 1–5. Cycle repeated every 28 days. Prednisolone US, F, G, I, (generics) S, UK, J Pentostatin/ Pentostatin US, F, G, I, Pentostatin: cyclophos- (SuperGen/ S, UK, J 4 mg/m2 /d on day 1. Cycle phamide Wyeth/ repeated every Pfizer’s Nipent, 28 days. Nihonkayaku’s Coforin) (generics)

• Myelosuppression • Increased infection • Alopecia • Malaise/fatigue • Nausea/vomiting • Anorexia • Hemorrhagic cystitis (cyclophosphamide)

• Myelosuppression (mild) • Increased infection • Malaise/fatigue • Fever • Nausea/vomiting • Anorexia • Hemorrhagic cystitis (cyclophosphamide)

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TABLE 4. (continued) Regimen Components Regimen

Cladribine (single agent)

Alemtuzumab (single agent)

Agent

Availability

Cyclophosphamide US, F, G, I, (Bristol-Myers S, UK, J Squibb’s Cytoxan/ Endoxan/ Endoxana, Baxter’s Endoxan/ Endoxana, Shionogi’s Endoxan, Pfizer’s Cyclostin, generics) Cladribine US, F, G, I, (Ortho-Biotech’s S, UK, J Leustatin, Janssen-Cilag’s Leustatin/ Leustatine/ Leustat, generics) Alemtuzumab (Berlex’s Campath, Schering’s Mab Campath)

US, F, G, I, S, UK

Dose

Common Toxicities

Cyclophosphamide: 600 mg/m2 /d on day 1. Cycle repeated every 28 days.

Cladribine: 0.14 mg/kg/d on days 1–5.

Alemtuzumab: escalating dose of 3, 10, 30 mg/d for week 1, followed by 30 mg/d for 3 days per week.

• Myelosuppression • Increased infection • Malaise/fatigue • Fever • Rash • Headache • Myelosuppression • Increased infection • Anemia • Fever • Nausea/vomiting • Rash • • • •

Fatigue Urticaria Rigors Transient Cytopenia

a. Prednisolone is used where prednisone is not available. IV = Intravenous; MAB = Monoclonal antibody. US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan.

Chlorambucil (Single Agent) Overview. For the past 40 years, Þrst-line treatment of CLL has been limited mainly to the alkylating agent chlorambucil (GlaxoSmithKline’s Leukeran). Although newer agents are being used with increasing frequency, chlorambucil still garners a signiÞcant patient share in the United States, France, Germany, Italy, Spain, and the United Kingdom. This popularity is the result of its preferable toxicity proÞle, which is advantageous in CLL—a disease with an elderly population that is less able to cope with high toxicities.

354

CHRONIC LYMPHOCYTIC LEUKEMIA (CLL)

Cl

COOH N

Cl FIGURE 2. Structure of chlorambucil.

Mechanism of Action. Chlorambucil (Figure 2) is an alkylating agent. Alkylation of DNA results in breaks in the DNA molecule as well as cross-linking of the twin strands, thus interfering with DNA replication and transcription of RNA and ultimately causing cell death. Clinical Performance. A variety of schedules are in use. Chlorambucil is an agent that can be administered with or without the corticosteroids prednisone or prednisolone (both generic). No evidence from randomized trials exists to prove that adding a corticosteroid enhances the efÞcacy of chlorambucil or other alkylating agents. However, physicians often include them in cases of bulky disease requiring a rapid response or when chlorambucil produces severe hematologic toxicity (Pangalis GA, 2002). First-line chlorambucil therapy (plus or minus prednisone) in untreated CLL patients produces complete response rates of 4–12% and overall response rates of approximately 40–60% (Rai K, 2000; Robak T, 2000). Recent trials have shown that Þrst-line ßudarabine therapy (discussed later) provides superior response rates compared with chlorambucil (Rai K, 2000); however, the two compounds’ long-term survival rates are not signiÞcantly different. Chlorambucil does have a less toxic side-effect proÞle and is therefore the preferred therapy in older patients (65 or older) and those with a poor performance status who are unable to tolerate the severe myelosuppression caused by ßudarabine. Another well-established beneÞt of chlorambucil is its oral formulation, although an oral formulation of ßudarabine has become available in some countries. Fludarabine (Single Agent) Overview. The clinical efÞcacy of purine analogues Þrst became apparent in the late 1980s (Keating MJ, 1989), and a progressive shift toward their use in Þrst-line therapy has occurred. Fludarabine (Schering AG/Berlex’s Fludara) is approved for the treatment of CLL in patients who have failed prior therapy, but physicians report this agent forms the basis of Þrst-line regimens in patients with a good performance status. Fludarabine is the leading cytotoxic agent throughout our study period in terms of market share. Mechanism of Action. Fludarabine (Figure 3) is a purine analogue and is metabolized rapidly to F-Ara-ATP, which restricts DNA synthesis by inhibition

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FIGURE 3. Structure of fludarabine.

of DNA polymerases and prevents elongation of DNA strands through direct incorporation into the DNA molecule. Clinical Performance. Three large-scale, randomized trials have investigated single-agent ßudarabine in previously untreated CLL patients. A three-arm study compared ßudarabine (25 mg/m2 on days 1–5, every 28 days), chlorambucil (40 mg/m2 every 28 days), and a ßudarabine (20 mg/m2 on days 1–5 every 28 days)/chlorambucil (20 mg/m2 every 28 days) combination in untreated patients with intermediate- and high-risk disease (Rai KR, 2000). The overall response rate was markedly improved in patients receiving ßudarabine compared with chlorambucil (63% versus 37%). Complete and partial responses were 20% and 43%, respectively, in the ßudarabine arm compared with 4% and 33%, respectively, in the chlorambucil arm. The median duration of response was 25 and 14 months, and median survival was 66 months and 56 months in patients receiving ßudarabine versus chlorambucil, respectively. However, a signiÞcantly higher incidence of grade 3 and 4 toxicities, speciÞcally neutropenia and infection rate, was observed in patients receiving ßudarabine compared with chlorambucil. The combination of ßudarabine and chlorambucil produced response rates similar to those of ßudarabine alone but with greater toxicity. Despite the improved response rates, at a median follow-up of Þve years, overall survival was not signiÞcantly different between the three arms of the study. The response rates for single-agent ßudarabine in untreated patients are in agreement with results gained from two other large-scale trials (Johnson S, 1996; Leporrier M, 1999). The dose and schedule of chlorambucil in the Rai study have been criticized as not optimal (Proctor SJ, 2001), yet a study in which high-dose chlorambucil (15 mg daily for up to six months) was administered produced a median survival of 68 months, which is not signiÞcantly different from the 56 months achieved in the three-arm study (Jaksic B, 1997; Rai K, 2001). A more recent study compared high-dose chlorambucil (10 mg/m2 daily for 18 weeks) with ßudarabine and found complete remissions (CRs) of 47% versus 33%, respectively, and

356

CHRONIC LYMPHOCYTIC LEUKEMIA (CLL)

partial remissions (PRs) of 41% versus 42% (Jaksic B, 2000). Hematological toxicity was signiÞcantly higher in the chlorambucil arm, and infections were more common in the ßudarabine arm. Survival data are not yet available. In addition to its efÞcacy as a Þrst-line agent, ßudarabine works in the secondline setting, following failure with alkylating and/or anthracycline agents. More than 15 studies including a total of 1,400 patients have published results showing a wide range of response rates due to the heterogeneity of patient characteristics. Overall response rates are approximately 40%, including 10% CR and 20% nodular PR (nPR) (Pangalis GA, 2002). The major limitation of ßudarabine is the high level of myelotoxicity, resulting in an increased infection rate. Elderly patients and those with a poor performance status are often unable to cope with these adverse side effects and are therefore limited to chlorambucil therapy. Fludarabine/Cyclophosphamide (FC) Overview. Several recent studies have investigated the combination of ßudarabine with cyclophosphamide (Bristol-Myers Squibb’s Cytoxan, Baxter’s Endoxan/Endoxana, PÞzer’s Neosar/Cyclostin, generics), and this regimen is used in both Þrst and subsequent lines of treatment. Mechanism of Action •

•

Fludarabine (Figure 3) is a purine analogue and is metabolized rapidly to FAra-ATP, which inhibits DNA synthesis by inhibition of DNA polymerases and prevents elongation of DNA strands through direct incorporation into the DNA molecule. Cyclophosphamide (Figure 4) is an alkylating agent. These agents alkylate DNA bases, thereby producing “cross-links” that covalently link the two DNA strands and prevent cell replication.

Clinical Performance. A trial published in 2003 investigated this combination in both treated and untreated patients of performance status 0, 1, or 2 (Schiavone EM, 2003). The untreated patient cohort contained 15 patients with

FIGURE 4. Structure of cyclophosphamide.

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Rai stage III-IV CLL or stage II with bulky disease. Within this group, 60% of patients achieved CR and 40% achieved PR status. Seventeen patients had received previous therapy, 6 of whom received ßudarabine or ßudarabine combinations. Response rates were lower in the previously treated group: 29% achieved CR and 59% achieved PR. The median time to progression (TTP) (median follow-up = 24 months) for the entire population was 25 months (median TTP in untreated patients had not yet been reached versus 18 months in treated patients). The median survival was 35 months (median survival in the untreated group had not yet been reached versus 20 months in treated patients). The program of six courses of ßudarabine/cyclophosphamide was administered in 75% of patients and reduced in 19% of patients because of severe myelosuppression and/or sepsis, which was fatal in one patient. Two patients discontinued therapy after two courses because of primary resistance; both patients had been heavily pretreated and had been refractory to previous chemotherapy. Toxicities included severe neutropenia in 31% of patients, which delayed subsequent courses of therapy and required the use of prophylactic granulocyte colony-stimulating factor (G-CSF). Red cell transfusions were required in 22% of cases because of decreased hemoglobin. No cases of grade 3/4 thrombocytopenia were observed. Infections, including seven cases of pneumonia and two of sepsis—one of which was fatal—occurred in 28% of patients. In another study, the efÞcacy of ßudarabine/cyclophosphamide in a population of CLL patients was accurately determined by analyzing response rates according to the patients’ treatment history (O’Brien SM, 2001). Patients were divided into four cohorts: • • • •

No prior treatment. Prior therapy with alkylating agents either alone or in combination. Prior therapy with alkylating agents and ßudarabine. Initial response to ßudarabine therapy followed by relapse. Prior therapy with alkylating agents and ßudarabine. Patients relapsed after or were refractory to alkylating agents and failed to achieve a PR with their last ßudarabine-based therapy.

Patients who had not received prior therapy (n = 34) had an overall response rate of 88%, which comprised 35% CR, 29% nPR, and 24% PR. Their median TTP and overall survival had not yet been reached after a follow-up of 41 months. Previously untreated patients receiving ßudarabine monotherapy reportedly achieved an overall response rate of 60–80% (Rai K, 2000; Keating MJ, 1998), similar to the data presented in this trial. The CR rate was also not signiÞcantly different from that reported for ßudarabine alone. However, these authors suggested that CRs achieved with ßudarabine/cyclophosphamide may be more durable because the percentage of patients with CD5+ B cells still present in their bone marrow at the time of CR was less than that found in patients receiving ßudarabine with or without prednisone (8% versus 33%).

358

CHRONIC LYMPHOCYTIC LEUKEMIA (CLL)

The combination of ßudarabine/cyclophosphamide was signiÞcantly more efÞcacious as salvage therapy for previously treated patients than was single-agent ßudarabine. Previous Phase II studies have reported response rates of 45–65% in patients receiving ßudarabine following failure of therapy with alkylating agents. In one such study (Pangalis G, 2002). In this study, patients (n = 20) achieved an overall response rate of 85% (15% CR, 25% nPR, 45% PR) and estimated median overall survival of 38 months. In the group of patients who had previously received both drug therapies, those who were ßudarabine-sensitive (n = 46) showed an overall response rate of 80% (12% CR, 17% nPR, 51% PR) and a median overall survival of 21 months. Fludarabine-resistant patients have an extremely poor prognosis and a median survival of less than one year, with most combination regimens providing less than or equal to 15% response. In this trial, patients achieved a response rate of 38% and a median overall survival of 12 months. However, the majority of these remissions were partial (3% CR, 13% nPR, 26% PR), and median TTP was less than a year. SigniÞcant toxicities were observed with this regimen, resulting in dose reductions of cyclophosphamide and 25% of patients unable to complete the planned number of six courses of therapy. Both alkylating agents and purine analogues are known to cause myelosuppression, which can result in a high infection rate. The dose of cyclophosphamide was reduced from 500 mg/m2 to 300 mg/m2 daily for three days because of the occurrence of grade 3/4 neutropenia occurring in 88% of patients and grade 3/4 thrombocytopenia in 30% of patients at the highest dose. Even at 300 mg/m2 of cyclophosphamide combined with ßudarabine, 75% of patients had grade 3 and 48% grade 4 neutropenia. Infections were common, and serious infections including bacteremia and/or pneumonia occurred in 25% of patients. Fludarabine/Cyclophosphamide/Mitoxantrone (FCM) Overview. The ßudarabine, cyclophosphamide, and mitoxantrone (Serono/ Wyeth/Takeda’s Novantrone, Baxter’s Onkotrone, generics) regimen is under investigation in clinical trials and is used as the standard of care in some hospitals in Germany and Spain. Mechanism of Action. •

•

•

Fludarabine (Figure 3) is a purine analogue and is metabolized rapidly to FAra-ATP, which inhibits DNA synthesis by inhibition of DNA polymerases and prevents elongation of DNA strands through direct incorporation into the DNA molecule. Cyclophosphamide (Figure 4) is an alkylating agent. These agents alkylate DNA bases, thereby producing “cross-links” that covalently link the two DNA strands and prevent cell replication. Mitoxantrone (Figure 5) is a synthetic antineoplastic anthracenedione. It is a DNA-reactive agent that intercalates with DNA by hydrogen bonding,

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H N OH

O

HN

OH

O

HN

OH

OH N H

FIGURE 5. Structure of mitoxantrone.

causing cross-links and strand breaks. It also reacts with RNA and is a potent inhibitor of topoisomerase II. Clinical Performance. In vitro studies have demonstrated ßudarabine’s synergistic effect with both cyclophosphamide and mitoxantrone (Bellosillo B, 1998; Bellosillo B, 1999). On the basis of these results, a clinical trial was designed to test the efÞcacy of these agents in previously treated CLL patients (Bosch F, 2002). Of the 60 patients treated with FCM, 34 had received one prior therapy, 18 had received two, and 8 had received three or more. Previous therapies included chlorambucil with or without prednisone (70%), cyclophosphamide, doxorubicin, vincristine, prednisone (CHOP) (32%), and ßudarabine monotherapy (8%). FCM was administered following relapse in 25 cases (42%) and following development of resistance to prior therapy in 35 (58%) patients. Thirty patients (50%) achieved a CR and 17 patients (28%) achieved a PR. Of the 30 patients in CR, 10 were found to be negative for minimal residual disease (MRD) by ßow cytometry and polymerase chain reaction (PCR; socalled molecular remission). The CR and PR rate was signiÞcantly higher among patients who had relapsed (32% and 40%, respectively) compared with those who were resistant to prior treatment (6% and 28%, respectively). None of the Þve patients previously treated with ßudarabine achieved a CR, although they did respond to FCM therapy. The median duration of response was 19 months in patients who achieved a CR, similar to that achieved with other ßudarabine combinations, and was not signiÞcantly different if patients were MRD-negative (17 versus 21 months) or between relapsed and resistant cases. Hematologic toxicities and infections were the most signiÞcant side effects, as reported in other ßudarabine-based regimens. Sixty-three percent of patients suffered from grade 3 or 4 neutropenia, and this toxicity was greater in patients who had prior chlorambucil treatment (81% versus 47%). Grade 3/4 thrombocytopenia and anemia were observed in 16% and 17% of cases, respectively, and grade 3/4 infections/fever occurred in 23% of patients. Although no large-scale trials describing the use of FCM in Þrst-line therapy of CLL have been performed, the regimen is being used in this setting in some European countries.

360

CHRONIC LYMPHOCYTIC LEUKEMIA (CLL)

Cyclophosphamide, Doxorubicin, Vincristine, Prednisone (CHOP) Overview. The CHOP regimen incorporates cyclophosphamide (Bristol-Myers Squibb’s Cytoxan, Baxter’s Endoxan/Endoxana, PÞzer’s Neosar/Cyclostin, generics), doxorubicin (PÞzer’s Adriamycin/Adriblastine, Bristol-Myers Squibb’s Rubex, Kyowa’s Adriacin, generics), vincristine (Eli Lilly’s Oncovin), and prednisone (generics). This combination is frequently used in non-Hodgkin’s lymphoma; its use is becoming less widespread in CLL. Mechanism of Action •

•

•

•

Cyclophosphamide (Figure 3) is an alkylating agent. These agents alkylate DNA bases, thereby producing “cross-links” that covalently link the two DNA strands and prevent cell replication. Doxorubicin (Figure 6) is an anthracycline. Anthracyclines interact with several different cellular targets, most importantly topoisomerase II. By inhibiting this DNA regulation enzyme, anthracyclines exert their cytotoxic effect. Another mechanism leading to cell death, known as DNA intercalation, involves insertion of the anthracycline molecule between base pairs. This phenomenon causes single- and double-stranded breaks in DNA that inhibit cell proliferation. The free radicals generated by the reductive metabolism of anthracyclines may also damage cellular structures. Vincristine (Figure 7) is a vinca alkaloid. Vinca alkaloids interact with tubulin and disrupt microtubular function in the mitotic spindle. This action leads to metaphase arrest, resulting in mitotic arrest and cell death. Prednisone (Figure 8) is a corticosteroid and reduces inßammatory responses and suppresses the immune system.

Clinical Performance. The dosage of cyclophosphamide and doxorubicin in CHOP regimens can vary (300–750 mg/m2 cyclophosphamide and 25–50 mg/m2

FIGURE 6. Structure of doxorubicin (R = OCH3 , R1 = OH, R2 = OH, R3 = H, R4 = OH).

CURRENT THERAPIES

N

OH CH3

NH H3CO H3CO

361

N

O

H N R

CH3 OAc O

H HO H3CO

FIGURE 7. Structure of vincristine (R = CHO).

CH2OH C O

H3C

O OH

H3C

O FIGURE 8. Structure of prednisone.

doxorubicin). When lower doses of these drugs are used, it may be referred to as mini-CHOP. A randomized clinical trial was designed to compare the anthracyclinecontaining regimens CHOP (incorporating the lower doses of cyclophosphamide and doxorubicin) and CAP (cyclophosphamide, doxorubicin, prednisone) with ßudarabine in 938 previously untreated Binet stage B and C CLL patients (Leporrier M, 2001). Patients were randomly assigned to receive six monthly courses of CHOP, CAP, or ßudarabine. Those patients who received ßudarabine or CAP and had stable or progressive disease after three cycles were switched to either CAP or ßudarabine, respectively. The CHOP regimen used in this study consisted of lower doxorubicin doses, so responses were assessed after six courses of treatment and patients with stable or progressive disease were switched to ßudarabine. The response rates obtained following six courses of therapy (or at switch during the six courses) showed that both ßudarabine and CHOP were clearly superior to CAP. CR rates of 29.6%, 15.2%, and 40.1% and PR rates of 41.9%, 43%, and 31% were achieved in patients receiving CHOP, CAP, and ßudarabine, respectively, regardless of stage. Accrual to the CAP arm was prematurely closed

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when results of the Þrst interim analysis showed signiÞcantly reduced response rates compared with CHOP and ßudarabine. The median follow-up in this study was 70 months and median survival was 67 months, 70 months, and 69 months in the CHOP, CAP, and ßudarabine arms, respectively, with Þve-year survival rates of 57.3%, 59.8%, and 58.4%, respectively. Although distinct differences in remission rates were observed, they did not translate into differences in survival. This effect was generated by the fact that patients who did not respond to their initial therapy were switched to another arm of the study, and the majority of patients received subsequent therapy over the course of their disease, thus obscuring the interpretation of survival data. Similar conclusions have been outlined in other large, prospective multicenter trials (Rai K, 2000). A higher frequency and severity of nausea, vomiting, and hair loss in patients receiving anthracycline-containing regimens were observed, whereas myelosuppression predominated in the ßudarabine group. However, the increased rate of infection associated with myelosuppression in ßudarabine-treated patients reported in previous trials was not observed in this study. The authors suggested that this absence may be explained by the fact that this patient group was previously untreated. Additional studies have shown that standard doses of CHOP did not result in any advantage in terms of response duration or survival when compared with chlorambucil plus prednisone (Hansen MM, 1991; Kimby E, 1991). A metaanalysis of 2,022 CLL patients in ten trials of combination chemotherapy (CHOP or COP [cyclophosphamide, oncovin, prednisone]) versus chlorambucil plus or minus corticosteroids was carried out to compare overall long-term survival rates (CLL Trialists’ Collaborative Group, 1999). No beneÞt was seen for combination chemotherapy over single-agent chlorambucil in terms of Þve-year survival rates. The combination of the toxicity associated with CHOP and the response rates gained with other agents have convinced physicians that CHOP should be reserved for those patients who fail chlorambucil or ßudarabine therapy. Cyclophosphamide, Vincristine, Prednisone (CVP) Overview. CVP incorporates cyclophosphamide (Bristol-Myers Squibb’s Cytoxan, Baxter’s Endoxan/Endoxana, PÞzer’s Neosar/Cyclostin, generics), vincristine (Eli Lilly’s Oncovin), and prednisone (generics). As with CHOP, this older regimen is losing favor to ßudarabine-based regimens. Mechanism of Action •

•

Cyclophosphamide (Figure 4) is an alkylating agent. These agents alkylate DNA bases, thereby producing “cross-links” that covalently link the two DNA strands and prevent cell replication Vincristine (Figure 7) is a vinca alkaloid. Vinca alkaloids interact with tubulin and disrupt microtubular function in the mitotic spindle. This action leads to metaphase arrest, resulting in mitotic arrest and cell death.

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Prednisone (Figure 8) is a corticosteroid and reduces inßammatory responses and suppresses the immune system.

Clinical Performance. The majority of randomized trials comparing CVP (also known as COP) treatment with chlorambucil as Þrst-line therapy report equivalent response rates with the two treatment strategies. An Eastern Cooperative Oncology Group (ECOG) study randomized patients with advanced CLL to receive either chlorambucil and prednisone or CVP (Raphael B, 1991). A median follow-up of seven years determined that chlorambucil/prednisone and CVP invoked no signiÞcant differences in survival (4.8 versus 3.9 years), CR rate (25% versus 23%), and duration of response (2.0 versus 1.9 years). These results were also comparable to those obtained using the CHOP regimen. The French Cooperative Group on CLL obtained similar results in an analysis of 291 Binet stage B patients, randomized to receive either chlorambucil or CVP (French Cooperative Group on CLL, 1990). No signiÞcant differences were observed either in the three- and Þve-year survival rates (69% and 44% in the chlorambucil group and 73% and 43% in the CVP group, respectively) or in median survival times (58 months versus 57 months, respectively). A large-scale meta-analysis of 2,022 CLL patients in ten trials conÞrmed that the combination chemotherapy regimens CVP and CHOP offer no beneÞt in long-term survival when compared with chlorambucil and were signiÞcantly more toxic (CLL Trialists’ Collaborative Group, 1999). Pentostatin/Cyclophosphamide Overview. Pentostatin (SuperGen{/Wyeth/PÞzer}’s Nipent) was Þrst administered to patients with acute leukemia in the 1970s, but it was associated with severe extramedullary toxicity, and enthusiasm for its use declined. However, patients with hairy-cell leukemia and CLL have shown responses at lower doses, so pentostatin is being investigated again as part of a combination regimen. This agent will garner an increasing, but modest, off-label share of sales throughout the study period owing to current investigation into its use in CLL by investigators at the Mayo Clinic College of Medicine and the Memorial Sloan-Kettering Cancer Center. HO N NH

HO

N O

N

OH FIGURE 9. Structure of pentostatin.

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Mechanism of Action •

•

Pentostatin (Figure 9), a purine analogue, is a potent, irreversible inhibitor of adenosine deaminase (ADA) produced by Streptomyces antibioticus. ADA is a vital enzyme in the purine salvage pathway, and its inhibition results in the accumulation of adenosine and deoxyadenosine metabolites, which inhibit ribonucleotide reductase. The depletion of the nucleotide pool leads to the inhibition of DNA synthesis and subsequent toxicity to the cell. ADA is a ubiquitous enzyme but is found in higher concentrations in lymphoid tissue. Cyclophosphamide (Figure 4) is an alkylating agent. These agents alkylate DNA bases, thereby producing “cross-links” that covalently link the two DNA strands and prevent cell replication.

Clinical Performance Pentostatin is thought to be the least myelosuppressive of the purine analogues in use to treat CLL. Its usage, though, is not widespread and few trial data are available. A study published in 2003 describes the combination of pentostatin with cyclophosphamide in 21 patients with intermediate- or high-risk CLL and 2 patients with small-cell lymphoma who had received substantial prior therapy (median of three regimens) including ßudarabine (87% of patients), chlorambucil (78%), and rituximab (26%) (Weiss M, 2003). Responses were seen in 17 patients and included 4 CRs (17%), 1 nPR (4%), and 12 PRs (57%). When analyzing the responses in comparison with patients’ responses to previous therapies, where sufÞcient data were available, investigators in this trial determined that seven patients achieved their best response on this regimen. In addition, 13 patients who had previously failed ßudarabine-based treatment responded, including 1 CR. (This latter group of patients is particularly difÞcult to treat and is left with few therapy options.) The median response duration of all 17 responders was 7 months (13 months for those in CR and 6 months for those in PR). As expected, median survival was longer for responders versus nonresponders (17 versus 8.5 months). Mild, asymptomatic tumor lysis syndrome (the release of cellular breakdown products into the blood) was detected in 39% of patients, demonstrating the rapid cytotoxic effects of this combination, which resulted in a sharp decline in white blood cell count following the Þrst cycle of chemotherapy. Pentostatin’s different mechanism of action may be clinically relevant in ßudarabine-treated patients. The incidence of response was similar among both sensitive and refractory patients, although a higher rate of CR occurred in ßudarabine-sensitive patients (30% versus 8%, respectively). One of the major reasons for using pentostatin versus ßudarabine is the potential for reducing myelosuppression. Grade 3/4 thrombocytopenia, neutropenia, and serious infections were observed in 30%, 35%, and 9% of patients in this study. The rates of thrombocytopenia and neutropenia are higher and lower, respectively, than those observed in a study of ßudarabine/cyclophosphamide

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NH2 N HO

N

N N

Cl

O

OH FIGURE 10. Structure of cladribine.

(19% and 75%) (O’Brien SM, 2001), and the rate of infection is lower. However, it is difÞcult to compare these trials because of different variables, including the use of G-CSF support in the pentostatin/cyclophosphamide study. Ongoing studies are attempting to determine the efÞcacy of pentostatin combinations in CLL patients, particularly combination therapy with monoclonal antibodies. Cladribine (Single Agent) Overview. Cladribine (Ortho Biotech’s Leustatin, Janssen-Cilag’s Leustatin/ Leustatine/Leustat, generics; also known as chlorodeoxyadenosine and 2-CDA) (Figure 10) is primarily used to treat hairy-cell leukemia. Cladribine is used much less frequently than ßudarabine for CLL because many more, and larger, randomized trials have evaluated the latter agent. It has now been approved. Mechanism of Action. Cladribine is a purine analogue that closely resembles ßudarabine; it inhibits DNA synthesis by interfering with DNA polymerases, thereby preventing the elongation of DNA strands. Clinical Performance. A multicenter, randomized prospective study analyzed the efÞcacy and toxicity of cladribine plus prednisone versus chlorambucil plus prednisone in previously untreated patients with progressive or symptomatic CLL (Robak T, 2000). As with other study designs, patients were randomized to receive either therapy; evaluated after a certain number of courses (in this case, three); and switched to the alternate therapy arm if they had not responded. As mentioned previously, this method obscures the interpretation of long-term survival data. However, in the initial evaluation following three courses of therapy, 59/126 (47%) patients receiving cladribine plus prednisone achieved a CR and 50/126 (40%) achieved a PR, giving a total response rate of 87%. In the chlorambucil plus prednisone arm, 12/103 (12%) patients achieved a CRs and 46/103 (45%) of patients achieved a PR, giving an overall response rate of 57%. In addition, early relapses occurred more frequently in the chlorambucil arm than in the cladribine arm. Patients who did not respond to Þrst-line treatment with either regimen were switched to the alternate arm of the study. Second-line treatment with cladribine

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plus prednisone was found to be more effective (10/43 CR,19/43 PR) than chlorambucil plus prednisone (0/26 CR and 7/26PR). Response rates in the cladribine arm were comparable to responses obtained in patients receiving ßudarabine. Although both the overall response rates and progression-free survival improved in the cladribine arm, this improvement did not translate into a signiÞcant difference in terms of overall survival rates. This effect may be associated with the switching of patients from one arm to another early on in the study. Analysis of the drug-induced toxicity conÞrmed cladribine’s myelosuppressive effects, which resulted in a high incidence of neutropenia and infections compared with patients in the chlorambucil arm. Thrombocytopenia occurred with equal frequency in both arms. Several Phase II studies have examined the efÞcacy of cladribine in combination chemotherapy in regimens such as cladribine/cyclophosphamide and cladribine/cyclophosphamide/mitoxantrone. In untreated patients, the former regimen produced overall response rates of 88%, with a CR of 29% (Robak T, 2002). Large-scale, randomized trial data are lacking for this drug. Alemtuzumab (Single Agent) Overview. Alemtuzumab (Schering AG’s Mab Campath, Berlex Laboratories’ Campath) was launched for third-line therapy of CLL in the United States and Europe in 2001. Early pilot studies indicated that alemtuzumab could cause tumor regression in advanced non-Hodgkin’s lymphoma. However, subsequent studies showed that the therapeutic effect was conÞned mainly to tumor cells in the blood and bone marrow rather than in lymph nodes, a Þnding that paved the way for trials in CLL. Patients who are refractory to ßudarabine are left with few treatment options. These difÞcult-to-treat patients are therefore candidates for therapies such as monoclonal antibodies (MAbs), whose uptake is growing. Mechanism of Action. Alemtuzumab is a chimeric, humanized MAb directed against the cluster of differentiation (CD) molecule 52, a glycosylphosphatidylinositol-anchored glycoprotein expressed on all mature lymphocytes, monocytes, and spermatozoa but not on hematopoietic stem-cell progenitors. The physiological function of CD52 is unknown. The binding of alemtuzumab to cell-surface CD52 leads to complement-mediated lysis, antibody-dependent cellular cytotoxicity, and opsonization, resulting in cell death. The efÞcacy of alemtuzumab has been correlated with the density of CD52 on the target cell surface. Clinical Performance. An international study investigated the efÞcacy of alemtuzumab in 93 CLL patients who had failed at least one alkylating-based regimen and ßudarabine treatment. FDA approval was granted largely on the basis of this study’s initial results (Keating MJ, 1999). Longer-term follow-up reported that 33 out of 93 patients responded (33%), experiencing 2 CRs (2%) and 29 PRs (31%) (Keating MJ, 2002). The authors noted that this result significantly exceeded their target response rate of 20%, and responses were seen in

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all prognostic subsets: both those who had failed ßudarabine and those who had previously had a short response to this agent. However, patients with Rai stage IV and those who had at least one lymph node greater than 5 cm in diameter were less likely to respond. Median TTP among responders was 9.5 months; median survival was 16 months in all patients, which is longer than for historical controls. Infusion-related reactions were commonly reported adverse events and were mostly grade 1/2. They included rigors (90% in total, 14% grade 3); fever (85% in total, 17% grade 3, 3% grade 4); nausea (53% grade 1/2); vomiting (38% in total, 1% grade 3); and rash (33% grade 1/2). These events declined over the time of treatment. Most patients experienced transient cytopenias. Neutropenia was most common during weeks 5 and 6 (30% of patients) and thrombocytopenia occurred in the Þrst two weeks. These problems had resolved in the majority of patients by two-month follow-up. This study recorded a high rate of infection (51 patients, 55%), although 53% of patients had a prior history of infection and 33% had infection in the month before alemtuzumab therapy. Twenty-Þve of 51 patients had a grade 3/4 infection. Grade 3/4 sepsis occurred in ten patients and led to death in two of these cases. A total of nine deaths occurred during treatment or within 30 days of the last alemtuzumab dose, of which Þve were related to treatment, comparing favorably with the 22% death rate observed in ßudarabine trials. Although alemtuzumab is approved as third-line therapy in CLL, the literature reports that it has been investigated in previously untreated CLL patients (Lundin J, 2002; Osterborg A, 1996). Forty-one patients, the majority (90%) of whom were in Rai stage II-IV, were enrolled in a dose-escalation Phase II study of subcutaneous alemtuzumab (Lundin J, 2002). Of the 38 evaluable patients, 7 (19%) achieved a CR and 26 (68%) achieved a PR, giving an overall response rate of 87%. The response rate was almost as high in Rai stage III-IV patients as it was in stage I-II, possibly because this antibody is effective in eradicating bone marrow disease, thereby improving or normalizing peripheral blood counts. In addition, the response rates were equally high among older patients. Acute administration-related reactions such as rigor, nausea, hypotension, and bronchospasm were rare or absent. These reactions are commonly seen when alemtuzumab is administered intravenously, and the reason for the discrepancy is unclear. Hematologic toxicity included transient grade IV neutropenia in 21% of patients; in some patients, repeated or more prolonged episodes required the use of G-CSF so that further treatment was not delayed. Alemtuzumab is also under investigation in combination with chemotherapy and other monoclonal antibodies. Data presented at the American Society of Clinical Oncology (ASCO) conference in 2003 described the combination of ßudarabine and alemtuzumab in relapsed CLL patients who had received a median number of two prior courses of therapy (Elter T, 2003). Of 14 patients, 9 (64%) achieved a CR and 3 (21%) achieved a PR. Transient grade 3 and 4 hematologic toxicities were observed. Other investigations include using alemtuzumab as consolidation therapy following chemotherapy to further improve responses and eradicate minimal residual disease.

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Alemtuzumab’s major limitation is the high level of hematologic toxicity and subsequent infection it induces in these already fragile patients. Physicians are wary of these side effects, but in light of alternative options, alemtuzumab is a popular therapy choice in the third-line setting. Continued investigation is evaluating its worth in earlier treatment settings. Nonpharmacological Approaches Currently, the only potentially curative option for CLL is stem-cell transplantation (SCT), which is still a highly experimental approach. There are two main types of SCT: autologous, in which the patient’s own stem cells are harvested and then returned to his or her body, and allogeneic, in which a related or unrelated donor is the source of stem cells. Allogeneic SCT carries the risk of the patient developing graft-versus-host disease (GVHD), a condition in which the donated stem cells trigger an immune response against the patient. This complication can be fatal; indeed, allogeneic SCT carries a high risk of mortality. However, allogeneic SCT also provides a higher chance of cure, partly because of the graft-versus-leukemia (GvL) effect, whereby the donor’s stem cells trigger an immune response against the patient’s own leukemia cells. There is no risk of GVHD in autologous SCT, but neither does the beneÞcial GvL occur. Another disadvantage of autologous SCT is that harvested and donated stem cells may be contaminated with tumor cells, which are then returned to the patient. The relapse rate for patients treated with autologous SCT is high. Current research is investigating allogeneic SCT in a nonmyeloablative, rather than fully ablative, setting. Nonmyeloablative SCT uses less intensive conditioning regimens that rely on immunosuppression rather than cytotoxicity. In general, SCT is used in a minority of patients who are young and have poor prognostic factors or as a last-chance option for patients with advanced disease. EMERGING THERAPIES The emerging therapy market for B-cell chronic lymphocytic leukemia (CLL) is extremely sparse. Only two agents are in Phase III development, and of the agents in Phase II development, data are available on only a minority. Further, progress on agents such as Novartis’s protein kinase inhibitor midostaurin (PKC412) and Bioenvision/Ilex’s antimetabolite clofarabine (Clofarex) has not been published. Two immunotherapeutic approaches are in Phase II development for CLL: the University of Southampton in the United Kingdom is developing a DNA vaccine that produces anti-idiotype antibodies conjugated to tetanus toxin, and Immuno-Designed Molecules is developing IDM-4, a macrophage-activated killer-cell bispeciÞc antibody. No clinical data are available on either of these agents. The proteasome inhibitor bortezomib (Millennium’s Velcade) has been extremely successful in the treatment of multiple myeloma and is under investigation for non-Hodgkin’s lymphoma. However, clinical trial data demonstrating its effect in CLL have not been published. For now, the focus remains Þxed on

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monoclonal antibodies (MAbs) to improve both response and overall survival rates in CLL. Table 5 summarizes drug therapies in development for CLL. Monoclonal Antibodies Overview. The launch of alemtuzumab (Schering AG’s Mab Campath/Berlex’s Campath) for the treatment of CLL has clearly paved the way for the success of MAbs in this disease. Clinicians are very enthusiastic about this class of drug and interested in developing additional MAbs against other targets. Mechanism of Action. MAbs target cell surface proteins and can interfere with receptor/ligand interactions, thereby affecting downstream signaling and subsequent growth and proliferation. They can also alert the immune system to target the cell for death via cell antibody-dependent cellular cytotoxicity. Rituximab. Rituximab (Rituxan, MabThera) is under development by Biogen Idec and Genentech in collaboration with Roche, Chugai, and Zenyaku Kogyo. This antibody is in Phase III clinical trials in the United States and Europe. Rituximab is a mouse/human chimeric MAb directed against the cluster of differentiation (CD) 20 molecule. CD20 is a calcium channel that interacts with the B-cell immunoglobulin receptor complex (Bubien JK, 1993) and is expressed on both normal and malignant B cells, making it an ideal target for MAb therapy in B-cell disorders. After binding to CD20, rituximab is thought to deplete B cells in a number of ways, including antibody-dependent cellular cytotoxicity (ADCC), complement-dependent cytotoxicity (CDC), and alteration of calcium ßux and factors involved in apoptosis (Lin T, 2003). This antibody has been launched for the treatment of relapsed or refractory low-grade or follicular, CD20-positive B-cell non-Hodgkin’s lymphoma (B-NHL). Rituximab is under investigation in many CLL clinical trials both as a single agent and in combination with chemotherapy as well as in Þrst- and subsequentline settings. The role for rituximab as a single agent in CLL is controversial. Previous studies showed an overall response rate of only 11% (Winkler U, 1999) and 25% (Huhn D, 2001), comparing poorly with the rate of 40–60% reported for follicular NHL (McLaughlin P, 1998). However, a recent Nordic multicenter study yielded improved results. Twenty-four CLL patients of median age 57 (47–72) with active disease (3 Binet A, 7 Binet B, 14 Binet C) who had previously been heavily treated with a variety of chemotherapy regimens were given the standard dose of 375 mg/m2 rituximab once weekly for four doses. The primary objectives this study addressed were response rate, quality, and duration; secondary objectives were to analyze the feasibility and tolerability of rituximab therapy. Eight of 23 evaluable patients (35%) achieved a partial response (PR), with a median duration of 12.5 weeks. A drop of at least 50% in blood lymphocyte count occurred in 17/21 (81%) patients who had pretreatment lymphocytosis, and 10 patients achieved a normal blood lymphocyte count (< 3 × 109 L−1 ). Of the

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TABLE 5. Emerging Therapies in Development for Chronic Lymphocytic Leukemia Compound Monoclonal antibodies Rituximab (Rituxan, MabThera) United States Europe Japan Lumiliximab (IDEC-152) United States Europe Japan Antisense oligonucleotides Oblimersen (Genasense) United States Europe Japan Cell-cycle inhibitors Alvocidib (Flavopiridol) United States Europe Japan Immunostimulatory therapies Xcellerate United States Europe Japan ISF-154 United States

Development Phase

Marketing Company

III III —

Biogen Idec/Genentech Roche/Chugai/Zenyaku Kogyo —

II — —

Biogen Idec

III — —

Genta/Aventis — —

II III —

National Cancer Institute — —

I/II

Invitrogen (formerly with Xcyte Technology) — —

— — II

Europe — Japan — Immunotoxins Denileukin diftitox (Ontak) United States II Europe — Japan — Apoptosis inducers SDX-101 United States Ib/IIa Europe — Japan — Motexafin gadolinium (Xcytrin) United States II Europe — Japan — Selective apoptotic antineoplastic drugs OSI-461 United States IIa Europe — Japan —

— —

Tragen/University of California at San Diego — —

Ligand Pharmaceuticals — —

Salmedix — — Pharmacyclics — —

OSI Pharmaceuticals — —

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15 patients who did not achieve a PR with rituximab, 9 had at least a 50% drop in lymphocyte count and 3 achieved a normal count. Seventy-Þve percent of patients experienced rituximab-related side effects, half of which were related to the Þrst infusion only. The most common toxicities were World Health Organization (WHO) grade 1/2 chills and grade 2 fever. In previous studies, the severe infusion-related toxicities reported had been speciÞcally linked to a high tumor burden (Huhn D, 2001; Winkler U, 1999). Mainly mild/moderate side effects were observed in this study, even in patients with extremely high lymphocytosis (223 × 109 L−1 ). This study demonstrates that single-agent rituximab does have some activity in heavily pretreated CLL patients, although the response is minor and of short duration. The reasons rituximab is more effective in NHL than in CLL are unclear. Circulating soluble CD20 and a high tumor burden, both of which “mop up” rituximab, are potential mechanisms/states by which the antibody is rapidly cleared from the blood; this theory is supported by the observation of altered pharmacokinetics and increased response rates with higher doses of rituximab in CLL (Byrd JC, 2001). In addition, CLL cells have a much lower density of surface CD20 than do NHL cells, although no correlation between density and response to therapy has been found (Byrd JC, 2001). Rituximab has been used as a Þrst-line, single-agent therapy, and limited clinical data suggest it may be more effective than as second- or third-line therapy. In one trial, treatment-naive patients with stage II–IV small lymphocytic lymphoma (SLL) or CLL received 375 mg/m2 rituximab weekly for four doses (Hainsworth JD, 2003). Patients who achieved an objective response (PR or complete response [CR]) or stable disease at reevaluation after six weeks continued maintenance courses of rituximab using the standard four-week schedule every six months for a maximum of four courses. Twenty-two of forty-three patients (51%) had an objective response at week 6, and the remaining patients had stable disease. Twenty-eight patients (65%) went on to receive maintenance rituximab therapy. With a median follow-up of 24 months, the response rate was 58% (9% CR). Median progression-free survival (PFS) was 19 months with a one- and two-year actuarial PFS of 63% and 49%, respectively. Two patients had a reversible grade 3 infusion-related toxicity with the Þrst course of rituximab. The increase in overall response rate is encouraging, but the small CR indicates that single-agent rituximab will not result in long-term survival in CLL (Lin TS, 2003). Treatment for CLL is generally reserved for patients with symptoms of advanced disease, although rituximab therapy may be effective in early-stage disease for those at risk of progression. The overall response rate in 21 evaluable patients with Rai stage 0–II and beta-2 microglobulin levels ≥2 mg/dL was 90% (19% CR, 19% nodular PR [nPR], 48% PR) (Thomas DA, 2001). The clinical signiÞcance of these results is unclear because a longer follow-up is required to analyze time to progression and long-term survival (Lin TS, 2003). The dose and schedule of administration for single-agent rituximab therapy as both Þrst and subsequent lines of therapy are under investigation in doseescalation studies in an attempt to increase response rates. Researchers have

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reported using doses of up to 2,000 mg/m2 /week in four patients (Jain D, 2003). Such studies are ongoing to optimize clinical responses. The most active area of research involving rituximab is in combination with chemotherapy. A randomized Phase II study of ßudarabine in combination with concurrent rituximab versus sequential rituximab was conducted in 104 previously untreated CLL patients (Byrd JC, 2003[a]). The treatment schedule for sequential therapy involved patients receiving 25 mg/m2 ßudarabine for 5 days, repeated every 28 days for six cycles. Four weekly doses of 375 mg/m2 rituximab were administered to patients who achieved stable disease or better, following a two-month rest period and restaging. The concurrent schedule followed the same pattern as the sequential schedule, with the addition of rituximab to each ßudarabine cycle. It is important to note that patients receiving concurrent administration received 11 doses of rituximab (seven in combination with ßudarabine and four as consolidation after this therapy) compared with only 4 doses in the sequential arm. Concurrent administration of these two agents demonstrated superior response rates when compared with the sequential arm (47% CR versus 28% CR, 43% PR versus 49% PR, respectively). Neutropenia was more common in the concurrent arm, but infectious complications occurred at similar frequencies in both schedules. Additional data presented at the American Society of Hematology (ASH) meeting in 2003 determined that adding rituximab to ßudarabine did not signiÞcantly increase the risk of infection (Morrison VA, 2003). This encouraging study establishes that concurrent administration of rituximab and ßudarabine produces CR rates superior to those achieved with ßudarabine alone. To date, the impact of rituximab on improving progression-free survival and overall survival compared with ßudarabine monotherapy has not been analyzed in a randomized trial. A retrospective comparison with data from 179 patients enrolled in the North American Intergroup Study CALGB 9011 who received ßudarabine monotherapy showed that CR, PR, and two-year performance-free and overall survival rates were signiÞcantly superior in the ßudarabine/rituximab group (Byrd JC, 2003[b]). The triple-drug regimen ßudarabine/cyclophosphamide/rituximab (FCR) is also under intense investigation. In one study, 202 previously untreated CLL patients received FCR (25 mg/m2 /day F for three days; 250 mg/m2 /day C for three days; 375–500 mg/m2 R on day 1) (Keating MJ, 2003). Results showed 68% CR, 18% nPR, and 14% PR. The study also analyzed patients for the presence of minimal residual disease (MRD) and found that the FCR regimen produced a high level of MRD-negative complete remissions. A longer follow-up will determine whether MRD-negative CR is more durable than MRDpositive CR. At the 2003 ASH meeting, the results of a sequential FCR program also were presented (Lamanna N, 2003). Thirty treatment-naive CLL patients received six cycles of standard ßudarabine therapy, then 3 g/m2 cyclophosphamide every three weeks for three cycles, and Þnally standard rituximab therapy. CR and PR rates of 57% and 29% (10% nPR and 19% PR), respectively, were achieved.

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The FCR regimen has also succeeded in patients with relapsed or refractory CLL. In one trial, 179 patients who had already received between one and three courses of therapy were treated with FCR and achieved responses of 25% CR, 16% nPR, and 32% PR (Wierda W, 2003[a]). MRD (analyzed by polymerase chain reaction) was absent in 33% of CR patients. Therapy was well tolerated, and 62% of patients completed four or more cycles of this regimen. Forty-one percent of patients experienced fever and chills with the Þrst rituximab infusion, and a minority experienced hypotension, nausea, and dyspnea (6%, 9%, and 3%, respectively). Hematologic toxicities included neutropenia in 30% of cycles and thrombocytopenia in 12%. A comparative, retrospective analysis of patients treated with ßudarabine (plus or minus prednisone), ßudarabine/cyclophosphamide, or FCR demonstrated increased CR, overall response, and median survival in patients treated with FCR (Wierda W, 2003[b]). The purine analogue pentostatin (SuperGen Warner-Lambert’s Nipent) has shown signiÞcant activity and minimal toxicity when combined with cyclophosphamide in CLL patients (Weiss M, 2003). In one trial, rituximab was added to this combination (known as the PCR regimen) and administered to previously untreated CLL patients (Kay NE, 2003). Preliminary data on 15 patients presented at ASH 2003 revealed 40% CR, 13% complete clinical response, and 47% PR. Most toxicities were grade 1 or 2, although eight patients suffered grade 3 anemia and hypotension and one patient developed grade 4 sinus bradycardia. In another trial, 20 patients with relapsed or refractory disease were treated with the PCR regimen; the response rates were 20% CR, 10% nPR, and 50% PR (Weiss MA, 2003). Grade 3/4 neutropenia occurred in 45% of patients, grade 3/4 thrombocytopenia in 5%, and infections in 15%. Preliminary data suggest this regimen is well tolerated, but further analysis is needed to determine both response rates and toxicity proÞles compared with those associated with ßudarabine-containing regimens. The combination of rituximab and another MAb, alemtuzumab is under investigation for relapsed and refractory CLL. Nine patients underwent treatment with this combination, and preliminary data showed a 44% CR and 23% PR rate (Faderl S, 2003). Nonhematologic toxicities were grade 2 or less, and infection occurred in 44% of patients. Another study presented at ASH 2003 failed to show any complete or partial remissions in 11 patients with relapsed or refractory CLL who were treated with alemtuzumab and rituximab in combination (Nabhan C, 2003). Further investigation into the combination of these antibodies is needed to determine their potential efÞcacy. In an attempt to improve upon the success seen in FCR, the M.D. Anderson Cancer Center is pioneering a trial examining a regimen consisting of cyclophosphamide, ßudarabine, alemtuzumab, and rituximab (known as the CFAR regimen). Only two relapsed/refractory patients have completed all courses, and both achieved PRs (Wierda W, 2003[c]). Four patients on continuing therapy were evaluated after three courses, and responses included one CR, one nPR, and two PRs. Seven patients came off therapy because of treatment failure (n = 2),

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infection (n = 1), noncompliance (n = 1), or at their own request (n = 2); one death occurred as a result of disease-related liver failure. Early analysis indicates good response with substantial but expected toxicities. Rituximab enjoys extensive off-label usage in the United States, mainly in the Þrst- and second-line CLL settings in combination with chemotherapy. In Europe, however, the use of rituximab is restricted by a lack of reimbursement owing to its experimental status and high cost. Lumiliximab. The primatized anti-CD23 MAb lumiliximab (Biogen Idec’s IDEC-152) is under development for CLL, asthma, and rhinitis. The drug is now in Phase II clinical trials for CLL. CD23, the low-afÞnity IgE receptor, is expressed constitutively on CLL cells and is therefore a target of interest for MAb therapy in this disease. Lumiliximab has been developed as a macaque-human chimera; it has no mouse component to minimize immunogenicity. At the ASH conference in 2003, preliminary data were presented from an ongoing, Phase I, dose-escalating, multicenter study assessing the safety, efÞcacy, and pharmacokinetics of lumiliximab for relapsed or refractory CLL (Byrd JC, 2003[c]). Twenty-Þve patients were evaluable. All had progressive CLL after a median of three prior treatment regimens. Decreased absolute lymphocyte counts (ALC) were observed in 24 patients, and reduced lymphadenopathy was reported. The most common study-related adverse events included fatigue, nausea, headache, cough, and increased sweating; two patients suffered from dose-limiting toxicities of grade 4 neutropenia and grade 4 headache at the highest-dose tested (500 mg/m2 ). These early data suggest that lumiliximab therapy is safe and may have activity in heavily pretreated CLL patients. Accrual and response evaluations are ongoing. Preclinical data have demonstrated the induction of apoptosis in lumiliximabtreated CLL cells in vitro. Western blot analysis revealed alterations in the levels of proteins controlling apoptosis as well as activation of the caspase pathway (Pathan N, 2003). Additional in vitro data suggest synergy with other MAbs and chemotherapeutic drugs. Antisense Oligonucleotides Overview. No antisense oligonucleotides are on the market yet for cancer. Despite the compounds’ high speciÞcity and low toxicity, this drug class has been plagued with disappointing results, such as those reported for ISIS-3521 (Isis Pharmaceuticals/Eli Lilly’s AfÞnitak) in non-small-cell lung cancer. Their appeal is further limited by the fact that they require long periods of intravenous administration. Mechanism of Action. Antisense oligonucleotides are short sequences of single-stranded DNA that bind to a speciÞc region of target messenger RNA (mRNA). This binding, or hybridization, triggers enzymatic degradation of the mRNA, thereby blocking the translation of the mRNA and the generation of the corresponding protein.

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Oblimersen. Genta/Aventis’s oblimersen (Genasense) is in clinical trials for a variety of cancers, including malignant melanoma, multiple myeloma, prostate, lung, breast, colorectal, and leukemia/lymphoma. Enrollment is complete for Phase III trials in multiple myeloma, malignant melanoma, and CLL; the FDA awarded both fast-track and orphan drug designations for all three programs. Positive trial data from the malignant melanoma studies allowed Genta and Aventis to Þle the Þrst part of a rolling new drug application (NDA) in December 2003 with a request for priority review status, and preregistration approval took place in February 2004. However, in early May 2004, the FDA Advisory Committee voted not to recommend oblimersen for marketing approval for malignant melanoma. The committee stated that the data presented did not provide substantial evidence for effectiveness, as measured by response rate and progression-free survival, to outweigh the increased toxicity endured by the patients receiving this in vivo drug. Genta subsequently announced that it was withdrawing the NDA for oblimersen therapy in malignant melanoma. It plans to meet with the FDA to discuss key issues and the next steps in developing oblimersen for malignant melanoma. Oblimersen is an antisense oligonucleotide speciÞc for mRNA transcribed from the bcl-2 gene. Bcl-2 is involved in the prevention of apoptosis and is overexpressed in CLL. Preclinical data have demonstrated that oblimersen induces apoptosis in primary CLL cells in vitro and its activity is potentiated when combined with rituximab, alemtuzumab, or cytotoxic chemotherapy (Auer RL, 2001; Cotter FE, 2003). Ongoing trials are investigating these observations in CLL patients. In July 2003, a randomized Phase III study comparing ßudarabine and cyclophosphamide with or without oblimersen in patients with relapsed or refractory CLL completed enrollment of 200 patients at 60 centers in the United States, Canada, and Europe. Its objectives are to compare CR and nPR, overall response, response duration, survival, and time to progression. No data are yet available. Other ongoing clinical trials include a two-part Phase I/II study analyzing oblimersen as a monotherapy in CLL patients who have failed a median of three prior therapies. Twenty-six patients, with a median age of 61, in Rai stages II-IV and two patients with Richter’s transformation entered the trial (Rai K, 2002). Cell-Cycle Inhibitors Mechanism of Action. Progression through the cell cycle depends on numerous signaling pathways and checkpoints (e.g., the cyclin families of proteins, the cyclin-dependent kinases [CDKs]). Deregulation of these key cell-cycle checkpoints is observed in most cancer cells. Inhibition of these enzymes can result in cell-cycle arrest and, ultimately, cell death, usually by apoptosis. Targeting the machinery involved in the cell cycle aims to directly attack cells with a high turnover, such as cancer cells. Alvocidib. Aventis and the National Cancer Institute (NCI) were developing alvocidib (Flavopiridol) together until Aventis discontinued its involvement in

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February 2004. This synthetic ßavenoid inhibits CDKs and is the Þrst CDK inhibitor to enter trials; it is in Phase II trials for CLL. It has reached Phase III in France. Preclinical research has demonstrated alvocidib’s ability to enhance both the radio- and chemosensitivity of a variety of cell lines and animal tumors. Ongoing experiments are analyzing this mechanism of action as well as further delineating alvocidib’s activity when administered alone. In a syngeneic mouse model of lymphoma, alvocidib enhanced the tumor response to radiation (Mason KA, 2003), and in H460 human lung cancer cells, alvocidib enhanced the effects of docetaxel and radiation (Kim JC, 2003). Researchers examined the molecular pathway of alvocidib activity and found that this agent interacts synergistically with the tumor necrosis factor–related apoptosis-induced ligand (TRAIL). Alvocidib does so through a mechanism involving the downregulation of XIAP (X-linked inhibitor of apoptosis protein), resulting in apoptosis of human leukemia cells (Rosato RR, 2003). A Phase II study of alvocidib in patients with ßudarabine-refractory CLL is complete. This trial was an open-label, multicenter study aiming to accrue up to 37 patients. Those registered before September 2000 received intravenous (IV) alvocidib continuously on days 1–3, and treatment was repeated every 14 days for a total of 12 courses in the absence of disease progression or unacceptable toxicity. Patients who were registered after September 2000 received IV alvocidib over one hour daily on days 1–3. Treatment was repeated every three weeks for a total of eight courses. Patients were to be followed every three months for the Þrst year and then every six months for Þve years. The study aimed to assess toxicity, CR, PR, progression-free and overall survival, and the impact of alvocidib on normal T-cell subsets and immunoglobulin levels. No data from this trial have been published. A Phase I dose-escalation study is examining alvocidib combined with ßudarabine and rituximab in patients with various lymphoproliferative disorders, including CLL. No data have been published. Given the lack of data on the use of alvocidib for CLL, we examined results from Phase I and II studies of alvocidib as a single agent or in combination with chemotherapy in the treatment of other cancers. In a Phase II trial of single-agent alvocidib in patients with hormone-refractory metastatic prostate cancer, no objective responses were observed and only 11% of patients achieved stable disease as the best response (King DM, 2003). In addition, alvocidib administered at 40 or 50 mg/m2 /day for a total of 72 hours was associated with signiÞcant toxicities that resulted in a high withdrawal rate from the study. A single-agent Phase II study of alvocidib therapy involving 33 renal-cell carcinoma patients resulted in 1 CR, 2 PRs, and 13 stable disease, but the study’s response criteria were not met (Van Veldhuizen PJ, 2003). Alvocidib is being investigated in a variety of combination regimens with agents such as irinotecan, docetaxel, 5-FU and leucovorin, and paclitaxel and carboplatin. The majority of trials completed have set out to establish alvocidib’s safety and maximum tolerated dose.

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Immunostimulatory Therapies Overview. The subject of immune surveillance as a defense against cancer has been debated since the 1950s. Knowledge of the genetic basis of cancer and immune processes has expanded signiÞcantly since that time and allowed researchers to manipulate tumor-speciÞc characteristics as targets for the immune system. The idea of activating a speciÞc immune response against cancer is an attractive one, and scientiÞc research in this area is making its way into the clinic. Mechanism of Action. The immune system has developed a safety mechanism that allows it to mount appropriate responses to pathogens while maintaining tolerance to self-tissue. When this tolerance breaks down, autoimmunity occurs. The safety mechanism works by allowing only a speciÞc subset of cells known as antigen-presenting cells (APCs) to induce an immune response by delivering two signals. The primary signal comprises an antigen bound to a major histocompatibility (MHC) molecule, which engages the T-cell-receptor (TCR) complex on the surface of the responding T cell. The secondary, or costimulatory, signal results when molecules on the surface of the APC interact with their ligands on the T-cell surface. These initial steps must occur to drive an immune response; the delivery of one signal without the other can result in anergy (a state of nonresponsiveness). In cancer, it is thought that the immune system ignores malignant cells because one or both signals are missing. Xcellerate. Xcyte Therapies and the University of Chicago were developing Xcellerate, a cocktail of immunostimulatory antibody-coated beads, which are used to stimulate patients’ T cells ex vivo before being reinfused. Xcellerate is in Phase I/II studies for prostate cancer, multiple myeloma, and CLL in the United States. Xcyte technologies has eventually sold this technology to Invitrogen. Inadequate or impaired antigen-presentation and/or the delivery of costimulatory signals appear to play a role in the failure of the immune system to detect and eradicate cancer cells. This scenario is particularly true in CLL, where the number of circulating T cells is reduced and those that do exist can be anergic (nonresponsive) due to the tumor burden. Xcellerate aims to overcome these problems of immune inaction by delivering such signals ex vivo. Blood is collected from the patients and sent to Xcyte Therapies’ cell manufacturing facility, where Xcellerate-activated T cells are generated by stimulation with CD3 (part of the TCR complex) and CD28 (costimulatory signal) antibody-coated beads. The product is then returned and administered in an outpatient setting. Three clinical trials in the United States are investigating Xcellerate. A Phase I trial in hormone-refractory prostate cancer patients has been completed, and Phase I/II trials in multiple myeloma and CLL are ongoing. In 2003, at ASH, researchers presented data from the Phase I/II study of Xcellerated T cells in CLL patients (Kipps TJ, 2003). Eleven patients received varying doses of activated T cells. There were no grade 3 or 4 infusion toxicities. Lymph node and spleen size fell by more than 50% in 10/11 patients, although

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this decline was not associated with a corresponding fall in circulating lymphocyte count. Further follow-up is needed to determine the clinical signiÞcance of these data. ISF-154. ISF-154 is a personalized, cellular therapy being developed by the University of California at San Diego (UCSD) and Tragen for B-cell malignancies. It is in Phase II trials for CLL in the United States. B cells are one of the three types of professional APCs, but CLL B cells are known to be poor at presenting antigen because of their lack of surface costimulatory molecules. In an attempt to overcome this defect and potentially induce an immune response against the malignant cells themselves, ISF-154 therapy involves collecting CLL cells from the patient and transducing them with CD40L, a critical molecule for T-cell activation, using a recombinant adenovirus vector. These transduced cells are then readministered to the patient. Phase II trial data were presented at ASH in 2003. In this multidose trial, seven patients received Þve courses of 3 − 6 × 108 ISF-154 transduced cells given at two-week intervals, and trial endpoints aimed to evaluate safety and relative tumor load reduction (Saville W, 2003). All patients had intermediateor high-risk disease classiÞed by the modiÞed Rai system. Six of seven patients received all Þve courses of therapy; one patient withdrew after suffering grade II adverse events consisting primarily of ßu-like symptoms after the second course of therapy. All patients (7/7) responded to therapy, experiencing transient falls in B-cell counts and lymph node size reductions of more than 50%. Durable responses were achieved in 5/6 patients who received all Þve courses of therapy. Two of these Þve patients achieved a PR; a third patient’s disease declined progressively over six months as the circulating B-cell count fell by 60% following the last infusion. Immunotoxins Overview. Immunotoxins use MAb technology conjugated to natural toxins. MAbs are well established in cancer therapy. Immunotoxins have a lack of encouraging data in non-Hodgkin’s lymphoma, but their activity in blood-based tumors versus solid tumors may prove more efÞcacious. Mechanism of Action. Immunotoxins comprise peptides, usually an antibody or growth factor, which are linked to toxins such as diphtheria toxin, Pseudomonas exotoxin, or ricin. This “magic bullet” mechanism is designed to target the toxic moiety to a speciÞc cell by binding the growth factor/antibody portion of the immunotoxin to its cell surface receptor. Upon internalization, the toxin is cleaved into an active form and causes cell death. Denileukin Diftitox. Ligand Pharmaceuticals is developing denileukin diftitox (Ontak) for the treatment of CLL. Phase II trials are underway in the United States. This agent has already been launched in the United States and preregistered in Western Europe for the treatment of cutaneous T-cell lymphoma.

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Denileukin diftitox is an interleukin-2 (IL-2) diphtheria toxin fusion protein; it is designed to direct the cytocidal action of diphtheria toxin to cells that express the IL-2 receptor. The human IL-2 receptor exists in three forms: low (CD25), intermediate (CD122/CD132), and high (CD25/CD122/CD132) afÞnity, the last of which is expressed on activated T and B lymphocytes and activated macrophages. Researchers presented data from a Phase II trial in which heavily pretreated, ßudarabine-refractory CLL patients received denileukin diftitox at a dose of 18 µg/kg/day or 9 µg/kg/day (if older than 70) intravenously for 5 days every 21 days (Desai H, 2003; Frankel AE, 2003). The mean age of patients enrolled in this study was 61.8 years and included 14 males and 4 females with Rai stages I (n = 2), II (n = 6), and IV (n = 10) who had received a mean of 4.5 prior treatments. Eleven out of 12 patients who received three or more courses of denileukin diftitox showed a reduced CLL cell count, and 6 of the 11 showed a ≥95% reduction. Seven of 12 patients showed a reduction in all lymph node diameters, with 1 patient showing 60% and another 80% shrinkage. Pre- and post–bone marrow biopsies performed on 11 patients showed a reduction in CLL marrow index; 7/11 had>50% reduction, including ≥98% in 3 patients. In the study, denileukin diftitox produced 2/12 (17%) PRs and 7/12 (58%) minimal responses. Progression-free intervals ranged from 1 to more than 19 months. Toxicities were mild to moderate and included asymptomatic transient transaminasemia, fever, hypoalbuminemia, nausea, vomiting, elevated creatinine kinase, and vascular leak syndrome. The results suggest this agent has some biologic activity in this difÞcult-to-treat group of patients. Previous published data have reported that immunotoxins directed against CD25 are more effective in treating malignancies where the expression of surface CD25 is high, such as in hairy-cell leukemia (Robbins DH, 2000). Research has determined that CLL cells can be sensitized to CD25-directed immunotoxins by treating them with an agent that upregulates CD25 expression (Decker T, 2002). A follow-up study is investigating denileukin diftitox therapy in combination with bexarotene (Ligand’s Tagretin), a retinoid X receptor (RXR) selective agonist known to upregulate IL-2 receptor expression. Another study of denileukin diftitox therapy also presented data on a small number of heavily pretreated ßudarabine-refractory patients (Seymour JF, 2003). Their median age was 63, and patients had received three to eight prior therapies. Of the Þve evaluable patients, the median number of denileukin diftitox cycles received was three. Therapy was stopped after one cycle because of grade 4 hepatitis, diarrhea, and disseminated herpes simplex with fatal pneumonia and after two cycles because of grade 4 vascular leak with microangiopathy/cardiac tamponade or lack of efÞcacy. Other toxicities included grade 1 and 2 fatigue and myalgias; grade 4 anorexia; and grade 1, 2, and 4 raised liver function. Two of the Þve patients achieved a PR, both of which were ongoing at 12+ months, and two achieved minor responses (1 month and ongoing at 6+ months). CD25positive and -negative patients were treated, and PR was observed in both groups. Furthermore, therapy with denileukin diftitox resulted in reduced lymphocytosis

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(median pre = 55 and post = 3.7) and a more normalized range of hemoglobin, neutrophils, and platelets. Apoptosis Inducers Overview. Protein members of the apoptotic pathway are found to be dysregulated in many cancers. CLL cells, in particular, overexpress antiapoptotic proteins, thus avoiding susceptibility to cell death. CLL’s biology, therefore, makes it an attractive target for apoptosis-inducing agents. Mechanism of Action. Apoptosis is the mechanism by which cells undergo programmed cell death. It is a highly ordered and complex cascade with many regulatory checkpoints. Characteristic features include DNA condensation and membrane blebbing. SDX-101. Salmedix’s SDX-101 is a small-molecule, orally administered, proapoptotic agent that is the R-isomer of etodolac, a marketed anti-inßammatory drug. The marketed drug is a mixture of SDX-101 and its isomer. Salmedix has completed a Phase Ib/IIa nonrandomized, multicenter, dose-escalation trial of SDX-101 in CLL in the United States. Researchers presented data on 40 patients split into cohorts of 6–8, treated with doses of 600, 800, 1,000, 1,200, 1,800, or 2,400 mg twice daily for eight weeks, with a four-week post-treatment follow-up (Jenson M, 2003). The median age was 62.9, and 25% were Binet stage A–progressive, 63.5% Binet B, and 12.5% Binet C. Both untreated (55%) and previously treated (32% one prior therapy, 10% two, and 3% three) patients were enrolled, and the majority (82.5%) had a good performance status. Clinically relevant reductions (deÞned as ≥25%) in absolute lymphocyte count (ALC) were observed in 50%, 86%, 88%, and 63% of patients receiving 1,000, 1,200, 1,800, or 2,400 mg SDX-101, respectively. Reductions of 42–53% in ALC were achieved within four weeks. No consistent changes in lymphadenopathy or organomegaly were observed, so, based on the NCI’s Working Group criteria, no responses were noted. In addition, no clinically relevant changes in hemoglobin or platelet levels were observed. Adverse events included mild to moderate gastrointestinal toxicity (worse at doses of 1,800 and 2,400 mg), skin rash, and elevated liver enzymes (no dose relationship). Dose-limiting toxicities included deep vein thrombosis in 1 patient at 600 mg and elevated liver enzymes and painful adenopathy in 1 patient at 1,200 mg, all of which resolved upon withdrawal of the drug. Other dose-limiting toxicities included skin rash at 1,800 mg (requiring treatment with antihistamines) and diarrhea at 2,400 mg. No clinically signiÞcant myelosuppression, anemia, thrombocytopenia, or gastrointestinal bleeding occurred. Preclinical data presented at the American Association for Cancer Research (AACR) conference in July 2003 demonstrated that SDX-101 can induce apoptosis in primary CLL and multiple myeloma cells as well as in various lymphoma cell lines (Leoni LM, 2003). SDX-101 displayed an additive activity when combined with ßudarabine, chlorambucil, or rituximab. In CLL cells, SDX-101

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altered several proteins, including downregulation of the antiapoptosis protein Mcl-1. In addition, the concentration of SDX-101 required to kill cancer cells did not affect normal blood or bone marrow mononuclear cells. Motexafin Gadolinium. MotexaÞn gadolinium (Pharmacyclics’ Xcytrin) is the Þrst in a class of investigational drugs known as texaphyrins. The compound is under investigation for several cancers, including CLL, NHL, pancreatic cancer, non-small-cell lung cancer, and brain metastases. A Phase II clinical trial involving patients with refractory or relapsed CLL began in the United States in December 2003. Texaphyrins’ mechanism of action is still under investigation, but motexaÞn gadolinium is known to target cells that have increased rates of metabolism, such as cancer cells. Once inside the cell, motexaÞn gadolinium promotes oxidative stress by generating reactive oxygen species; tumor cells treated with this agent undergo apoptosis through the mitochondrial-mediated intrinsic pathway (Magda D, 2003). MotexaÞn gadolinium is also a radio- and chemosensitizing agent. Eleven CLL patients who had failed multiple prior treatment regimens received motexaÞn gadolinium daily for Þve days, every three weeks, for two cycles or until disease progression. Pharmacyclics presented preliminary data at the International Congress of Hematologic Malignancies in Canada in March 2004 (Pharmacyclics, press release, March 12, 2004). Researchers observed regression of involved lymph nodes and spleen and a decline in circulating tumor cells in several patients. MotexaÞn gadolinium was well tolerated. Based on these results, Pharmacyclics plans to expand its trials in CLL and other non-Hodgkin’s lymphomas to additional centers. Preclinical data presented at ASH 2003 demonstrated that treatment of lymphoma and myeloma cell lines with motexaÞn gadolinium resulted in growth inhibition, with a concomitant increase in apoptosis (Naumovski L, 2003). Lymphoma cell lines were also treated with motexaÞn gadolinium in combination with other therapeutic agents, including rituximab. Results showed at least additive cytotoxicity, indicating that motexaÞn gadolinium has the potential to be used as a single agent or in combination. Selective Apoptotic Antineoplastic Drugs Overview. OSI Pharmaceuticals is pioneering a new approach to the prevention and treatment of cancer with the development of agents termed selective apoptotic antineoplastic drugs (SAANDs). These drugs aim to selectively induce apoptosis in precancerous and cancerous cells. Mechanism of Action. SAANDs exert their apoptotic effect by inhibiting cyclic GMP phosphodiesterase (cGMP PDE), an enzyme know to be overexpressed in precancerous and cancerous cells. Inhibition results in an increase in cellular cGMP, a second messenger molecule that can induce apoptosis through a series of steps involving the activation of protein kinase G, reduction in β-catenin, and activation of caspases.

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OSI-461. OSI-461 is a more potent analogue of OSI Pharmaceuticals’ Þrstgeneration SAAND exisulind (Aptosyn), which is in Phase III trials for nonsmall-cell lung cancer. In 2001, the company initiated three Phase IIa trials of OSI-461 for CLL, renal cell carcinoma, and hormone-refractory prostate cancer. Fifteen previously untreated CLL patients received 400 mg OSI-461 daily for up to 30 weeks (Whitehead CM, 2003). Four of the 15 patients experienced a decline in absolute lymphocyte count (ALC) of more than 50%, and 3 achieved a 25–50% decline in ALC. Responses were observed two to six weeks after initiating treatment. Six patients maintained stable disease and two experienced disease progression. Six additional patients were enrolled in this study at the higher dosage of 800 mg daily. No further clinical data are available at this time. In February 2004, OSI Pharmaceuticals announced that it had expanded an ongoing Phase I dose-escalating and pharmacokinetic trial of OSI-461 in patients with advanced solid tumors. The company reported that preclinical data demonstrated that administering OSI-461 with food increased the drug exposure levels. The ongoing Phase I study was amended to allow OSI Pharmaceuticals to explore these data. The company stated that although initial antitumor activity was demonstrated in CLL, it does not believe OSI-461’s optimal dose has been determined. REFERENCES American Cancer Society (ACS). Cancer Facts and Figures, 2003 . Atlanta, GA: American Cancer Society; 2003. Auer RL, et al. Bcl-2 antisense (Genasense) induces apoptosis and potentiates activity of both cytotoxic chemotherapy and rituximab in primary CLL cells. Meeting of the American Society of Hematology. 2001. Abstract 3358. Bellosillo B, et al. Mitoxantrone, a topoisomerase II inhibitor, induces apoptosis of B-chronic lymphocytic leukemia cells. British Journal of Hematology. 1998;100(1): 142–146. Bellosillo B, et al. In vitro evaluation of ßudarabine in combination with cyclophosphamide and/or mitoxantrone in B-cell chronic lymphocytic leukemia. Blood. 1999; 94(8):2836–2843. Binet JL, et al. A new prognostic classiÞcation of chronic lymphocytic leukemia derived from a multivariate survival analysis. Cancer. 1981;48(1):198–206. Bosch F, et al. Fludarabine, cyclophosphamide, and mitoxantrone in the treatment of resistant or relapsed chronic lymphocytic leukemia. British Journal of Haematology. 2002;119:976–984. Bray F, et al. Estimates of cancer incidence and mortality in Europe in 1995. European Journal of Cancer. 2002;38:99–166. Bubien JK, et al. Transfection of the CD20 cell surface molecule into ectopic cell types generates a Ca2+ conductance found constitutively in B lymphocytes. Journal of Cell Biology. 1993;121(5):1121–1132. Byrd JC, et al. Rituximab dose-escalation trial in chronic lymphocytic leukemia. Journal of Clinical Oncology. 2001;19(8):2165–2170.

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Byrd JC, et al. Randomized Phase II study of ßudarabine with concurrent versus sequential treatment with rituximab in symptomatic, untreated patients with B-cell chronic lymphocytic leukemia: results from Cancer and Leukemia Group B 9712 (CALGB 9712). Blood. 2003;101(1):6–14. [a] Byrd JC, et al. The addition of rituximab to ßudarabine signiÞcantly improves progressionfree and overall survival in previously untreated chronic lymphocytic leukemia (CLL) patients. 2003. Proceedings of the American Society of Hematology. Abstract 245. [b] Byrd JC, et al. Interim results from a Phase I study of lumiliximab (IDEC-152, anti-CD23 antibody) therapy for relapsed or refractory CLL. Meeting of the American Society of Hematology. 2003. Abstract 248. [c] Caligaris-Cappio F, et al. New insights into the biology of B-chronic lymphocytic leukemia. Hematology. 1999. www.hematology.org/education/hema99/caligaris-cappio. pdf. Call TG, et al. Incidence of chronic lymphocytic leukemia in Olmsted County, Minnesota, 1935 through 1989, with emphasis on changes in initial stage at diagnosis. Mayo Clinic Proceedings. 1994;69:323–328. Catovsky D, et al. Prognostic factors in chronic lymphocytic leukemia: the importance of age, sex, and response to treatment in survival: a report from the MRC CLL 1 trial. MRC Working Party on Leukaemia in Adults. British Journal of Hematology. 1989;72(2):141–149. Cheson BD. The chronic lymphocytic leukemias. In: Devita VT. Cancer: Principles and Practice of Oncology. New York, NY: Lippincott, Williams & Wilkins; 2001. Chevallier P, et al. CD38 expression and secondary 17p deletion are important prognostic factors in chronic lymphocytic leukemia. British Journal of Hematology. 2002;116(1): 142–150. Chronic Lymphocytic Leukemia Trialists’ Collaborative Group. Chemotherapeutic options in CLL: a meta-analysis of the randomized trials. Journal of the National Cancer Institute. 1999;91(10):861–868. Cotter FE, et al. Oblimersen sodium (G-3139) sensitizes malignant B cells to alemtuzumab (Ab)-induced apoptosis. Proceedings of the American Society of Clinical Oncology. 2003. Abstract 910. Criel A, et al. Further characterization of morphologically deÞned typical and atypical CLL: a clinical, immunophenotypic, cytogenetic and prognostic study on 390 cases. British Journal of Hematology. 1997;97:383–391. Damle RN, et al. Ig V gene mutation status and CD38 expression as novel prognostic indicators in chronic lymphocytic leukemia. Blood. 1999;94(6):1840–1847. Decker T, et al. Sensitization of B-cell chronic lymphocytic leukemia cells to recombinant immunotoxin by immunostimulatory phosphorothioate oligodeoxynucleotides. Blood. 2002;99(4):1320–1326. Del Poeta G, et al. Clinical signiÞcance of CD38 expression in chronic lymphocytic leukemia. Blood. 2001;98(9):2633–2639. Demidem A, et al. Chimeric anti-CD20 (IDEC-C2B8) monoclonal antibody sensitizes a B-cell lymphoma cell line to cell killing by cytotoxic drugs. Cancer Biotherapy and Radiopharmaceuticals. 1997;12:177–186. Desai H, et al. A Phase II study of DAB389IL-2 in ßudarabine-refractory chronic lymphocytic leukemia patients. Proceedings of the American Society of Clinical Oncology. 2003. Abstract 2372.

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Dighiero G, et al. Is chronic lymphocytic leukemia one disease? Haematologica. 2002;87: 1233–1241. Durig J, et al. Expression of ribosomal and translation-associated genes is correlated with a favorable clinical course in chronic lymphocytic leukemia. Blood. 2003;101(7): 2748–2755. East Anglian Cancer Registry, 2004. Personal communication with the East Anglian Cancer Intelligence Unit. February 27, 2004. Elter T, et al. Development of new four-weekly schedule (FluCam) with concomitant application of Campath-1H and ßudarabine in patients with relapsed/refractory CLL. Proceedings of the American Society of Clinical Oncology. 2003. Abstract 2331. Est`eve J, et al. Relative survival and the estimation of net survival: elements for further discussion. Statistics in Medicine. 1990;9:529–538. Faderl S, et al. Continuous IV infusion (C.I.) followed by subcutaneous (S.C.) alemtuzumab plus rituximab is active in patients with relapsed/refractory chronic lymphoproliferative disorders (LPD). Proceedings of the American Society of Hematology. 2003. Abstract 2505. Ferlay J, et al. Globocan 2000.Cancer Incidence, Mortality, and Prevalence Worldwide. Lyon, France: International Agency for Research on Cancer. World Health Organization. IARC Press; 2001. Frankel AE, et al. Phase II study of DT fusion protein denileukin diftitox in patients with ßudarabine-refractory chronic lymphocytic leukemia. Clinical Cancer Research. 2003; 9:3555–3561. French Cooperative Group on Chronic Lymphocytic Leukemia. A randomized clinical trial of chlorambucil versus COP in stage B chronic lymphocytic leukemia. Blood. 1990;75:1422–1425. Gorgun G, et al. Immunomodulatory effects of RXR rexinoids: modulation of high-afÞnity IL-2R expression enhances susceptibility to denileukin diftitox. Blood. 2002;100(4): 1399–1403. Greene FL, et al., eds. AJCC Cancer Staging Manual . Boston, MA: Springer; 2002. Hainsworth JD, et al. Rituximab as Þrst-line and maintenance treatment for patients with chronic lymphocytic leukemia (CLL) or small lymphocytic lymphoma (SLL): A Minnie Pearl Cancer Research Network Phase II Trial. Proceedings of the American Association of Oncology. 2003. Abstract 2332. Hamblin TJ, et al. Unmutated Ig V(H) genes are associated with a more aggressive form of chronic lymphocytic leukemia. Blood. 1999;94(6):1848–1854. Hamblin TJ, et al. CD38 expression and immunoglobulin variable region mutations are independent prognostic variables in chronic lymphocytic leukemia, but CD38 expression may vary during the course of the disease. Blood. 2002;99(3):1023–1029. Hansen MM, et al. CHOP versus chlorambucil/prednisone in chronic lymphocytic leukemia. Leukemia Lymphoma. 1991;5(suppl 1):97–100. Harris NL, et al. A revised European-American classiÞcation of lymphoid neoplasms: a proposal from the International Lymphoma Study Group. Blood. 1994;84(5): 1361–1392. Huhn D, et al. Rituximab therapy of patients with B-cell chronic lymphocytic leukemia. Blood. 2001;98(5):1326–1331.

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Klein U, et al. Gene expression proÞling of B-cell chronic lymphocytic leukemia reveals a homogeneous phenotype related to memory B cells. Journal of Experimental Medicine. 2001;194(11):1625–1638. Lamanna N, et al. Sequential therapy with ßudarabine, high-dose cyclophosphamide, and rituximab induces a high incidence of complete response in patients with chronic lymphocytic leukemia. Proceedings of the American Society of Hematology. 2003. Abstract 1603. Leoni LM, et al. Activity of SDX-101 (R-etodolac), a pro-apoptotic compound for the treatment of chronic lymphocytic leukemia, multiple myeloma, and lymphoma. Proceedings of the American Association of Cancer Research. 2003;44. Abstract 1701. Leporrier M, et al. Randomized clinical trial comparing two anthracycline-containing regimens (CHOP and CAP) and ßudarabine (FDR) in advanced chronic lymphocytic leukemia (CLL). Blood. 1999;94(suppl 1):603a. Leporrier M, et al. Randomized comparison of ßudarabine, CAP, and CHOP in 938 previously untreated stage B and C chronic lymphocytic leukemia patients. Blood. 2001;98(8):2310–2325. Lin TS, et al. Rituximab in B-cell chronic lymphocytic leukemia. Seminar in Oncology. 2003;30(4):483–492. Lundin J, et al. Phase II trial of subcutaneous anti-CD52 monoclonal antibody alemtuzumab (Campath-1H) as Þrst-line treatment for patients with B-cell chronic lymphocytic leukemia (B-CLL). Blood. 2002;100(3):768–773. Magnac C, et al. Predictive value of serum thymidine kinase level for Ig-V mutational status in B-CLL. Leukemia. 2003;17(1):133–137. Mason KA, et al. Flavopiridol enhances tumor response to radiotherapy. Proceedings of the American Society of Clinical Oncology. 2003. Abstract 3478. Mauro FR, et al. Clinical characteristics and outcome of young chronic lymphocytic leukemia patients: a single institution study of 204 cases. Blood. 1999;94:448–454. McLaughlin P, et al. Rituximab chimeric anti-CD20 monoclonal antibody therapy for relapsed indolent lymphoma: half of patients respond to a four-dose treatment program. Journal of Clinical Oncology. 1998;16(8):2825–2833. Micheli A, et al. Cancer prevalence in European registry areas. Annals of Oncology. 2002; 13:840–865. Molica S, et al. Clinico-prognostic implications of simultaneous increased serum levels of soluble CD23 and beta-2-microglobulin in B-cell chronic lymphocytic leukemia. European Journal of Hematology. 1999;62(2):117–122. Montserrat E, et al. Lymphocyte doubling time in chronic lymphocytic leukemia: analysis of its prognostic signiÞcance. British Journal of Hematology. 1986;62(3):567–575. Montserrat E, et al. Presenting features and prognosis of chronic lymphocytic leukemia in younger adults. Blood. 1991;78:1545–1551. Montserrat E. Chronic lymphoid leukemias. In: Degos L, et al., eds. Textbook of Malignant Hematology. London: Martin Dunitz Ltd.; 1999. Morrison VA, et al. Adding rituximab to ßudarabine therapy for patients with untreated chronic lymphocytic leukemia (CLL) does not increase the risk of infection: Cancer and Leukemia Group B (CALGB) Study 9712. Proceedings of the American Society of Hematology. 2003. Abstract 1606.

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Chronic Myelogenous Leukemia

ETIOLOGY AND PATHOPHYSIOLOGY Overview Chronic myelogenous leukemia (CML; also called chronic myeloid leukemia and chronic granulocytic leukemia) is a slowly progressing blood and bone marrow disease in which too many bone marrow stem cells develop into granulocytes, a type of white blood cell. Over time, the granulocytes and blast cells (immature white blood cells) crowd out normal white blood cells, red blood cells, and platelets. Reduced numbers of normal white cells can increase the risk of infection; reduced platelet levels can lead to increased bruising or bleeding; and low numbers of red blood cells can cause weakness and fatigue. CML is characterized by a chronic phase that can last for months or years and is often asymptomatic. Eventually, CML progresses to an accelerated phase, during which the leukemia cells proliferate more quickly, then to a blastic phase. Once the person enters the blastic phase (also known as a “blast crisis”), median survival is usually less than one year. The disease is easily diagnosed because the leukemic cells of at least 95% of patients have a distinctive cytogenetic abnormality—the Philadelphia (Ph) chromosome. Pathophysiology Leukemias are cancers of the hematopoietic (blood-producing) system. The word leukemia derives from the Latin word for white blood and refers to the proliferation of white blood cells (leukocytes) in people with this disease. Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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FIGURE 1. The hematopoietic cascade: development of mature blood cells from pluripotent stem cells.

Mature blood cells (red cells, white cells, and platelets) are normally produced in the bone marrow from primitive hematopoietic stem cells. (Figure 1 shows the different lineages of blood cells and the stages involved in their maturation.) The blood cells mature and differentiate through a sequence of steps involving a series of complex—and incompletely understood—interactions with growth factors, cytokines, and other cells in the bone marrow. Once mature, the blood cells leave the bone marrow and enter the general circulation, where they have a limited life span. Leukemia occurs when a genetic mutation arises in a single cell that interferes with the normal maturation and differentiation of developing leukocytes. These

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changes produce “immortal” white blood cells, in which the mechanisms of programmed cell death (apoptosis) are inactivated. The white blood cells proliferate without limitation, eventually replacing normal bone marrow cells and entering the peripheral bloodstream. Unlike acute leukemias (which have a sudden onset, progress rapidly, and, if untreated, can be fatal in as little as two months from the onset of symptoms), chronic leukemias are indolent. They have an insidious onset, progress slowly, and can remain asymptomatic and require no treatment for months or years. Leukemia is a heterogeneous disease; in other words, the leukemic mutations may affect any stage of hematopoietic differentiation, and the type of leukemia can be characterized by the type of affected cell. If the mutations affect the maturation of lymphoid cells, lymphoid leukemias result (e.g., chronic lymphocytic leukemia, acute lymphocytic leukemia [ALL]); maturation arrest in myeloid differentiation results in myeloid leukemias (e.g., CML, acute myelogenous leukemia [AML]). In CML, maturation arrest in myeloid differentiation is caused by a genetic mutation that results in increasing numbers of circulating myeloid cells: neutrophils, basophils, and eosinophils, collectively known as granulocytes (which are themselves a type of leukocyte). The main function of granulocytes is to Þght infection by bacteria and fungi. Granulocytes also regulate allergic reactions. In the chronic phase of the disease, these cells are functionally mature and the initial clinical features of the disease are a result of high levels of these granulocytes. Among patients who are symptomatic at presentation, symptoms of CML include fatigue, weight loss, fever, night sweats, bruising, aches in bones and joints, and

TABLE 1. Laboratory Features Characteristic of Chronic Myelogenous Leukemia Diagnostic Modality Blood count

Blood smear

Features Characteristic of CML • Presence of leukocytosis (white blood count usually >25,000 per mm3 ). • Elevated basophils and granulocytes (particularly myelocytes). Promyelocytes and myeloblasts present in small numbers unless patient has presented during blast crisis. • Erythrocyte and platelet counts may also be increased. • Mild anemia is present in 50% of cases at presentation. • Platelet count is abnormally high in 30–50% of cases. • Morphology of white and red cells is normal. • Platelet morphology is usually normal but giant platelets may be present.

Bone marrow biopsy

• Marrow is hypercellular; an increase in the number of myeloid white blood cells occurs, particularly early myeloid forms.

Cytogenetic/molecular analysis

• Presence of Philadelphia chromosome/BCR-ABL fusion gene.

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swollen lymph nodes. Other patients are diagnosed following a routine blood test. Table 1 lists laboratory features characteristic of CML. In addition to increasing the number of circulating granulocytes, CML affects the maturation of bone marrow stem cells and causes an increase in blasts in the bloodstream and marrow. Blasts cannot carry out the functions of the mature granulocytes, resulting in anemia and increased risk of infection. CML is primarily a disease of adulthood. The median age at diagnosis is 55–60 years; less than 10% of cases occur in people under the age of 20 (Cortes J, 1996). The disease in children is similar in behavior to that in adults, but the outcome of treatment with progenitor stem-cell transplantation—the only potentially curative therapy for CML—is better in these younger individuals. Although the median survival is Þve to seven years (based on patients treated in the pre-imatinib era), the range is wide: some patients die within one year of diagnosis, while others live for more than 15 years. The ratio of CML cases by sex is 1.4 male cases to 1 female case, but the clinical course is similar in both sexes. Staging. The clinical progression of CML can be divided into the following three phases: • • •

Chronic (blasts represent 5% or less of cells in blood and bone marrow). Accelerated (blasts represent 6–30% of cells in blood and bone marrow). Blastic (blasts represent 30% or more of cells in blood and bone marrow).

The chronic phase is characterized by a slow accumulation of granulocytes. This accumulation can be easily controlled, but not cured, with medication (inducing hematologic remission, i.e., normalization of blood cell counts and spleen size). Most patients are diagnosed while still in this phase, which varies in duration depending on the maintenance therapy used. The chronic phase usually lasts three to Þve years before it evolves into accelerated or blastic-phase disease. In two-thirds of patients, the disease transforms gradually into the accelerated phase, at which point the leukemia is more difÞcult to manage and symptoms become more severe (the remaining one-third of patients progress straight to the blastic phase). During the accelerated phase, symptoms are caused by an increase in the number of granulocytes and blasts in the bloodstream. The survival of patients diagnosed in this phase averages 1–1.5 years. The accelerated phase is a poorly deÞned stage for which no universally accepted deÞnition exists. Table 2 shows three published classiÞcations of accelerated phase. The criteria proposed by researchers (Kantarjian HM, 1988) from the M.D. Anderson Cancer Center in Houston, Texas, have been used most frequently in the recent studies of imatinib (Novartis’s Gleevec/Glivec), the gold-standard therapy for CML. The criteria of the International Bone Marrow Transplant Registry (IBMTR) are most frequently used in the bone marrow transplantation literature. The most recent classiÞcation, proposed by the World Health Organization (WHO), is not often employed.

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After 3–18 months, the accelerated phase progresses to an acute blastic transformation, or “blast crisis.” The WHO recently proposed a change in the deÞnition of the blastic phase; the organization suggested using a blast level in the blood of 20% as the threshold for diagnosis of this stage of disease (Vardiman JW, 2002). However, most of the available literature uses the standard cutoff of a 30% blast level in the blood. Skin or tissue inÞltration by blasts also characterizes the blastic phase. Cytogenetic evidence of another Ph-positive clone or clonal evolution is usually present. At the blastic phase, the disease resembles acute leukemia. In the majority of patients, CML then transforms into a condition resembling AML; TABLE 2. Comparison of Three Classifications for Accelerated-Phase Chronic Myelogenous Leukemia

Characteristic Blasts (peripheral blood white cells or bone marrow cells) (%)a Blasts + promyelocytes (%)a Basophils (%)a Platelets (× 109/L)

WBC (× 109/L)

Cytogenetic evidence of clonal evolution Anemia

M.D. Anderson Cancer Center (MDACC)

International Bone Marrow Transplant Registry (IBMTR)

World Health Organization (WHO)

≥15

≥10

10–19

≥30

≥20

NA

≥20 <100

≥20 (+ eosinophils) Increasing and unresponsive to therapy or persistent decrease Difficult to control, or doubling in less than five days Present

NA

Present

NA

Splenomegaly

NA

Other

NA

Frequency of classifications’ use

High

Unresponsive to therapy Increasing

Chloromas, myelofibrosis

Used in some bone marrow transplantation literature and clinical trials

a As a percentage of cells in blood and bone marrow.

NA = Not applicable. WBC = White blood cell.

≥20 <100 unrelated to therapy or >1,000 unresponsive to therapy Increasing and unresponsive to therapy Present

NA Increasing and unresponsive to therapy Megakaryocyte proliferation, fibrosis, granulocytic dysplasia Rare

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in about 25% of patients, the leukemia takes on the appearance of ALL. Whether the blast cells seen in blastic-phase disease are of myeloid or lymphoid lineage is an important distinction because it predicts the likely response to treatment and prognosis; patients with lymphoid blastic-phase disease have a better prognosis. Blastic-phase disease is generally resistant to treatment and is fatal within three to six months. Cytogenetic Changes. BCR-ABL–Positive Patients. At least 95% of patients with CML have an acquired distinctive cytogenetic abnormality of chromosome 22, known as the Philadelphia (Ph) chromosome (Shteper JP, 2001). The shortened Ph chromosome is the result of a reciprocal translocation of chromosomes 9 and 22, a process that produces the BCR-ABL fusion gene. The normal BCR gene is located on chromosome 22q11.21. The gene encodes a phosphoprotein associated with serine/threonine kinase activity and shows autophosphorylation activity as well as transphosphorylation activity for several protein substrates (Maru Y, 1991). The c-ABL gene, mapping on chromosome 9q34, codes for a tyrosine kinase with nuclear localization. The DNA-binding activity of the ABL protein is regulated by CDC2-mediated phosphorylation, suggesting a cell-cycle function for ABL. The gene is also implicated in processes of cell differentiation, cell division, cell adhesion, and stress response. The tyrosine kinase activity of nuclear ABL is regulated in the cell cycle through a speciÞc interaction with the Rb protein (Welch PJ, 1993). ABL activity is negatively regulated by its SH3 domain, and deletion of the SH3 domain turns ABL into an oncogene (Shore SK, 1990). The translocation t(9;22)(q34;q11), which transposes the ABL gene from chromosome 9 to the center of the BCR gene on chromosome 22, results in a head-to-tail fusion of these two genes and the formation of the Ph chromosome (Bernards A, 1987). Although the position of the breakpoint, or rearrangement, on chromosome 9 varies considerably, the breakpoint on chromosome 22 is clustered in an area called the breakpoint cluster region. Investigators have demonstrated a correlation between the site of the breakpoint within the breakpoint cluster region on chromosome 22 and the length of time between presentation and onset of accelerated-phase CML. Researchers have discovered that patients with a 5’ breakpoint have a fourfold longer chronic phase than do those with a 3’ breakpoint (Mills KI, 1989). The BCR-ABL fusion gene (also known as the BCR-ABL rearrangement) expresses an enzyme, the BCR-ABL oncoprotein, which has abnormally high tyrosine kinase activity compared with normal cells. This enzyme phosphorylates a number of substrates, activating a series of signal transduction pathways and, in turn, alters stem-cell function. The result is the chronic phase of CML. The remaining 5% or less of patients with CML do not have a shortened chromosome 22 but possess the BCR-ABL rearrangement (Ph-negative, BCRABL–positive), which can be detected by sensitive molecular techniques such as reverse transcription polymerase chain reaction (RT-PCR) or ßuorescence in situ hybridization (FISH).

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A few patients do not exhibit the BCR-ABL rearrangement but, because they have clinical and morphological features suggestive of CML, are still given a diagnosis of CML (discussed further on). Although the BCR-ABL rearrangement is the hallmark of CML, it is not unique to the condition and is present in up to 10% of patients with ALL and AML who have no evidence of antecedent CML. Accordingly, the diagnosis of CML is not based simply on the presence of the chromosome abnormality but requires the presence of typical clinical and morphological features (Table 1). The presence of the BCR-ABL rearrangement is used to monitor the response of a patient to therapy; patients who have a complete cytogenetic response (i.e., no detectable BCR-ABL expression) following therapy have a good prognosis. Clonal Evolution. Scientists hypothesize that genetic changes secondary to the BCR-ABL rearrangement drive the transition from the chronic phase to the accelerated and blastic phases. Indeed, the frequency of additional cytogenetic changes (clonal evolution [CE]) increases with advancing stage, rising from 30% in the accelerated phase up to 80% in the blastic phase. Given that chronic-phase CML is a manageable condition carrying no mortality, whereas transformation heralds certain death, understanding the process behind transformation is a key area of genetic research. The genetic abnormalities that have been identiÞed to date are diverse, and it remains unclear whether common blockable pathways exist. On average, a patient in blastic crisis carries three cytogenetic abnormalities out of a wide range reported so far. Chromosomes 8, 17, 19, and 22 are most often involved in CE. This involvement is almost entirely the result of the frequent occurrence of trisomy 8 (trisomy means having three copies of a given chromosome in each somatic cell rather than the normal number of two), trisomy 19, an extra Ph chromosome, and isochromosome 17, which is closely associated with the myeloid type of blast crisis. At least one of these four major cytogenetic changes is found in 71% of Ph-positive cases of CML (Mitelman F, 1993). Minor cytogenetic changes have also been documented: trisomies of chromosomes 17 and 21; monosomies of 7, 17 and Y; and translocation t(3;21) (q26;q22) (Faderl S, 1999). At least one of these cytogenetic changes can be detected in 15% of all Ph-positive CML patients. BCR-ABL–Negative Patients. As previously mentioned, approximately 5% of patients with clinical and morphological features consistent with CML have no evidence of the BCR-ABL rearrangement. These patients are generally older and often have thrombocytopenia, lower blood cell counts, greater monocytosis, lower bone marrow myeloid:erythroid ratios, and less basophilia than BCRABL–positive patients do. In fact, patients in this group do not have CML, even though they may be given this diagnosis, but rather have one of a variety of hematologic disorders, such as chronic myelomonocytic leukemia. Although these patients have a low risk of blastic transformation, their median survival is shorter than that of BCR-ABL–positive CML patients. The natural history of BCR-ABL–negative patients is characterized by progressive leukemic burden, extramedullary leukemic inÞltrates, poor response to chemotherapy, and

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bone marrow failure. Because of the low incidence of BCR-ABL–negative disease, it does not warrant further discussion. Prognostic Factors. Patients being treated for CML can be monitored by hematologic, cytogenetic, and molecular tests such as FISH and RT-PCR. Based on these assays, therapy outcome may be deÞned in terms of hematologic remission (normalization of blood cell counts and spleen size), varying degrees of cytogenetic response (decline in percentage of Ph-positive cells), or a molecular remission (PCR-negative). Several studies have shown that PCR-negativity indicates complete eradication of the leukemic clone, and PCR-positivity is associated with relapse in T-depleted transplanted patients and those undergoing transplantation in the advanced phase. PCR-positive patients undergoing transplantation in the chronic phase or those receiving nonmanipulated bone marrow have a slightly higher risk of relapse than do PCR-negative patients. In these cases, competitive PCR has shown that a low number of BCR-ABL transcript molecules are associated with prolonged complete remission, and patients with an increasing number of transcript molecules are subject to earlier relapse (Gaigner A, 1988; Lin F, 1996). Table 3 presents clinical characteristics related to the host or the disease. It lists features related to therapy response that have been identiÞed as having prognostic signiÞcance. These factors have been incorporated into scoring systems that enable clinicians to classify patients into groups with different prognoses. One commonly used tool is the Sokal score, which uses a patient’s age, spleen size, platelet count, and percentage of peripheral blasts in a complicated formula to determine the patient’s risk category (Sokal JE, 1984). This classiÞcation divides patients into three groups with four-year survival rates of 62% (low risk), 43% (medium risk), and 33% (high risk). Because studies have found this model to be inaccurate for patients undergoing treatment with interferon-alpha (IFNα, the most common therapy used in the treatment of CML), an alternative scoring system, sometimes called the Hasford score, has been developed for these patients (Hasford J, 1998). In addition to the variables used in the Sokal TABLE 3. Poor Prognostic Factors for Chronic-Phase Chronic Myelogenous Leukemia Clinical Older age Male sex Enlarged liver Enlarged spleen Hematologic Elevated blast cells in peripheral blood or marrow Elevated basophils in peripheral blood or marrow Elevated platelet count or decreased platelet count Elevated promyelocytes plus myelocytes in peripheral blood Elevated eosinophils in peripheral blood Nucleated red blood cells in peripheral blood

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score, the Hasford score uses the level of eosinophils to calculate a patient’s risk category. Investigators have proposed new prognostic models based on imatinib-treated patients, but these models have yet to be validated in large trials. In one study of patients who received imatinib after IFN-α failure, four variables were associated with a lower probability of achieving a major cytogenetic response: hematologic resistance to IFN-α, time from diagnosis to treatment >12 months, bone marrow basophils ≥5%, and a percentage of Ph-chromosome metaphases >90% (Kantarjian HM, 2002[b]). This study identiÞed three risk groups based on the number of adverse features. The Þrst group (zero to one adverse features) has a 93% probability of achieving a major cytogenetic response, the second (two adverse features) a 53% probability, and the third (three to four adverse features) a 34% probability. Another model based on two adverse features has been proposed: (1) development of neutropenia < 1 × 109/L between days 45 and 90 of imatinib therapy, and (2) failure to achieve at least a major cytogenetic response at three months (Marin D, 2003). In this model, three groups were deÞned with progression-free survival probabilities at two years of 100% (no adverse features), 66% (one adverse feature), and 15% (two adverse features). However, when this model was tested, the two-year progression-free survival rates for the three groups were 98%, 80%, and 96%, respectively (Kantarjian H, 2004). Etiology CML is an acquired rather than an inherited condition; familial cases are rare and little evidence exists linking hereditary factors to CML (Lawler SD, 1977). The offspring of patients with CML do not have a higher incidence of CML than does the general population, and there is no correlation in monozygotic twins. In the great majority of patients, a causative factor cannot be identiÞed. Nevertheless, it is well known that ionizing radiation is a predisposing factor, as shown by studies of survivors of the Nagasaki and Hiroshima atomic bombs and of patients who have received radiotherapy for conditions such as cervical cancer (Heyssel R, 1960; Moloney WC, 1987). CURRENT THERAPIES Allogeneic stem-cell transplant (allo-SCT) is the only potentially curative therapy for chronic myelogenous leukemia (CML). This aggressive approach is appropriate only for young (less than 55 years of age), Þt patients with matched donors—a proÞle that accounts for less than one-third of the CML population. Since its launch in 2001, the tyrosine kinase inhibitor imatinib (Novartis’s Gleevec/Glivec), alone or in combination with other agents, has been regarded as the treatment of choice for patients not destined for immediate allo-SCT. Previously, the standard treatment for patients with newly diagnosed chronicphase CML who were ineligible for allo-SCT was interferon-alpha (IFN-α), either alone or in combination with low-dose cytarabine (LDAC) (PÞzer’s Cytosar-U, generics). IFN-α largely replaced hydroxyurea (Bristol-Myers Squibb’s Hydrea,

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generics) in the mid-1990s, when clinical trials demonstrated that it induces major cytogenetic responses in about one-third of patients and achieves an overall survival advantage of one to two years compared with hydroxyurea. When disease progresses to the accelerated and blastic phases, more aggressive chemotherapeutic regimens may be employed (e.g., an anthracycline and cytarabine, high-dose cytarabine alone). In select cases, autologous transplantation is attempted. However, once CML starts to progress, no treatment is particularly effective. Purely palliative interventions include radiotherapy, splenectomy, and leukapheresis (the mechanical removal of white blood cells). With the exception of the purely palliative interventions, which we do not discuss further in this report, the agents and procedures employed in the treatment of CML are described in the following sections. Table 4 lists brand names, marketing companies, dosage, and market availability of agents commonly used in the treatment of CML. Table 5 illustrates the achievements of current treatment modalities in various patient populations. Because of the long median survival of patients in chronic-phase CML, primary endpoints in studies involving these patients are often hematologic and cytogenetic response rates. These response rates act as an indicator of length of survival. As novel drugs are achieving complete hematologic and complete cytogenetic responses, molecular response is becoming an increasingly popular trial end point owing to the fact that a complete molecular response indicates disease eradication. The following types of responses are generally measured: TABLE 4. Current Therapies Used for Chronic Myelogenous Leukemia Agent

Company/Brand

Daily Dose

Availability

Protein tyrosine kinase inhibitors Imatinib Novartis’s Gleevec, Glivec

400 mg

US, F, G, I, S, UK, J

Interferons Interferon-alpha-2a Interferon-alpha-2b

Roche’s Roferon-A Schering-Plough’s Intron-A Sigma -Tau’s Humoferon, GlaxoSmithKline’s Wellferon, Sumitomo’s Sumiferon Janssen-Cilag’s Cilferon-a, Otsuka’s Oif

5 × 106 IU/m2 5 × 106 IU/m2

US, F, G, I, S, UK, J US, F, G, I, S, UK, J

5 × 106 IU/m2

I, S, J

5 × 106 IU/m2

I, J

Pfizer’s Cytosar-U, generics Bristol-Myers Squibb’s Hydrea, generics GlaxoSmithKline’s Myleran

20 mg/m2

US, F, G, I, S, UK, J

40–50 mg/kg

US, F, G, I, S, UK, J

0.1 mg/kgb

US, F, G, I, S, UK, J

Interferon-alpha-N1

Interferon-alpha Cytotoxic agents Cytarabine Hydroxyurea Busulfana

a Busulfan is now rarely used in the treatment of CML but is included in the table for historical reasons. b When used in myeloablative regimens prior to allogeneic progenitor stem-cell transplantation, a dose of

8–16 mg/kg is given over four days. US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan.

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CHRONIC MYELOGENOUS LEUKEMIA

TABLE 5. Achievements of Current Therapies Used for Chronic Myelogenous Leukemia, 2005: Benchmarks for Evaluation of Emerging Therapies

Setting Late chronicphase CML after failure with IFN-α therapy Early chronicphase CML Early chronicphase CML Chronic phaseCML Early chronicphase CML

Treatment

Four-Year Survival/ Ten-Year Survival (%)

CHR(%) CCR(%)

PFS After 18 Months/ 42 Months (%)

Median Survival (months)

Imatinib 400 mg dailya

—

95

41

89/—

—

Imatinib 400 mg dailyb IFN-α 5 MIU/m2 dailyd IFN-α 9 MIU/m2 dailyf IFN-α + LDAC (10 mg)h

—

97

76

97/90c

—

—

80

26e

—

89

—/47g

—

10

—

104g

70/—

92

50

—

—

a Kantarjian H, 2002. b O’Brien SG, 2003. c Guillhot F, 2004. A 42-month follow-up showed PFS of 90% for patients who had achieved CCR within

12 months of beginning therapy. For patients not achieving CCR, PFS was 75%. d Kantarjian HM, 1996. e Five-year survival for patients who achieved CCR was 90%. f The Italian Cooperative Study Group, 1998. g Data for Sokal’s low-risk patients. h Kantarjian HM, 1999.

CCR = Complete cytogenetic response. CHR = Complete hematologic response. CML = Chronic myelogenous leukemia. IFN-α = Interferon-alpha. LDAC = Low-dose cytarabine. PFS = Progression-free survival. Note: Full source citations appear in ‘‘References.’’

•

•

Hematologic response. A complete hematologic response (CHR) is the absence of disease-related symptoms and splenic enlargement, normalization of the white blood cell count (i.e., a count between 4,000 and 11,000 per mm3 ), and a normal differential white blood cell and platelet count. If only some of these criteria are met, the response may be classed as partial. Cytogenetic response. A complete cytogenetic response (CCR) is the absence of detectable Ph-positive cells (cells expressing the Philadelphia chromosome) in metaphase. If a percentage of Ph-positive cells is detectable, the response may be classed as a major cytogenetic response (MCR) (or partial cytogenetic response) (1–34% Ph-positive cells); minor cytogenetic response (35–94% Ph-positive cells); or no cytogenetic response (95–100% Ph-positive cells). Cytogenetic responses are clinically important because they are associated with better prognosis.

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Molecular response. A molecular response is the disappearance or reduction in quantities of BCR-ABL transcripts (i.e., amount of BCR-ABL oncoprotein). Monitoring the level of BCR-ABL is a way of predicting long-term patient outcomes. A thousandfold (≥3 log) reduction in levels of BCR-ABL is deÞned as a major molecular response (MMR). The authors of a study (involving more than 1,000 CML patients) estimated that 100% of CML patients who have achieved a CCR and who achieve a MMR at 12 months will remain progression-free after another year (Hughes TP, 2003). A complete molecular response (CMR) is when there is no evidence of BCR-ABL transcripts, indicating disease eradication. The levels of BCR-ABL transcripts are usually measured by a quantitative real-time polymerase chain reaction (PCR) assay.

Protein Tyrosine Kinase Inhibitors Overview. Protein tyrosine kinases (PTKs) are enzymes that catalyze the phosphorylation of tyrosine residues. These enzymes are involved in cellular signaling pathways and regulate key cell functions such as proliferation, differentiation, anti-apoptotic signaling, and neurite outgrowth. Unregulated activation of these enzymes, through mechanisms such as point mutations or overexpression, can lead to various forms of cancer as well as to benign proliferative conditions. More than 70% of the known oncogenes and proto-oncogenes involved in cancer encode PTKs. A number of PTK inhibitors have been developed and approved for cancer treatment. These include inhibitors of c-Abl (imatinib, for treatment of CML); HER2 (trastuzumab [Genentech/Roche’s Herceptin], for treatment of breast cancer); vascular endothelial growth factor (VEGF) receptor (bevacizumab [Genentech/Roche’s Avastin], for treatment of metastatic colorectal cancer); and the epidermal growth factor receptor (EGFR) geÞtinib (AstraZeneca’s Iressa, also known as cetuximab (ImClone/Merck & Co./BMS’s Erbitux), for treatment of lung and colorectal cancer, respectively. Mechanism of Action. The rationale for developing tyrosine kinase inhibitors for the treatment of cancer is based on the observation that tyrosine kinase enzymes are critical components of the cellular signaling apparatus and are regularly mutated or otherwise deregulated in human malignancies. Novel tyrosine kinase inhibitors are designed to exploit the molecular differences between tumor cells and normal tissues. In CML, affected cells have a consistent cytogenetic abnormality, the Philadelphia chromosome, which carries a BCR-ABL fusion gene encoding a tyrosine kinase oncoprotein. Imatinib mesylate is a speciÞc inhibitor of this oncoprotein. Imatinib. Imatinib mesylate (Novartis’s Gleevec/Glivec, formerly STI-571) (Figure 2) was Þrst launched in the United States in May 2001 for the treatment of blastic- and accelerated-phase CML and chronic-phase disease after failure of IFN-α therapy. Imatinib had previously been awarded fast-track status for the

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FIGURE 2. Structure of imatinib.

myeloid blastic phase indication of CML and granted orphan drug designation in the United States, European Union, and Japan. In December 2002, the FDA approved the product for Þrst-line therapy in all phases of CML, after data from the imatinib arm of the International Randomized Study of Interferon Versus ST-1571 (IRIS; discussed subsequently) showed high CCR rates and delay in disease progression, suggesting that imatinib improves long-term survival. The dose of 400 mg per day of imatinib administered orally, the same dose used in the IRIS trial, is considered standard therapy for patients with newly diagnosed CML in the chronic phase. In February 2002, the FDA also approved imatinib for the treatment of inoperable and metastatic malignant gastrointestinal stromal tumors (GISTs). The product is also being investigated for the potential treatment of other cancers that express tyrosine kinases, including acute lymphocytic leukemia (ALL) and certain solid tumors. Imatinib is a 2-phenylamino-pyrimidine derivative that speciÞcally inhibits the tyrosine kinase activity of the ABL proteins c-ABL and BCR-ABL. The BCRABL fusion gene present in CML encodes an oncoprotein, p210BCR-ABL, that has dysregulated tyrosine kinase activity that is central to the pathogenesis of CML (Buchdunger E, 1996). Imatinib competitively inhibits the interaction of adenosine triphosphate (ATP) with these oncoproteins, thereby lessening their ability to phosphorylate and activate downstream target proteins (Schindler T, 2000). The initial approval of imatinib was based on data from Phase II studies involving approximately 1,230 patients in 32 centers located in Þve countries. The trial endpoints included hematologic and cytogenetic response rates. In one study, a total of 532 patients with late chronic-phase CML in whom previous therapy with IFN-α had failed were treated with 400 mg of oral imatinib daily (Kantarjian H, 2002 [a]). Imatinib induced MCRs in 60% (69% of these patients displayed a CCR) and CHRs in 95% of the patients. The time to onset of an MCR ranged from 2.4 months to 19 months, and the median time to a CHR was 0.7 months. After a median follow-up of 18 months, CML had not progressed

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to the accelerated or blastic phases in an estimated 89% of patients, and 95% of the patients were still alive. Only 2% of patients discontinued treatment because of drug-related adverse events, and no treatment-related deaths occurred. Data from some ongoing Phase III IRIS trials demonstrated superior response rates in imatinib-treated patients compared with IFN-α (O’Brien SG, 2003). The IRIS study was the largest study of CML patients ever conducted, enrolling 1,106 patients (553 randomized to each treatment arm) with newly diagnosed Phpositive CML between June 2000 and January 2001 in 16 countries. The study compared imatinib at 400 mg per day with IFN-α plus subcutaneous low-dose cytarabine (LDAC) (IFN+LDAC) as Þrst-line treatments; patients were allowed to cross over to the other treatment arm if they experienced loss of response, lack of response, or intolerance to the treatment. Patients were evaluated for hematologic and cytogenetic responses, toxic effects, and rates of progression. After a median follow-up, the estimated rate of an MCR at 18 months was 87% in the imatinib group and 35% in the IFN+LDAC–treated group. The estimated rates of CCR were 76% and 14%, respectively. At 18 months, the estimated rate of freedom from progression to accelerated or blastic-phase CML was 97% in the imatinib group and 91% in the combination-therapy group. Imatinib was better tolerated than IFN+LDAC. It is worth noting that 89% of patients receiving IFN+LDAC had already switched to imatinib therapy after a median of only 8 months into the study. Therefore, the survival beneÞt with imatinib compared with IFN+LDAC has not yet become apparent with long-term follow-up because most patients treated with IFN+LDAC are beneÞting early on from the added sequential imatinib therapy. An additional follow-up to the IRIS trial at 42 months conÞrmed durable response with Þrst-line imatinib therapy while demonstrating the effect of cytogenetic response on long-term outcomes (Guilhot F, 2004). Of newly diagnosed patients treated with imatinib, 98% had achieved CHR, while 91% had achieved an MCR, and 84% had achieved a CCR. For patients who had achieved CCR and a thousandfold (3 log) or greater reduction in BCR-ABL transcript level (i.e., a molecular response) at 12 months, the probability of remaining progression-free was 98% at 42 months. This probability compared with 90% for patients with CCR and less than a thousandfold reduction in BCR-ABL transcript level, and 75% for patients who had not achieved CCR. Responses to imatinib were found to be durable at the 42-month follow-up; an estimated 91% of patients maintained CHR, 91% of patients maintained MCR, and 87% of patients maintained CCR. A follow-up study monitored the molecular response for a median of 42 months in all 28 patients enrolled in the IRIS trial in Australia and New Zealand who commenced imatinib as their Þrst-line therapy (Branford S, 2004). The study’s aim was to determine if the BCR-ABL levels continued to decrease after 24 months. A CCR (approximately equivalent to a greater than 2-log reduction of BCR-ABL) was achieved in 24 of the 28 patients. Of the four patients without a CCR, all had disease progression, and in one patient a BCR-ABL mutation was detected, followed by rapid progression to blastic-phase disease. The data demonstrate that, although the frequency of achieving an MMR increased

404

CHRONIC MYELOGENOUS LEUKEMIA

between 12 and 42 months, most of the improvement occurred between 12 and 24 months. Thirteen patients achieved an MMR by 12 months, and all 13 achieved a 4-log reduction (equivalent to undetectable levels of BCR-ABL transcripts) at 42 months. These results suggest that, in patients achieving an MMR by 12 months, leukemic cell mass is still decreasing after 3.5 years of imatinib therapy. Common side effects of imatinib treatment are superÞcial edema, nausea, and muscle cramps. Some patients may experience severe toxicity, leukopenia, thrombocytopenia, and anemia. The most common adverse events experienced in the IRIS trial were hematologic and hepatic toxicities and included severe (NCI grades 3/4) neutropenia (16.2%), anemia (4.0%), thrombocytopenia (9.3%), and elevated liver enzymes (5.4%). Other drug-related adverse events occurred in 15.8% of patients. Another study, conducted by researchers at the M.D. Anderson Cancer Center in Houston, Texas examined the optimal dose of imatinib therapy (Cortes J, 2004). In this trial, 222 previously untreated early chronic-phase CML patients were split into two groups. One group of patients was treated with the 400 mg daily dose of imatinib, while another group was treated with 800 mg daily. Patients in the higher-dose group had an estimated progression-free survival rate of 99% at 12 months compared with 92% in the standard dose group. Researchers concluded that the 800 mg daily imatinib dose resulted in higher rates of CCRs and MMRs. Extramedullary toxicity (toxicity outside the bone marrow) was similar in the two groups, but myelosuppression was more common with the higher dose. At 12 months, the median actual dose for the high-dose group was still 800 mg daily, with 36% of evaluable patients having required dose reduction, compared with 14% of those treated with the standard dose. Acquired resistance to imatinib among patients with chronic-phase disease appears to be rare and can often be overcome by increasing the dose. In a followup study, 261 patients with CML in chronic-phase post–IFN-α failure received an escalated daily dose of 600–800 mg of imatinib orally after demonstrating a poor response or relapse at the standard dose (400 mg daily) (Kantarjian HM, 2002[b]). Among patients treated for hematologic resistance or relapse, 65% achieved a complete or partial hematologic response. Among patients treated for cytogenetic resistance or relapse, 56% achieved a complete or major cytogenetic response. In contrast, 70% of patients in myeloid blast crisis exhibit resistance to imatinib. Furthermore, all patients in lymphoid blast crisis relapse within six months of responding to imatinib. This resistance appears to arise from a variety of mechanisms, including acquired mutations in the ABL kinase domain, BCRABL overexpression, P-glycoprotein overexpression reducing the cellular uptake of imatinib, selection of preexisting mutant cells, and possibly, excessive degradation of the BCR-ABL protein (Mahon FX, 2000; Gorre ME, 2001). Several studies have shown that imatinib is not as effective in the treatment of accelerated and blastic-phase CML as it is in the treatment of chronic-phase disease. A Phase II study investigated the hematologic and cytogenetic responses of 260 patients in myeloid blast crisis treated with 400–600 mg imatinib daily

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(Sawyers CL, 2002). Imatinib induced hematologic responses in 52% of patients and sustained hematologic responses lasting at least four weeks in 31% of patients, including CHRs in 8%. In patients with a sustained response, the estimated median response duration was 10 months. Imatinib induced MCRs in 16% of patients, and 7% of the responses were complete. Median survival time was 6.9 months. Drug-related adverse events led to discontinuation of therapy in 5% of patients, most often because of cytopenia, skin disorders, or gastrointestinal reactions. Another Phase II study involving 235 patients showed that imatinib 400–600 mg daily induced hematologic and cytogenetic responses in accelerated-phase CML. Imatinib induced a hematologic response in 82% of patients and sustained hematologic responses lasting at least four weeks in 69%, and complete responses in 34%. The rate of MCR was 24%; complete responses were achieved by 17%. Estimated 12-month progression-free and overall survival rates were 59% and 74%, respectively. In comparison with 400 mg, imatinib doses of 600 mg led to more cytogenetic responses (28% compared with 16%), longer duration of response (79% compared with 57% at 12 months), time to disease progression (67% compared with 44% at 12 months), and overall survival (78% compared with 65% at 12 months) with no clinically relevant increase in toxicity. Several groups have investigated the combination of imatinib plus LDAC using the hypothesis that resistance to imatinib would be less frequent. The CML French Group performed a Phase II trial to determine the safety and tolerability of the combination in 30 previously untreated patients in chronic-phase CML (Gardembas M, 2003). Treatment was administered in 28-day cycles. Patients were treated continuously with imatinib at a dose of 400 mg daily. LDAC was given on days 14 to 28 of each cycle at an initial dose of 20 mg/m2 /day via subcutaneous injection. Adverse events were frequently observed: grade 3 or 4 hematologic toxicities and nonhematologic toxicities occurred in 53% and 23% of patients, respectively. At 6 months, 100% of patients achieved a CHR, and the cumulative incidence of CCR at 12 months was 83%. The researchers concluded that the combination was safe and promising, given the rates of response. The STI-571 Prospective International Randomized Trial (SPIRIT) is a Phase III study underway to compare imatinib monotherapy, imatinib plus cytarabine, and imatinib plus IFN-α as Þrst-line treatment in randomized, newly diagnosed CML patients. Interferons Overview. The body’s immune system normally rids itself of aberrant cells before they have a chance to multiply, colonize, or metastasize. Immunotherapy takes advantage of these natural host defense mechanisms through several classes of agents: monoclonal antibodies that target aberrant proteins on cancer cells; cytokines that increase or activate host immune cells, such as T cells, macrophages, and natural killer (NK) cells (large granular lymphocytes that bind to and kill cells by releasing cytotoxins); or vaccines that activate speciÞc host T-cell or antibody responses that attack the cancer cells.

406

CHRONIC MYELOGENOUS LEUKEMIA

The only immunotherapy agent in current use for CML is the cytokine interferon-alpha (IFN-α). Cytokines are low-molecular-weight secreted proteins produced by many different cell types. These proteins are involved primarily in regulating the immune response and are necessary for the biochemical communication involved in wound healing, immunosurveillance, and inßammatory responses. The IFNs are a family of naturally occurring cytokines produced in response to viral infections, antigens, mitogens, and other cytokines. Three types of interferons have been identiÞed: interferon-alpha (produced by leukocytes and macrophages), interferon-beta (produced by Þbroblasts), and interferon-gamma (produced by T lymphocytes). Mechanism of Action. The direct antitumor effect of IFNs derives from their ability to inhibit cell-cycle progression, induce a reduction in tumor-cell protein synthesis, and inhibit late progenitor colonies. IFNs can also produce indirect antitumor effects by modulating immunomodulatory and anti-angiogenic responses (Baron S, 1991). Their immunomodulatory role involves inducing the expression of major histocompatibility antigens and modulating the expression and function of T cells, monocytes, and natural killer cells. Both the direct and indirect effects of IFNs result from induction of a subset of genes called the IFN-stimulated genes (ISGs) (Chawla-Sarkar M, 2003). Interferon-Alpha. IFN-α is used to treat approximately 30 different diseases, both alone and in combination with chemotherapeutic and biological agents. Several types of IFN-α are available (Table 4) although not all are licensed in all countries for the treatment of CML. IFN-α was considered standard Þrstline treatment until December 2002, when the FDA approved imatinib for the Þrst-line treatment of CML. IFN-α may exert a direct antiproliferative effect. Alternatively, it may exert an indirect effect on the immune system through nonspeciÞc enhancement of antileukemic cell-mediated response. IFN-α may increase human leukocyte antigen (HLA) molecule expression on Ph-positive cells; this increased expression enables more-efÞcient recognition of the HLA-linked leukemic peptide by antigen presenting cells (APCs) and T-lymphocytes (Ten Bosch GJA, 1999). The binding of IFN-α to its membrane receptor activates a number of signaling pathways that eventually modulate the transcription of several genes (Platanias LC, 1999). Many of these pathways are the same as those that are constitutively activated by the leukemia-speciÞc BCR-ABL tyrosine-kinase oncoprotein. An important difference between the effects of IFN-α and BCR-ABL is that ICSBP (interferon consensus sequence binding protein) is downregulated in BCR-ABL-expressing cells and upregulated by IFN-α. Mice lacking ICSBP expression develop a myeloproliferative syndrome resembling chronic-phase CML. Why some Ph-positive cells are or become resistant to IFN-α is unknown. Clinical observation indicates that IFN-α’s therapeutic effect decreases over time. IFN-α’s effect is much greater in early rather than in late chronic-phase disease, and it is minimal in disease that progresses from chronic to the accelerated or blastic phases (Faderl S, 1999). This declining effect implies the development

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of other genomic abnormalities that make the cells more resistant to IFN-α. Sensitivity to IFN-α may depend primarily on the amount of BCR-ABL tyrosinekinase oncoprotein present in the blood (Pane F, 1999). The identiÞcation of other genomic abnormalities that play a role in determining response may provide a basis for a more rational use of IFN-α, either alone or in combination. The use of IFN-α in CML was Þrst evaluated by the Houston group at the M.D. Anderson Cancer Center almost 20 years ago (Talpaz M, 1987). Since then, several studies by single-institution or cooperative groups using recombinant IFN-α-2a, -2b, or -2c have conÞrmed the efÞcacy of IFN-α in CML. The M.D. Anderson Cancer Center used a dose of 5 MIU/m2 daily for the treatment of 274 patients with early chronic-phase CML (Kantarjian HM, 1996). Of these patients, 80% achieved a CHR and 58% displayed a cytogenetic response (complete, 26%; major, 38%). The median survival was 89 months. Achieving a cytogenetic response after 12 months of therapy was associated with a statistically longer survival. At Þve years, the survival rates were 90% for CCR, 88% for MCR, and 76% for minor response. Several randomized studies have investigated the relationship of duration of the chronic phase and survival to IFN-α treatment dose and schedule. An Italian study involving 322 patients demonstrated a 6-year survival rate of 50% with IFN-α compared with 29% with chemotherapy (Italian Cooperative Group on Chronic Myeloid Leukemia, 1994). The median survival of the 218 patients who were assigned to IFN-α and the 104 patients who were assigned to chemotherapy was 76 months and 52 months, respectively. The proportion of the patients who were projected to be alive after ten years was 29% in the IFN-α arm and 17% in the chemotherapy arm. The median time from diagnosis to progression was 74 months with IFN-α and 46 months for chemotherapy. The median survival and the 10-year survival rate of Sokal’s low-risk patients achieving hematologic and cytogenetic responses were 104 months and 47%, respectively, in the IFN-α arm versus 64 months and 30%, respectively, in the chemotherapy arm (Italian Cooperative Group on Chronic Myeloid Leukemia, 1998). A worldwide collaborative overview of 1,554 patients randomized and assigned to treatment in seven trials aimed to establish whether patients beneÞt from treatment with IFN-α (Chronic Myeloid Leukemia Trialists’ Collaborative Group, 1997). The regimens that involved IFN-α produced statistically significantly better survival than those that involved hydroxyurea or busulfan. The Þve-year survival rates were 57% with IFN-α and 42% with chemotherapy. Two French multicenter trials studied the potential beneÞt of combining IFNα and LDAC (IFN+LDAC) as Þrst-line treatment (Guilhot F, 1997[a]; Guilhot F, 1997[b]). Patients were randomized to receive IFN-α 5 MU/m 2 or the same dose of IFN-α plus monthly courses of LDAC at a dosage of 10 mg/m2 /day for 10 days. Of 207 evaluable patients after a median follow-up of 85 months, 29 of 103 patients (28%) in the IFN+LDAC group achieved a CCR, whereas 21 of 104 patients (20%) treated with IFN-α obtained this result; median survival was 77 months in the IFN+LDAC group and 65 months in the IFNα group.

408

CHRONIC MYELOGENOUS LEUKEMIA

In the second study, the dose of cytarabine was increased to 20 mg/m2 /day. The trial enrolled 810 patients; 721 were studied (360 randomly assigned to the IFN+LDAC group and 361 to the IFN-α group). An update of this trial showed that the probability of achieving an MCR at 24 months was signiÞcantly higher in the IFN+LDAC group. In addition, the patients in the IFN+LDAC group survived signiÞcantly longer than those in the IFN-α group. At three years, the estimated survival rates were 85.7% and 79.1% for the IFN+LDAC group and the IFN-α group, respectively. The Italian Cooperative Study Group on CML conducted a similar randomized trial. Five hundred and forty evaluable patients were randomized to IFN+LDAC (275) and to IFN-α alone (265). At 12 months, the combination resulted in a higher MCR rate (28% versus 18%) (Baccarani M, 2002). Although this difference was signiÞcant, it did not translate to better survival. A Phase III trial using genetic randomization compared allo-SCT with “best available therapy”—IFN-α (Hehlmann R, 2000[a]). Genetic randomization uses the availability of a sibling donor or a matched unrelated donor to determine the treatment of the patient—all patients with an appropriate donor would receive allo-SCT. A preliminary report of this trial found that the projected three-year survival was 64% for the related donors and 80% for the drug therapy group; in the drug therapy group, the three-year survival for patients with a low-risk Sokal or Hasford score was 90%. These results support the Þndings of a registry-based study concluding that the survival beneÞt of allografting over IFN-α becomes apparent only between Þve and ten years after treatment. Prior to this time, patients treated with IFN-α have superior survival (Hehlmann R, 2000[b]). IFN-α has several shortcomings. It has an extremely slow onset of action, it is poorly tolerated (patients often develop ßu-like symptoms), and it requires daily injections. Therapy must be discontinued in 15–25% of patients and the dose reduced in up to 50%. Because hematologic responses take several months to occur, other agents such as hydroxyurea are often used prior to IFN-α therapy to rapidly reduce the leukocyte counts. Cytotoxic Agents Overview. Chemotherapy uses anticancer or “cytotoxic” drugs to destroy cancer cells by disrupting their growth. Cytarabine is commonly used in combination with IFN-α and in chemotherapy regimens to treat blastic-phase CML. The cytotoxic agents hydroxyurea and busulfan were the treatments of choice until IFN-α was introduced into the CML market twenty years ago. The older agents are less costly than IFN-α or imatinib and are orally administered. Once CML enters the blastic phase, no treatment is particularly effective for controlling the disease. Clinicians use a wide variety of multidrug regimens that are based on the treatments for acute myelogenous leukemia and acute lymphocytic leukemia. Commonly used cytotoxic agents are idarubicin (PÞzer’s Zavedos/Idamycin), daunorubicin (SanoÞ-Aventis’s Cerubidin, Bedford’s Cerubidine, generics), and vincristine (Eli Lilly’s Oncovorin, generics).

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Mechanism of Action. Chemotherapeutic drugs work by damaging cancer cells as they undergo division, or mitosis, and preventing their further reproduction. Cells that are at rest (e.g., most normal cells) are much less vulnerable to chemotherapeutic damage. The cell cycle is composed of four distinct phases, G1 , S, G2 , and G0 , during which the cell prepares for and undergoes mitosis. Combination chemotherapy includes drugs that damage cells at different stages in the process of cell division. Using more than one drug increases the chance of killing a greater number of cells. Chemotherapy also affects healthy body tissues that grow constantly (e.g., skin, hair, digestive system), a factor that explains the side effects, such as hair loss, suffered by patients. Cytarabine. In the treatment of chronic-phase CML, cytarabine (PÞzer’s Cytosar-U, generics) is generally used in conjunction with IFN-α. The addition of cytarabine to IFN-α increases the toxicity of the treatment; whether the combination of the two therapies increases response rates is controversial. Cytarabine is metabolized intracellularly into its active triphosphate form (cytosine arabinoside triphosphate). This metabolite then damages DNA via multiple mechanisms including the inhibition of alpha-DNA polymerase, inhibition of DNA repair through an effect on beta-DNA polymerase, and, most importantly, incorporation into DNA. Cytotoxicity is highly speciÞc for the S phase of the cell cycle. One study analyzed the efÞcacy of daily treatment with IFN-α (5 MU/m2 ) combined with LDAC (10 mg) (IFN+LDAC) in 140 patients with Ph-positive early chronic CML (Kantarjian HM, 1999). Results were compared with those in patients receiving IFN-α with or without intermittent LDAC (seven days per month). CHRs were observed in 92% of patients treated with IFN+LDAC, and cytogenetic responses were observed in 74% (major in 50%, complete in 31%). The estimated four-year survival rate was 70%. The incidence of CHR was higher with IFN+LDAC than with intermittent or no LDAC (92% versus 84% versus 80%, respectively); similar results were noted for cytogenetic response (74% versus 73% versus 58%, respectively). Also, the time to achievement of a major cytogenetic response was signiÞcantly shorter than was obtained with previous IFN-α regimens. Another study randomized 721 patients with Ph-positive, early chronic-phase CML to receive treatment with either hydroxyurea (50 mg/kg) and IFN-α (5 MU/m2 /d) or hydroxyurea, IFN-α, and monthly courses of LDAC (20 mg/m2 for 10 days/month) (Guilhot F, 1997[b]). The rate of CHR was 66% in the IFN+LDAC group versus 55% in the IFN-α/hydroxyurea group. The cytogenetic response rate was 66% in patients treated with LDAC (major in 41%, complete in 15%), which was signiÞcantly higher than the 52% response rate (major in 24%, complete in 9%) among patients treated with IFN-α/hydroxyurea. Patients in the IFN+LDAC group had a signiÞcantly better survival rate than patients in the IFN-α/hydroxyurea group: three-year survival rates were 86% versus 79%. Cytarabine is generally administered as an intravenous preparation. An oral form of cytarabine is available in Japan.

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Hydroxyurea. Hydroxyurea (also known as hydroxycarbamide) (Bristol-Myers Squibb’s Hydrea, generics) effectively produces rapid hematologic responses and controls overall tumor burden in CML, as assessed by white blood cell count and spleen size. Although up to 80% of patients achieve hematologic remissions with hydroxyurea, cytogenetic responses are rare, and when they occur, are normally transient. Hydroxyurea is superior to busulfan, but inferior to IFN-α therapy, in terms of both survival rates (median survival 56 months with hydroxyurea and 44 months with busulfan) and toxicity (Hehlmann R, 1993). Hydroxyurea is also used to treat acute myeloid leukemia, head and neck cancers (before radiotherapy treatment), and ovarian cancer. Hydroxyurea acts primarily as an inhibitor of ribonucleotide reductase. Inhibition of this protein leads to the depletion of essential DNA precursors. Another proposed mechanism of cytotoxicity involves direct chemical damage to DNA by hydroxyurea or a metabolite. Hydroxyurea also inhibits repair of DNA damaged by chemotherapy or radiation, offering potential synergy between hydroxyurea and radiation or alkylating agents. Laboratory studies suggest that hydroxyurea acts selectively against the episomes responsible for drug resistance. Hydroxyurea is speciÞc for the S-phase of the cell cycle. Because of its rapid onset of action, hydroxyurea can be used to reduce the leukemic burden in newly diagnosed patients before initiating IFN-α or progenitor stem-cell transplantation. Hydroxyurea is also used in patients with refractory disease or those who are intolerant of Þrst-line therapy. Common side effects associated with hydroxyurea are threefold. They include a temporary drop in bone marrow function, causing a fall in white blood cell count that increases the risk of severe infection; a drop in red cell count (anemia), causing fatigue and shortness of breath; and a drop in platelet numbers in the blood, causing bleeding or bruising. Busulfan. Low-dose therapy with busulfan (GlaxoSmithKline’s Myleran) (Figure 3) was once the mainstay of treatment for CML. The superior survival rates produced by imatinib, IFN-α, and hydroxyurea have reduced busulfan’s role to that of myeloablative therapy prior to allo-SCT. This medication is also used to treat other disorders of the blood or bone marrow (e.g., myeloproliferative disorder, thrombocytosis, myeloÞbrosis). Busulfan is a bifunctional alkylating agent that has been in clinical use since 1959. Carbonium ions rapidly form after systemic absorption of busulfan, leading to alkylation of DNA. This alkylation results in breaks in the DNA molecule as well as cross-linking of the twin strands, thus interfering with DNA replication and transcription of RNA. The antitumor activity of busulfan is cell cycle phase–nonspeciÞc. Selective effects on granulocytopoesis are not well understood. H3C

O SO2

CH2 CH2 SO2 O CH3 CH2 CH2

FIGURE 3. Structure of busulfan.

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In the treatment of CML, busulfan is most commonly used in high doses as a myeloablative agent in patients receiving progenitor stem-cell transplantation. The dose of busulfan in this setting depends on the protocol, ranging from 8 to 16 mg/kg given over four days. The major dose-limiting effects of busulfan are myelotoxicity and pancytopenia. Myelotoxicity may be increased in patients who are recovering from the effects of prior chemotherapy or those who have received radioactive phosphorus or radiation to marrow-bearing bones. Busulfan can lower the body’s ability to Þght an infection, as well as prevent normal blood clotting. Other side effects associated with busulfan include hyperpigmentation, pulmonary toxicity, abnormal gonadal function, seizures, and veno-occlusive disease. Stem-Cell Transplantation Stem-cell transplantation (SCT) includes both bone marrow transplantation and peripheral blood cell transplantation. SCT initially involves the collection of precursor hematopoietic cells (stem cells) from either the bone marrow (resulting in a bone marrow transplant) or the peripheral blood (resulting in a peripheral blood cell transplantation). The stem cells can be harvested either from a healthy HLA-matched donor (allogeneic SCT [allo-SCT]) or from the patient (auto-SCT). The stem cells are preserved while high-dose myeloablative chemo- and radiotherapy is administered. The cells are then reinfused into the patient, whereupon they migrate to the bone marrow to aid hematopoietic recovery and “rescue” the patient from lethal myelotoxicity. In the treatment of leukemia, the aim of high-dose therapy and transplantation is to destroy the defective blood progenitor stem cells and replace them with healthy, nonleukemic cells. In autologous peripheral stem-cell transplantation (APSCT), the destruction of the patient’s bone marrow cells also destroys the patient’s own T cells, preventing the patient from rejecting the donor progenitor stem cells. In the context of this report, we consider PSCT to include both the myeloablative treatment and the reinfusion of progenitor stem cells. Myeloablative Regimens. Two common myeloablative regimens are used in the treatment of CML to destroy patients’ bone marrow prior to PSCT: total body irradiation and cyclophosphamide (TBI-CY) or busulfan and cyclophosphamide (BU-CY). In patients who undergo transplantation in the chronic phase of CML, the two regimens have comparable transplant-related mortality and efÞcacy rates, but the BU-CY regimen is reportedly better tolerated (Clift RA, 1999). Patients with more advanced disease who are treated with BU-CY have worse transplantrelated mortality and survival rates than those treated with TBI-CY (Ringd´en O, 1999). Allogeneic Stem-Cell Transplantation. Although high-dose therapy followed by allo-SCT is considered the only curative treatment for CML, the

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procedure’s toxicity has always restricted its use to younger (generally younger than 55), Þtter patients. The number of patients receiving allo-SCT is further limited by the number of available matched donors. As previously noted, although long-term survival rates are higher for allo-SCT than for IFN-α, the high early mortality associated with transplantation means that the survival beneÞt does not become apparent until Þve to ten years after transplantation. Allo-SCT is more successful if it is performed in the chronic phase rather than in the accelerated or blastic phases (long-term disease-free survival rates are 50–60%, 30–40%, and 20–25%, respectively; Kantarjian HM, 2001). Aside from the phase in which allo-SCT is performed, several other factors can increase the risk of mortality. Table 6 summarizes these factors, which can be used to calculate an overall risk score that may help determine whether allo-SCT should be undertaken (Tura S, 1999). Survival appears to be best among patients who receive their transplant within one year of diagnosis, are less than 40 years of age, have a male donor, and, along with their donor, are cytomegalovirus-seronegative (Gale RP, 1998). For TABLE 6. Prognostic Scoring System for Allogeneic Progenitor Stem-Cell Transplantation in Chronic Myelogenous Leukemia Factor

Score

Donor type HLA-identical sibling donor Matched unrelated donor

0 1

Disease stage Chronic phase Accelerated phase Blastic phase or higher chronic phase

0 1 2

Age of recipient <20 years 20–40 years >40 years

0 1 2

Sex combination For all except male recipient/female donor Male recipient/female donor

0 1

Time from diagnosis to transplant 12 months >12 months

0 1

Note: A patient’s overall risk score for allo-SCT (allogeneic stemcell transplantation) in CML is calculated by combining the individual risk scores above. If the total score is 1 or less, the patient is a good candidate for allo-SCT (treatment-related mortality [TRM] of approximately 20%). If the total score is 5 or more, the patient is a poor candidate for transplantation (TRM > 70%). For patients with a score of 2–4 (which accounts for 71% of patients), the choice of transplantation would be made on an individual basis depending on the Sokal score (Tura S, 1999). HLA = Human leukocyte antigen.

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such a cohort, Þve-year disease-free survival is 70% to 80%, and the relapse rate is 10% to 20% (Clift RA, 1996). The details of transplant procedure and the source of stem cells (marrow versus peripheral) also inßuence outcome (Savage DG, 1997). Engraftment appears to be more rapid following a peripheral blood stem-cell transplant, though the incidence of chronic graft-versus-host disease (GVHD) may be increased (Bensinger W, 2001). Acute and chronic GVHD is a signiÞcant cause of transplant-related mortality. A combination of cyclosporine and methotrexate is the usual prophylaxis treatment, but controversy surrounds the question of what is the best approach. Researchers have demonstrated that T-cell depletion of the graft substantially reduces the incidence of GVHD but also increases the risk of relapse (Goldman JM, 1988). Allo-SCT using T-cell depletion with the CD52 monoclonal antibody Campath results in an actuarial relapse rate of 60% to 70%. Other methods of T-cell depletion include the use of other antibodies and E-rosette formation/soybean lectin agglutination. They also reduce the incidence of GVHD while increasing the incidence of relapse to varying degrees (Ho VT, 2001). A study involving 22 patients showed that transplantation of highly puriÞed CD34+ peripheral blood stem cells with T-lymphocyte add-back does not appear to be associated with an increased risk of relapse; this strategy is being tested further (Elmaagacli AH, 2003). Studies have suggested that prior treatment with IFN-α is associated with inferior survival, although reports conßict on the matter (Hehlmann R, 1999, Lee SJ, 2001, Giralt S, 2000). However, preliminary data from the European Group for Blood and Marrow Transplantation (EBMT) suggest that treatment with IFN-α’s successor, imatinib, does not have a deleterious effect on subsequent transplantation (Deininger MW, 2001). This phenomenon requires further observation. Autologous Progenitor Stem-Cell Transplantation. Unlike allo-SCT, auto-SCT is not curative. However, auto-SCT can effectively induce short second chronic phases in patients with accelerated- or blastic-phase disease. The use of auto-SCT in chronic-phase patients is considered experimental. A study analyzed the outcome of 581 patients receiving auto-SCT for CML in the Þrst chronic phase reported to the EBMT between 1983 and 1998 (Olavarria E, 2002). Of 207 patients evaluable for cytogenetic analysis within six months of SCT, 17% achieved a CCR, 16% achieved a major response, 36% achieved a minor response, and 31% displayed no cytogenetic response. Results of the cytogenetic analysis within one to two years of SCT were available for 117 patients, the majority of whom received IFN-α post SCT. Of these patients, 15% achieved a CCR, 15% achieved a major response, 20% achieved a minor response, and 50% displayed no cytogenetic response. Patients in CCR or major remission at one to two years post SCT had a ten-year survival of 66% compared with 36% for patients in minor remission or those who displayed no response. The Þve-year survival for patients receiving IFN-α post SCT was 72% compared

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with 61% for patients not treated with IFN-α. Out of 155 patients refractory to IFN-α pre SCT, 70% achieved a cytogenetic response post SCT; this response was complete or major in 31%. EMERGING THERAPIES Following the entrance of the protein tyrosine kinase (PTK) inhibitor imatinib (Novartis’s Gleevec/Glivec) into the chronic myelogenous leukemia (CML) marketplace in 2001, the agent dramatically transformed the market, and PTK inhibitors have dominated research and development in the Þeld ever since. Since 2001, imatinib has entirely replaced interferon-α (IFN-α) as the agent against which all potential newcomers are measured. Several agents that were trialed against IFN-α prior to the launch of imatinib have had to be reevaluated for synergism with imatinib or for activity in imatinib-refractory patients—indeed, the high and durable response rates imatinib has achieved have caused several companies to discontinue development of competing therapies. However, imatinib’s shortcomings deÞne the nature and extent of unmet needs in the treatment of CML. The leaders among the emerging therapies for CML are the PTK inhibitors BMS-354825 and AMN-107, under development by Bristol-Myers Squibb and Novartis, respectively. Other novel promising agents include the heat-shock protein (HSP) inhibitors, peptide vaccines, farnesyl transferase inhibitors (FTIs), and hypomethylating agents. Table 7 lists the emerging therapies’ stages of development. Other drugs in development for the treatment of CML include rapamycin analogues, histone deacetylase inhibitors (HDACIs) such as suberoylanilide hydroxamic acid (SAHA), and anti-bcl-2 antisense oligonucleotides. A preclinical study has reported that the mammalian target of rapamycin (mTOR) inhibitor, rapamycin, synergized with imatinib against BCRABL–positive myeloid and lymphoid cells and increased survival in a murine CML model (Mohi G, 2004). In addition, rapamycin + imatinib combinations inhibited imatinib-resistant mutants of BCR-ABL. The possibility of combining HDACIs with the HSP-90 antagonist 17-AAG was raised in a recent study as a novel strategy against BCR-ABL–positive leukemias, including those resistant to imatinib (Rahmani M, 2004). Companies developing agents for CML may apply for orphan drug status for their molecule in any of the major markets. Orphan drug status was introduced in 1983 to give pharmaceutical companies a Þnancial incentive to develop products to treat rare diseases. In the United States, a condition is considered rare if it affects fewer than 200,000 people in the country. Manufacturers of compounds with orphan drug status receive several beneÞts: seven years of market exclusivity if the product is subsequently approved, a federal tax credit for clinical research costs, exemption from the FDA registration fee, and protocol assistance. They may also be eligible for additional grants. The introduction of orphan drug status has dramatically increased the number of drugs being developed to treat rare conditions.

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TABLE 7. Emerging Therapies in Development for Chronic Myelogenous Leukemia Compound

Development Phasea

Protein tyrosine kinase inhibitors BMS-354825 United States II Europe II Japan II AMN-107 United States Europe Japan

Marketing Company

Bristol-Myers Squibb Bristol-Myers Squibb Bristol-Myers Squibb

I I —

Novartis Novartis —

Ib — —

Kosan Biosciences/NCI/UK Institute of Cancer Research — —

Vaccines AG-858 (HSPPC-70-C) United States Europe Japan

II II —

Antigenics Antigenics —

PR1 peptide antigen United States Europe Japan

II — —

M.D. Anderson Cancer Center — —

Heat-shock protein inhibitors 17-AAG United States Europe Japan

BCR-ABL fusion peptide vaccine United States II Europe — Japan —

Memorial Sloan-Kettering Cancer Center — —

Farnesyl transferase inhibitors Tipifarnib (R-115777; Zarnestra) United States Europe Japan Lonafarnib (Sch-66336; Sarasar) United States Europe Japan

I — —

— —

Hypomethylating agents Decitabine (DAC) United States Europe Japan

II — —

SuperGen/MGI Pharma — —

I — —

Janssen Pharmaceutica/Johnson & Johnson — — Schering-Plough

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TABLE 7. (continued) Development Phasea

Marketing Company

Antineoplastic agents Homoharringtonine (Ceflatonin) United States Europe Japan

II — —

ChemGenex — —

Apoptosis stimulators Arsenic trioxide (Trisenox) United States Europe Japan

II — —

Cell Therapeutics — —

Compound

a Agent may be in later stages of development for other indications.

The long median survival of patients with chronic-phase CML means that initial response rates to investigational drugs are frequently used as surrogate measures of efÞcacy in clinical trials. These initial response rates have historically been the ability to achieve a complete hematologic response (a white blood cell count between 4,000 and 11,000 per mm3 ) or complete cytogenetic response (the absence of cells expressing the Philadelphia chromosome). However, because of the success of imatinib at achieving these endpoints, measuring molecular response (a reduction in quantities of BCR-ABL transcripts, as detected by polymerase chain reaction [PCR], is becoming increasingly important for evaluating novel therapies. A complete molecular response occurs when there is no evidence of BCR-ABL transcripts, indicating disease eradication. Protein Tyrosine Kinase Inhibitors Overview. PTKs are enzymes that catalyze the phosphorylation of tyrosine residues. These enzymes are involved in cellular signaling pathways and regulate key cell functions such as proliferation, differentiation, antiapoptotic signaling, and neurite outgrowth. Unregulated activation of these enzymes, through mechanisms such as point mutations or overexpression, can lead to various forms of cancer as well as benign proliferative conditions. More than 70% of the known oncogenes and proto-oncogenes involved in cancer encode PTKs. Two PTK inhibitors, Bristol-Myers Squibb’s BMS-354825 and Novartis’s AMN-107, are detailed in this section. Lack of published data precludes discussion of other PTK inhibitors in development for CML. Ariad Pharmaceuticals’ AP-23464, and Wyeth’s SKI-606 are two such agents. Mechanism of Action. The rationale for developing tyrosine kinase inhibitors for the treatment of cancer is based on the observation that tyrosine kinase enzymes are critical components of the cellular signaling apparatus and are regularly mutated or otherwise deregulated in human malignancies. Novel tyrosine kinase inhibitors are designed to exploit the molecular differences between tumor cells and normal tissues. In CML, affected cells have a consistent cytogenetic abnormality—the Philadelphia (Ph) chromosome, which carries a BCR-ABL

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fusion gene encoding a tyrosine kinase oncoprotein. Imatinib, the treatment of choice for CML, is a speciÞc inhibitor of this oncoprotein. The development of resistance to imatinib is a frequent setback in patients with advanced phases of the disease. This resistance is frequently the result of mutations in the BCR-ABL kinase domain and overexpression of tyrosine kinases that are not inhibited by imatinib. The majority of amino acid substitutions that cause resistance to imatinib impair the ability of the kinase to adopt the speciÞc closed conformation to which imatinib binds, while a small fraction of these substitutions directly interfere with drug binding. This insight suggests that other small-molecule ABL kinase inhibitors with less stringent requirements for binding could be effectively deployed to combat resistance. BMS-354825. Bristol-Myers Squibb’s BMS-354825 is an oral agent that inhibits Þve tyrosine kinase proteins. They include BCR-ABL, the protein that accounts for abnormal cell growth in CML, and steroid receptor coactivator (SRC) proteins that may play a role in imatinib resistance. The compound has completed Phase I trials at the M. D. Anderson Cancer Center in Houston, Texas, and the University of California at Los Angeles (UCLA) in patients with imatinibresistant CML, and is now in Phase II. The Phase I study involved patients with Ph chromosome–positive chronic, accelerated, and blastic-phase CML patients (36, 8, and 21, respectively) who were either resistant to or intolerant of imatinib (Sawyers CL, 2004; Talpaz M, 2004). Of the chronic-phase patients, 86% demonstrated a complete hematologic response (CHR). In 29 patients for whom cytogenetic data were available, 8 (28%) demonstrated a major cytogenetic response (MCR). In the acceleratedphase patient group, 75% demonstrated a hematologic response. At the time of presentation, no cytogenetic responses were seen in patients with accelerated disease. In the blastic-phase patient group, 79% demonstrated a hematologic response. Of 15 patients for whom cytogenetic data were available, 53% demonstrated a MCR. No patients discontinued the Phase I study because of toxicity, though several patients experienced serious side effects. In the chronic-phase population, 3 of 26 patients for whom data were available had grade 4 thrombocytopenia requiring treatment modiÞcation, and two patients were reported as having gastrointestinal bleeding possibly related to BMS-354825. Two patients in the blastic phase had evidence of tumor lysis syndrome (a serious complication of cancer therapy that causes metabolic abnormalities), and one patient with accelerated-phase CML had pneumonia possibly related to BMS-354825. Additional side effects reported in all phases during the study included arthralgia, pyrexia, fatigue, peripheral edema, headache, and diarrhea. In vitro studies of BMS-354825 have demonstrated more than 500-fold increased potency relative to imatinib versus BCR-ABL; more importantly, BMS354825 retained activity against 14 of 15 imatinib-resistant BCR-ABL mutants (Shah NP, 2004). The compound also proved equally effective against several preclinically and clinically derived tumor models of imatinib resistance (Lee FY, 2004).

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BMS-354825 is not a P-glycoprotein (Pgp) substrate, a feature that may eliminate the blood-brain-barrier problems experienced with imatinib. Instead, imatinib is a substrate of Pgp, the gene product of MDR1 (Hegedus T, 2002). The generally accepted action of MDR1 (MDR stands for multidrug resistance) is to reduce intracellular drug accumulation through Pgp-mediated efßux, thereby hampering the achievement of effective drug levels at the target site. One weakness of BMS-354825 is that, like imatinib (and AMN-107), it does not target the T315i mutation. This mutation has been identiÞed in patients who present with or progress to the accelerated or blastic phase and accounts for 15–20% of imatinib-resistant CML cases. If BMS-354825 progresses quickly through clinical trials, it could initially be approved for the treatment of CML in patients who have failed on imatinib therapy or who present in accelerated- or blastic-phase disease. Once the drug’s efÞcacy is proven in these subsets of patients, studies may be conducted to compare BMS-354825 with imatinib in Þrst-line therapy and to investigate its potential in combination with imatinib. However, considering the very high cytogenetic response rates achieved by imatinib and the absence of data demonstrating that BMS-354825 confers a complete molecular response rate (i.e., cure rate) superior to that of imatinib, BMS-354825 is unlikely to usurp imatinib’s role in Þrst-line therapy. AMN-107. Novartis’s AMN-107 is an oral novel aminopyrimidine ATPcompetitive inhibitor of the protein tyrosine kinase BCR-ABL and some mutant forms of this protein. The compound is in Phase I trials at the M.D. Anderson Cancer Center and the Johann Wolfgang Goethe University in Frankfurt, Germany. It is being studied in CML patients with accelerated- or blastic-phase disease that is imatinib-resistant. Once an effective dose has been determined, the study will expand to treat chronic-phase CML patients who have experienced imatinib failure. Sixty-Þve imatinib-resistant patients were involved in the Phase I study; 36 had accelerated-phase CML, 15 had myeloid blast-phase CML, 7 had lymphoid blastphase CML, and 7 had acute lymphocytic leukemia (ALL) (Giles F, 2004). Each patient received escalating oral doses of AMN-107, starting at 50 mg/day up to 1,200 mg/day. Cytogenetic responses were observed in 20% of accelerated-phase patients, 8% of myeloid blastic-phase patients, and 17% of lymphoid blastic-phase patients. Preclinical data reported at the 46th American Society for Hematology (ASH) meeting in 2004 showed that AMN-107 was 10- to 20-fold more potent than imatinib in vitro against wild-type BCR-ABL kinase and against BCR-ABL–dependent cell proliferation (Golemovic M, 2004). AMN-107 was also effective against clinically relevant imatinib-resistant mutations G250E, E255K/V, E292K, F317L, and M351T (Mestan J, 2004). Additional data presented at ASH show that AMN-107 is more effective than imatinib at inhibiting both BCR-ABL–positive cell lines (e.g., K562, KCL-22, Lama-84) and primary cells derived from two BCR-ABL–positive CML patients who were resistant

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to imatinib and one newly diagnosed chronic-phase CML patient (Le Coutre P, 2004). In a further study, AMN-107 was tested against BCR-ABL–positive cell lines (K562, Lama-84, AR230, BaF/BCR-ABL) and was found to inhibit these cells at nanomolar concentrations; these concentrations were 100-fold less than imatinib (Mahon FX, 2004). AMN-107 is unlikely to be used in combination with imatinib because both drugs have a similar mode of action. However, AMN-107, like imatinib and BMS-354825, does not inhibit the T315i mutation. This drawback presents a clinical problem because, as mentioned previously, this mutation is observed in 15–20% of patients who progress to the accelerated and blastic phases and become imatinib-resistant. Nonetheless, patients with chronic-phase CML who fail to achieve a CCR with imatinib (approximately 25% of patients), patients who present in the accelerated and blastic phases, and patients who relapse on imatinib (i.e., are no longer in cytogenetic remission) could beneÞt from AMN107, especially if its toxicity proÞle is found superior Heat-Shock Protein Inhibitors Overview. Heat-shock proteins (HSPs), also called stress proteins, are a group of proteins that are present in all cells. They are induced when a cell undergoes various types of environmental stresses such as heat, cold, and oxygen deprivation. HSPs are also present in cells under normal conditions. They act like “chaperones,” regulating the folding and degradation of key signaling molecules in cells. HSPs are also believed to play a role in the presentation of peptides or antigens on the cell surface to help the immune system recognize diseased cells. One particular HSP, HSP90, has emerged as an important drug target for anticancer agents because it functions as a chaperone of oncogenic proteins, which are critical to the proliferation and survival of tumor cells. As a result, HSP90-directed drugs have the potential to inhibit the growth of a wide range of cancer cells in both solid tumors and blood-based cancers. Mechanism of Action. Disruption of the function of HSP90 by HSP inhibitors reduces the stability of the CML oncogenic protein BCR-ABL and facilitates its proteosome-mediated degradation. This degradation, in turn, lowers BCRABL levels and induces apoptosis and differentiation of BCR-ABL–positive leukemia cells. 17-AAG. Note: at the time of composing this reference, this drug has become classiÞed under orphan drug status. 17-AAG (17-allylaminogeldanamycin) is a semisynthetic derivative of geldanamycin and a polyketide inhibitor of HSP90 under development by Kosan Biosciences in collaboration with the National Cancer Institute (NCI) and the U.K. Institute of Cancer Research. The compound is in Phase II development as a monotherapy for melanoma and in Phase Ib development as a combination therapy with imatinib for CML. KOS-953 is a 17-AAG variant in development by Kosan that replaces the dimethyl sulfoxide (DMSO)–egg lecithin vehicle

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CHRONIC MYELOGENOUS LEUKEMIA

used in the original formulation; the aim is to improve patient tolerability and provide greater stability. This molecule is in Phase I development for multiple myeloma (MM). An organic, solvent-free formulation of 17-AAG, CNF-1010, is being developed for the treatment of various cancers by Conforma. In October 2004, Kosan Biosciences’ 17-AAG was granted orphan drug status for the treatment of MM and CML. 17-AAG is not orally bioavailable; thus, the Institute of Cancer Research is also investigating orally bioavailable analogues (Phase I) in collaboration with RiboTargets. 17-AAG is an ansamycin antibiotic that binds with high afÞnity into the ATP binding pocket in HSP90, inhibiting its function and inducing the degradation of the BCR-ABL oncoprotein that requires this chaperone for conformational maturation. Investigators presented preclinical data on 17-AAG at the 44th ASH meeting in 2002. The data demonstrated that the combination of 17-AAG and imatinib in CML cells is synergistic in imatinib-resistant cells, including K562, EM-3, LAMA-84, and BV173 cells. The same combination also demonstrates synergy in imatinib-naive CML cells. Similar data were also presented in December 2004 at the 46th ASH meeting. (Bali P, 2004). Another preclinical study showed that Pgp activity was increased in imatinibresistant cells (Radujkovic A, 2004). Monotherapy with 17-AAG decreased Pgp activity in a dose-dependent fashion. The synergistic effect of both drugs in imatinib-resistant cells may thus be explained by increased intracellular levels of imatinib following 17-AAG treatment. Vaccines Overview. CML is among the malignancies most susceptible to immune recognition and eradication. The strongest evidence for this immune response is the graft-versus-leukemia effect, which is most clearly seen in CML after allogeneic stem-cell transplantation (allo-SCT) and donor lymphocyte infusion (DLI). The cancer vaccines being developed are designed to induce antigen-speciÞc antitumor immune responses against the speciÞc cancer cells present in the patient. Problems with production, storage, and administration could deter the acceptance and use of vaccines in the clinic, especially because of the availability of an effective oral drug, imatinib. Among patients treated with imatinib, though, only 39% achieve an MMR (deÞned as a reduction in BCR-ABL transcript levels of at least 3 log). Therefore, vaccines might instead be used to control and reduce minimal residual disease levels (the BCR-ABL transcript level) in patients being treated with drug therapy. Mechanism of Action. Some vaccines contain antigens or parts of antigens puriÞed from cancer cells obtained from the patient or from another individual. DNA vaccines are being tested that contain the DNA encoding the speciÞc antigen. In some approaches, cells are isolated in the laboratory and start making antibodies after the cancer antigen is inserted. In each case, the vaccines’ method of action is to render the cancer cells susceptible to immune attack by cytotoxic

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T lymphocytes (CTLs) by heightening the recognition of markers within cancer cells. The goal is to encourage the patient’s immune system to inhibit the growth of cancer cells. AG-858. Antigenics’ AG-858 (HSPPC-70-C) is a personalized HSP70 cancer vaccine. A Phase II trial of the agent in combination with imatinib is underway. AG-858 is a vaccine comprising HSP70-peptide complexes that have been isolated from individual patients’ cancer cells. Patients undergo a blood-Þltering process called leukopheresis, during which white blood cells (WBCs) are collected. The WBCs are then sent to Antigenics for production of the personalized vaccines. HSPs are a family of chaperone proteins that participate in the degradation process of intracellular proteins. Because it associates with multiple potentially antigenic peptides and cannot interact with antigen-presenting cells (APCs), HSP70 was proposed as an approach to develop tumor-speciÞc vaccines. When the patient is vaccinated, the HSP70-peptide complexes interact with the immune system’s APCs at the site of injection. The HSP complexes bind to the CD91 receptor on APCs, and are taken into the cells. The APCs then travel to the lymph nodes where they re-present the antigenic peptides on their surfaces. This process triggers a response by cytotoxic T-lymphocytes against antigens expressed in cells from which the HSP70 was derived. A Phase II study is evaluating the safety and efÞcacy of AG-858 in combination with imatinib in up to 120 patients in the United Kingdom and the United States with chronic CML who are receiving imatinib but have not achieved complete cytogenetic response (CCR). In a pilot trial, 11 CML patients who had failed to achieve an MCR after six months of imatinib therapy were treated with AG-858 and imatinib (Li Z, 2004). HSP was successfully puriÞed from all patients and, after vaccination, a CCR was achieved in all Þve patients who had completed the eight planned injections; two patients had a complete molecular response (CMR). PR1 Peptide Antigen The M.D. Anderson Cancer Center is investigating the human leukemiaassociated antigen, PR1, as a potential treatment for acute myeloid leukemia (AML) and CML. The agent is in Phase II clinical trials. PR1 is derived from proteinase 3, an aberrantly expressed protein in myeloid leukemia cells, and can be presented on HLA-A2 to CTLs that preferentially kill leukemic cells over normal hematopoietic progenitors. Results were presented at the 46th ASH meeting in 2004 from a trial in which 35 HLA-A2+ patients with AML, myelodysplastic syndrome (MDS), or CML were vaccinated with the PR1 peptide in an incomplete Freund’s adjuvant (an agent used in vaccine therapy to stimulate the immune system) and 75 mg of granulocyte-macrophage colony-stimulating factor (GM-CSF) (Muzaffar H, 2004). The CML patients either had not responded to Þrst-line treatment or had relapsed disease. Patients were assigned at random to receive 0.25 mg, 0.50

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mg, or 1.0 mg PR1 subcutaneously every three weeks for three total injections. Immune responses were assessed by PR1/HLA-A2 tetramer staining and intracellular interferon-gamma (IFN-γ ) production by CTLs. Clinical responses were assessed by bone marrow biopsy both before study entry and three weeks after the third vaccination. The patients had a median age of 50 and were treated for a median of 26 months from time of diagnosis. Follow-up lasted one to four years. Immune responses were elicited in 60% of the evaluable patients versus 0% among patients without an immune response. Progression-free survival for patients with and without immune response was 6.4 months versus 2.4 months, respectively. Of ten CML patients, one achieved a CCR. Three CML patients refractory to allo-SCT, IFN-α, and imatinib experienced stable disease with some hematologic improvement and were able to discontinue hydroxyurea. BCR-ABL Fusion Peptide Vaccine The Memorial Sloan-Kettering Cancer Center in New York is developing a series of peptides from the breakpoint of the BCR-ABL gene as vaccines for the potential treatment of CML. The peptides are in Phase II trials. The peptides are derived from the amino acid sequences crossing the b3a2 breakpoint of the BCR-ABL–generated 210 kDa oncogene product. The peptides have been shown to bind major histocompatibility complex (MHC) class I molecules, thereby eliciting CTL responses. In a Phase II trial, 14 patients with CML in the chronic phase with any HLA type and a b3a2 BCR-ABL breakpoint were vaccinated subcutaneously Þve times over a ten-week period using a preparation of six peptides mixed with the adjuvant Quillaja saponaria (Antigenics’ QS-21) (Cathcart K, 2004). All patients developed delayed-type hypersensitivity (DTH) and/or CD4 proliferative responses after beginning vaccinations, and 11 of 14 patients showed IFN-γ release by CD4 enzyme-linked immunospot (ELISPOT) at one or more time points. Four patients in hematologic remission had a decrease in Ph chromosome percentage; three patients in molecular relapse after allo-SCT became transiently PCR-negative after vaccination. In another Phase II trial, a total of 16 evaluable chronic-phase CML patients who had achieved MCR or CCR that had been stable for at least six months during conventional treatment with IFN-α or imatinib, and who had a b3a2 BCR-ABL breakpoint, were vaccinated subcutaneously six times over a 12-week period (every two weeks) using a preparation of Þve peptides mixed with QS-21 and GM-CSF (Bochia M, 2003). Patients who had an immune response received two booster vaccinations at four- and eight-month intervals after the date of the last vaccination. No signiÞcant toxic effects were observed. Thirteen patients developed CD4 proliferative responses, and 9 out of 16 developed DTH. Of 10 patients who had been previously treated with imatinib, a molecular response was noted in 6 (including three cases with undetectable levels of BCR-ABL). In 6 patients previously treated with IFN-α, 2 showed a response (one cytogenetic and one molecular).

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Farnesyl Transferase Inhibitors Overview. One of the best-recognized downstream events resulting from the tyrosine kinase activity of BCR-ABL in CML patients is the activation of ras. Ras, which is synthesized as an inactive protein in the cytoplasm, is activated through a prenylation process that allows attachment to the cellular membrane. This process is mediated most prominently by farnesyl transferase (FTase) and alternatively through geranyl protein transferase (Beaupre DM, 1999). Mutations of ras and Ras protein activation are frequent features of malignant transformation. Approximately 30% of human cancers have been associated with ras mutations. The frequency of these mutations varies in hematologic malignancies from 5% to 15% in acute lymphoblastic leukemia and up to 65% in CML. Therefore, inhibition of Ras activation has been investigated as an antineoplastic therapy. One approach to ras inhibition is inhibiting FTase via farnesyl transferase inhibitors (FTIs). Preclinical studies have demonstrated that FTIs have signiÞcant anti-CML activity (Peters DG, 2001; Reichert A, 2001). Mechanism of Action. The actual mechanism of action of FTIs is not yet clear because the inhibition of Ras farnesylation does not account for all of the FTIs’ actions. For example, FTIs do not require the presence of mutant Ras protein to produce antitumor effects (Karp JE, 2001). Several other proteins have been implicated as downstream targets that mediate the antitumorigenic effects of FTIs. The regulation of RhoB, a small GTPase that acts downstream of Ras and is involved in many cellular processes, including cytoskeletal regulation and apoptosis, has been proposed as a mechanism of FTI-mediated antitumorogenesis (Lebowitz PF, 1998). Additional proteins involved in cytoskeletal organization are also known to be farnesylated, including the centromere proteins CENPE and CENP-F, protein tyrosine phosphatase, and lamins A and B. Therefore, one possible mode of action of FTIs may be their inhibiting effects on cellular reorganization and mitosis. In addition to inhibiting cellular reorganization and mitotic pathways, it is known that FTIs indirectly modulate several important signaling molecules, including transforming growth factor (TGF) βRII, MAPK/ERK, PI3K/AKT2, Fas (CD95), and vascular endothelial growth factor (VEGF) (Zhang B, 2002). The regulation of these effectors can lead to the modulation of signaling pathways involving cell growth, proliferation, and apoptosis. Tipifarnib. Janssen Pharmaceutica and its parent company, Johnson & Johnson, are developing tipifarnib (R-115777, Zarnestra), an orally bioavailable nonpetidomimetic FTI for the treatment of various hematologic malignancies, including AML, MDS, and CML. Tipifarnib inhibits FTase, preventing Ras from being altered and locating in the cell membrane. Therefore, Ras is not activated, and the signal for cell growth is not transmitted. A Phase I study suggested that tipifarnib’s mechanism of action may involve inhibiting angiogenesis and decreasing the expression and secretion of VEGF (Cortes J, 2004). This study investigated the tolerability and efÞcacy of tipifarnib + imatinib in patients with chronic-phase CML who have failed imatinib (Cortes

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J, 2004). The starting dosage was tipifarnib 300 mg twice daily for 14 days every 21 days and imatinib 300 mg daily, with subsequent levels 300 mg + 400 mg once daily, 400 mg + 400 mg twice daily, and 500 mg + 400 mg three times daily, respectively. The median age was 63 years, and the median time from diagnosis was 68 months. All the patients had failed imatinib, nine had failed IFN-α, and six had failed other therapies. The maximally tolerated dose (MTD) was imatinib 400 mg daily and tipifarnib 400 mg twice daily. One patient died early (16 days on treatment) of unknown causes, nine discontinued therapy after a median of 12 three-week cycles, and six continued therapy after a median of 11+ cycles. Eleven patients started with abnormal WBC counts and nine achieved normalization during therapy. One patient achieved a CCR, one patient achieved an MCR (with a T315I mutation), and two patients displayed minor cytogenetic responses. Two patients lost the response after nine months, and two had ongoing responses more than 12 months after commencing therapy. The researchers concluded that this combination is well tolerated and demonstrates antileukemia activity. Lonafarnib. Schering-Plough’s lonafarnib (Sch-66336; Sarasar) is an orally bioavailable nonpetidomimetic FTI. The compound is in Phase II trials for a variety of difÞcult-to-treat solid tumors and leukemias and in Phase I for CML. Lonafarnib inhibits the proliferation of imatinib-resistant, BCR-ABL–positive cell lines as well as colony formation of cells from imatinib-resistant CML patients. It also sensitizes imatinib-resistant cells to apoptosis with imatinib. A Phase I study investigated lonafarnib in combination with imatinib for patients with CML who had failed imatinib therapy (Cortes J, 2004). The starting dosage for chronic-phase CML was imatinib 400 mg daily + lonafarnib 100 mg twice daily; for the accelerated and blastic phases dosage was 600 mg daily and 100 mg twice daily. A total of 22 patients were treated: 9 in the chronic phase, 10 in the accelerated phase, and 3 in the blastic phase. Prior therapy included imatinib (n = 22), IFN-α therapy (n = 16), and other agents (n = 7). Median age was 59 years, and median time from diagnosis was 51 months. Patients received therapy for a median of 23 weeks. Among 6 patients in the chronic phase evaluable for hematologic response, 1 patient had a CHR and 1 patient achieved an MCR. Among patients in the accelerated and blastic phases, 4 had hematologic responses: one CHR (with mutation F359V), one partial hematologic response (no mutation), and two hematologic improvements (one patient with lymphoid blastic-phase disease achieved marrow cytogenetic response, and one patient with accelerated-phase disease achieved CHR with incomplete platelet recovery). The researchers concluded that the combination of lonafarnib and imatinib is well tolerated and shows early evidence of activity in this refractory population. Hypomethylating Agents Overview. Hypermethylation of DNA in the regulatory area of selected genes has been shown to be common in neoplasia and is associated with tumor resistance or progression.

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Both 5-azacytidine (PÞzer’s Vidaza/Mylosar) and decitabine (Supergen and MGI Pharma’s Dacogen) are potent DNA methylation inhibitors and have shown signiÞcant antileukemic activity in myeloid malignancies, including AML and MDS (Santini V, 2001). In CML, these agents have been used mostly in the accelerated and blastic phases. Mechanism of Action. The Pa promoter of abl and several other genes that are central to the development of CML are hypermethylated in a signiÞcant proportion of patients, and methylation increases with disease progression. The p15 gene is hypermethylated with CML progression, and different patterns of hypermethylation for myeloid and lymphoid blastic phases have been reported (Nguyen TT, 2000). Therefore, hypomethylating agents may play a role in treating CML. Decitabine. Decitabine is a DNA methyltransferase inhibitor under development by Supergen and MGI Pharma for the potential treatment of a range of hematologic malignancies, solid tumors, and sickle cell disease. Decitabine has shown activity in accelerated-phase and blastic-phase CML as a single agent and is now being investigated in combination with other chemotherapeutic agents and imatinib. The agent is in Phase II trials for the treatment of CML. Decitabine is a cytidine analogue that exerts potent DNA hypomethylating effects through its covalent binding to DNA methyltransferase. Decitabine is cytotoxic at high doses, hypomethylating at low doses, and has clinical activity in myeloid malignancies that appears to be optimal at low doses. In a Phase II study, the activity of decitabine monotherapy (at 15 mg/m2 intravenously [IV] over one hour daily, Þve days a week for two weeks) in patients who were either imatinib-intolerant or had CML that was refractory to imatinib was evaluated (Issa J, 2004). Thirty-Þve patients were enrolled (12 in the chronic phase, 17 in the accelerated phase, and 6 in the blastic phase). CHRs were seen in 12 patients (34%), partial hematologic responses were seen in 7 patients (20%), and hematologic improvement was seen in 4 patients (11%); there was an overall hematologic response rate of 65% (83% in the chronic phase, 59% in the accelerated phase, and 50% in the blastic phase). MCRs were observed in 7 patients (20%), and minor cytogenetic responses were seen in 9 patients (26%), with an overall cytogenetic response rate of 46%. Median response duration was three months. The only common grade 3 or 4 toxicities observed were related to myelosuppression. There were two deaths in the study, both related to thrombocytopenia and hemorrhage. Another study showed synergy between imatinib and decitabine (La Rosee P, 2004). Ten patients who presented with untreated accelerated or blastic-phase CML were treated with a combination of decitabine (15 mg/m2 , IV, for 10 days) and imatinib (600 mg po daily). Of the ten patients enrolled, six were evaluable; two patients achieved CHR, and one patient achieved a minor cytogenetic response. If approved for the treatment of CML, decitabine will likely be used in accelerated-phase and blastic-phase disease that is unresponsive to imatinib therapy. The agent is unlikely to be used alone and will be used in combination with imatinib and other therapies.

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Antineoplastic Agents Overview. Although conventional cytotoxic drugs are the mainstay of therapy for most cancers, immunotherapies (and more recently, the PTK inhibitor imatinib) have played a more prominent role in CML. Accordingly, few antineoplastic agents are in active development for CML. We discuss only ChemGenex’s version of homoharringtonine (Ceßatonin) in this section. The Chinese Academy of Medical Science has also launched a version of homoharringtonine. Mechanism of Action. Conventional cytotoxic agents interrupt the DNA replication and repair processes required for functional cell division. They act in several ways, including alkylating DNA, resulting in strand breakage; inhibiting crucial enzymes required for DNA strand formation; and interfering with spindle formation. Homoharringtonine inhibits the initiation of protein, DNA, and RNA biosynthesis. Homoharringtonine. Homoharringtonine (ChemGenex’s Ceßatonin, formerly CGX-653) is an intravenous formulation of a natural product derived from the cephalotaxus evergreen tree. Homoharringtonine affects several cellular pathways, including the regulation of genes associated with apoptosis and angiogenesis. In the several years prior to the launch of imatinib, homoharringtonine was extensively investigated for chronic-phase CML, in Þrst- and second-line therapy, alone, and in combination with IFN-α and/or cytarabine (PÞzer’s Cytosar-U, generics). ChemGenex has initiated a combination Phase II trial of homoharringtonine and imatinib in CML patients who are developing resistance to imatinib. Phase II trials are ongoing in the acute myelogenous leukemia setting. A preclinical study in four paired imatinib-sensitive/resistant cell lines investigated the potency of homoharringtonine, cytarabine, daunorubicin, and hydroxyurea, alone or in combination with imatinib (Tipping AJ, 2002). Primary blasts from advanced-stage, imatinib-refractory CML patients were studied using semisolid media clonogenic assays to test the sensitivity of the tumor cells to homoharringtonine. Investigators found that homoharringtonine achieved major inhibition of CML cell-line proliferation. In a clinical trial, homoharringtonine, combined with IFN-α in the Þrst-line setting, achieved a CHR in 85% of patients, a CCR in 21% of patients, and an MCR in 49% of patients (O’Brien S, 2002). Combined with low-dose cytarabine as a second-line treatment, homoharringtonine induced a CHR among 72% of patients, MCR in 15% of patients, and CCR in 5% of patients (Kantarjian HM, 2000). In a triple-therapy study among patients with early chronic-phase CML, 90 patients received treatment with IFN-α, cytarabine, and homoharringtonine (O’Brien S, 2003). Patients received 5 million units (MU)/m2 IFN-α and cytarabine 10 mg, both subcutaneously daily, and homoharringtonine 2.5 mg/m2 by continuous infusion over 24 hours daily on days 1–5 every 28 days. After a median duration of 16.5 months of therapy, 78 patients switched to imatinib 400 mg orally daily. With the triple regimen, 94% of patients achieved a CHR and 74% achieved a cytogenetic response. The cytogenetic response was complete (Ph-positive cells

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0%) in 22% of treated patients and major in 46% of treated patients. SigniÞcant myelosuppression occurred, resulting in major dose reductions. After 12 months of therapy, the median IFN-α dosage was 1.6 MU/m2 daily, the median cytarabine dosage was 1.85 mg daily, and the median number of homoharringtonine-treated days was two every month. Only three patients developed blastic-phase disease while receiving the triple regimen. After switching to imatinib, and after a median follow-up of 46 months from the start of triple therapy, 63% of patients achieved a CCR and a further 13% achieved an MCR. Nine percent of patients had entered the blastic phase. Investigators estimate that Þve-year survival will stand at 88%. In a small Phase I/II trial, nine patients with Ph-positive accelerated-phase CML previously treated with imatinib received treatment with a semi-synthetic formulation of homoharringtonine by daily subcutaneous injection for seven days, every 28 days (Marin D, 2005). With a median follow-up of 12 months, 80% of patients had achieved a second chronic phase, and 67% of patients had achieved a complete hematologic response. No patient achieved a cytogenetic response. According to investigators, homoharringtonine was well tolerated with minimal nonhematologic toxicity. Grade IV neutropenia was observed in three patients, and grade IV thrombocytopenia requiring platelet support occurred in two patients. All patients were monitored for mutations in the ABL kinase domain, and, in one patient, a kinase domain mutation detected at the start of treatment was no longer detectable after six months of treatment. One problem facing older drugs in clinical trials for CML, such as homoharringtonine and arsenic trioxide, is that physicians would rather place patients in clinical trials testing the new tyrosine kinase inhibitors and use these older agents only as last resorts. However, the primary limitations of homoharringtonine are its hematologic toxicity and its relatively low incidence of CCRs. The hematologic toxicity has restricted the dose of homoharringtonine in clinical trials, and most patients ultimately received only two days, rather than the planned Þve days, of treatment per month. Homoharringtonine’s hematologic toxicity proÞle will probably preclude a role in combination with imatinib in the Þrst-line setting. Apoptosis Stimulators Overview. Apoptosis, or programmed cell death, represents a universal and efÞcient cellular suicide pathway. As an understanding of apoptosis’s vital role in normal cellular development has deepened, numerous genes that encode apoptotic regulators—some of which represent familiar oncogenes or tumor suppressor genes—have been identiÞed. Targeting apoptosis succeeds only if the therapeutic index is good enough to selectively destroy cancer cells rather than normal cells. In drug-curable malignancies, such as common pediatric leukemias and certain solid tumors, apoptosis is a prominent mechanism associated with the induction of tumor remission. Many cytotoxic agents’ ultimate mechanism of action takes place via the general apoptotic pathway.

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Mechanism of Action. Apoptosis is an evolutionarily conserved cell-death pathway that occurs in a variety of physiological situations. An apoptotic stimulus induces an initiation and commitment phase, followed by a degradation phase. This last stage is regulated by cysteine proteases (caspases 1–14) and follows a characteristic pattern of the following morphological changes: membrane rufßing; mitochondrial dysfunction; cytoplasmic and organelle shrinkage; nuclear contraction; and endonuclease activation, resulting in DNA fragmentation. Two central pathways mediate apoptosis: the type I extrinsic or death receptor pathway, which generates an apoptotic signal following the aggregation of death ligands; and the type II or intrinsic pathway, which signals through mitochondria. In some cases, type I activation may also proceed down the mitochondrial pathway. A drug that activates apoptosis might achieve a suitable therapeutic index in several ways. First, it might activate a death cascade via a drug target that is uniquely expressed in a cancer cell. Alternatively, it might be delivered to the target tissue in a manner that is selective for the cancer cell. A third possibility—and perhaps the most promising one—is that such a drug could exploit a pathway activated by oncogenes to provoke apoptosis selectively in cancer cells. It is now clear that oncoproteins can interact with apoptotic regulatory pathways. Thus, overexpression of Myc sensitizes cells to a wide assortment of apoptotic triggers, probably reßecting the role of apoptosis in the intracellular immunity that prevents normal cells from persisting in the body once they acquire cancer-causing genetic defects. However, many human tumors that overexpress Myc are highly resistant to apoptotic triggers, probably owing to a variety of downstream lesions that blunt the death pathway. Still, the recognition that oncogenes can sensitize cells to proapoptotic treatments suggests that if such lesions can be circumvented, drugs that induce cell death could prove highly selective for cancer cells. Arsenic Trioxide. Arsenic trioxide (Cell Therapeutics’ Trisenox) was commonly used in the treatment of leukemia during the Þrst half of the 20th century, prior to the advent of busulfan. Arsenic trioxide has been marketed for the treatment of acute promyelocytic leukemia (APL) in the United States since 2000 and in Europe since 2002. In the late 1990s, several trials investigated the potential of arsenic to treat chronic-phase CML in patients who have failed IFN-α. However, the launch of imatinib in 2001 rendered these data irrelevant almost before they were published. Phase II and initial Phase III data from these trials found that arsenic achieves a major cytogenetic response in 60% of IFN-α intolerant/resistant patients and in 80% of newly diagnosed patients (Tallman M, 2000). More recently, preclinical and clinical investigators have pursued a more clinically relevant role for arsenic trioxide by testing the agent in blastic-phase disease, in imatinib-resistant cells/patients, and in combination with imatinib. In a small Phase I/II study, six patients with chronic-phase imatinib-resistant or relapsed CML received arsenic trioxide and imatinib in an attempt to restore hematologic response and/or induce major cytogenetic response (O’Dwyer M, 2002). Resistant/relapsed disease was deÞned as either “lack of major cytogenetic

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response after six months of therapy” or “loss of previous response, with or without hematologic response.” Imatinib was administered at 400 mg per day, with arsenic trioxide given as a loading dose (0.25 mg/kg/day for Þve days) followed by maintenance therapy (0.25 mg/kg/day two days/week). According to investigators, treatment was well tolerated with predictable and manageable side effects. SigniÞcant toxicities included grade 3 shortness of breath, grade 2 elevations of aspartate amino transferase (AST) and/or amino alanine transferase (ALT), hyperglycemia, lower extremity edema, fatigue, and dyspepsia, each in three or fewer patients. No dose-limiting toxicities were encountered. Nonmyeloablative Transplants Nonmyeloablative transplants (NM-allo-SCTs; also known as mini transplants or reduced-intensity-conditioning hematopoietic stem-cell transplantation) are allogeneic stem-cell transplants (allo-SCT) that are preceded by a low-dose conditioning regimen. This regimen is not as intensive, nor as life-threatening, as traditional transplant regimens, which use high-dose chemotherapy. In nonmyeloablative transplants, the aim of the conditioning regimen is to destroy a sufÞcient number of the patient’s T cells to reduce the patient’s immune response and thereby prevent the donor bone marrow from being rejected. To this end, conditioning regimens containing immunosuppressive drugs such as ßudarabine and cyclophosphamide are used. The success of the transplant relies on the transplanted cells mounting an immunologic graft-versus-leukemia effect against the patient’s leukemic cells. Candidates for these transplants are those deemed at high risk of transplant complications (e.g., relatively older patients or those with compromised organ function). A study by P.A. McSweeney and colleagues used reduced total-body irradiation as the allo-SCT conditioning regimen and included nine patients with CML (McSweeney PA, 2001). Of the six chronic-phase patients receiving transplants, four achieved cytogenetic and molecular responses studied by real-time PCR. The molecular responses lagged behind cytogenetic responses by several months. In three accelerated-phase patients, two had progressive disease, and one achieved a molecular response. Two small studies reported on the outcome of patients following nonmyeloablative transplants (Slavin S, 2000; Sandmaier BM, 2000). In the larger trial (Slavin S, 2000), 21 CML patients in the Þrst or stable second chronic phase underwent NM-allo-SCT. An evaluation of the Þrst 17 patients with a median follow-up of 33 months provided a four-year probability of survival of 87%. Only one patient had a molecular recurrence, which was treated successfully with donor lymphocyte infusion (DLI). In the second trial (Sandmaier BM, 2000), 12 patients with CML were treated in a group of 88 patients with hematologic malignancies. Sixty-eight percent of patients were still alive after a median follow-up of 244 days (range 100–842). Although nonmyeloablative therapy has yet to be tested in a randomized controlled trial, it is already being used in the clinic. Graft rejection has been observed more frequently in nonmyoablative than in myoablative transplant, especially in

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patients with CML and MDS. This rejection is most likely the result of a relatively active donor immune system, which in CML has been spared exposure to cytotoxic and immunosuppressive therapy. Nonetheless, early results suggest that the nonmyeloablative approach may be quite useful in CML, especially for treating chronic-phase disease. Many lines of evidence suggest that CML is especially sensitive to immunologic effects (including the high relapse rates in syngeneic and T-cell-depleted transplants, the beneÞcial effects of DLI, and the effects of interferon). However, the question of whom to apply the nonmyeloablative approach to is problematic. Restricting the procedure to those patients who fail imatinib or to those who have a related or unrelated donor but are too old or too ill for conventional transplantation would conÞne the treatment to a very small patient population. One possible approach might be to add nonmyeloablative transplants as an immunologic adjuvant after initial therapy with imatinib, followed by the addition of imatinib after transplant to clear out any residual disease. REFERENCES Anstrom KJ, et al. Long-term survival estimates for imatinib versus interferon-α plus low-dose cytarabine for patients with newly diagnosed chronic-phase chronic myeloid leukemia. Cancer. 2004;101:2584–2592. Baccarani M, et al. A randomized study of interferon-α and low-dose arabinosyl cytosine in chronic myeloid leukemia. Blood. 2002;99:1527–1535. Bali P, et al. Molecular characterization of human CML cells with resistance to the heat shock protein (hsp) 90 inhibitor 17-allylamino-demethoxy geldanamycin (17-AAG). Proceedings of the American Society of Hematology; 2004. Abstract 1980. Baron S, et al. The interferons. Mechanisms of action and clinical application. Journal of the American Medical Association. 1991;266:1375–1383. Beaupre DM, Kurzrock R. RAS and leukemia: from basic mechanisms to gene-directed therapy. Journal of Clinical Oncology. 1999;17:1071–1079. Bensinger W, et al. Transplantation of bone marrow as compared with peripheral blood cells from HLA-identical relatives in patients with hematologic cancers. New England Journal of Medicine. 2001;344:175–181. Bernards A, et al. The Þrst intron in the human c-abl gene is at least 200 kb long and is a target for translocation in chronic myelogenous leukemia. Molecular and Cellular Biology. 1987;7:3231–3236. Bochia M. Chronic myeloid leukemia. European School of Hematology International Conference; September 28–30, 2003; Rapallo, Italy. Branford S, et al. BCR-ABL levels continue to decrease up to 42 months after commencement of standard dose imatinib in patients with newly diagnosed chronic phase CML who achieve a major molecular response. Proceedings of the American Society of Hematology; 2004. Abstract 274 Buchdunger E, et al. Inhibition of the Abl protein-tyrosine kinase in vitro and in vivo by a 2-phenylaminopyrimidine derivative. Cancer Research. 1996;56:100–104. Cathcart K, et al. A multivalent bcr-abl fusion peptide vaccination trial in patients with chronic myeloid leukemia. Blood. 2004;103(3):1037–1042.

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Chawla-Sarkar M, et al. Apoptosis and interferons: role of interferon-stimulated genes as mediators of apoptosis. Apoptosis. 2003;8(3):237–249. Chronic Myeloid Leukemia Trialists’ Collaborative Group. Interferon alpha versus chemotherapy for chronic myeloid leukemia: A meta-analysis of seven randomized trials. Journal of the National Cancer Institute. 1997;89:1616–1620. Clift RA, Storb R. Marrow transplantation for CML: the Seattle experience. Bone Marrow Transplantation. 1996;17(suppl 3):S1–S3. Clift RA, et al. Long-term follow-up of a randomized study comparing cyclophosphamide and total body irradiation with busulfan and cyclophosphamide for patients receiving allogeneic marrow transplants during chronic phase of chronic myeloid leukemia. Blood. 1999;94:3960–3962. Cortes J, et al. Result of interferon-alpha therapy in patients with chronic myelogenous leukemia 60 years of age and older. American Journal of Medicine. 1996;100:452–455. Cortes J, et al. High-dose imatinib mesylate treatment in patients (Pts) with previously untreated early chronic phase (CP) chronic myeloid leukemia (CML). Proceedings of the American Society of Hematology; 2004. Abstract 999. [a] Cortes J, et al. A Phase I study of tipifarnib in combination with imatinib mesylate (IM) for patients (Pts) with chronic myeloid leukemia (CML) in chronic phase (CP) who failed IM therapy. Proceedings of the American Society of Hematology; 2004. Abstract 1011. [b] Cortes J, et al. Phase I study of lonafarnib (SCH66336) in combination with imatinib for patients (Pts) with chronic myeloid leukemia (CML) after failure to imatinib. Proceedings of the American Society of Hematology; 2004. Abstract 1009. [c] Cortes JE. Natural history and staging of chronic myelogenous leukemia. Hematol Oncol Clin North Am. 2004;18(3):569–584. Crossman LC, O’Brien SG. Imatinib therapy in chronic myeloid leukemia. Hematology/Oncology Clinics of North America. 2004;18(3):605–617. Deininger MW, et al. Glivec prior to allografting is safe and not associated with increased transplant-related toxicity. Proceedings of the American Society of Hematology; 2004. Abstract 5146. Druker BJ, et al. Activity of a speciÞc inhibitor of the BCR-ABL tyrosine kinase in the blast crisis of chronic myeloid leukemia and acute lymphoblastic leukemia with the Philadelphia chromosome. New England Journal of Medicine. 2001;5;344(14): 1038–1042. [Erratum, New England Journal of Medicine. 2001;345:232.] East Anglia Cancer Registry. Cancer Incidence in East Anglia 1998. www.web.org/eacr/annrep.html. Accessed February 7, 2002. Elmaagacli AH, et al. Outcome of transplantation of highly puriÞed peripheral blood CD34+ cells with T-cell add-back compared with unmanipulated bone marrow or peripheral blood stem cells from HLA-identical sibling donors in patients with Þrst chronic phase chronic myeloid leukemia. Blood. 2003;101(2):446–453. Faderl S, et al. Mechanism of disease: the biology of chronic myelogenous leukemia. New England Journal of Medicine. 1999;341:164–172. Faderl S, et al. Monitoring of minimal residual disease in chronic myeloid leukemia. Hematology/Oncology Clinics of North America. 2004;18(3):657–670. Gaigner A, et al. Increase of BCR-ABL chimeric mRNA expression in tumor cells of patients with chronic myeloid leukemia precedes disease progression. Blood. 1988;72: 1237–1241.

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Gale RP, et al. Survival with bone marrow transplantation versus hydroxyurea or interferon for chronic myelogenous leukemia. Blood. 1998;91:1810–1819. Gardembas M, et al. Results of a prospective Phase II study combining imatinib mesylate and cytarabine for the treatment of Philadelphia-positive patients with chronic myelogenous leukemia in chronic phase. Blood. 2003;102(13):4298–4305. Giles F, et al. Phase I/II Study of AMN107, a novel aminopyrimidine inhibitor of Bcr-Abl, on a continuous daily dosing schedule in adult patients (pts) with imatinib-resistant advanced phase chronic myeloid leukemia (CML) or relapsed/refractory Philadelphia chromosome (Ph+) acute lymphocytic leukemia (ALL). Proceedings of the American Society of Hematology; 2004. Abstract 22. Giles FJ, et al. Accelerated and blastic phases of chronic myelogenous leukemia. Hematology/Oncology Clinics of North America. 2004;18(3):753–774. Giralt S, et al. Effect of short-term interferon therapy on the outcome of subsequent HLAidentical sibling bone marrow transplantation for chronic myelogenous leukemia: an analysis from the International Bone Marrow Transplant Registry. Blood. 2000;95: 410–415. Giralt S, et al. Melphalan and purine analog-containing preparative regimens: reducedintensity conditioning for patients with hematologic malignancies undergoing allogeneic progenitor cell transplantation. Blood. 2001;97(3):631–637. Goldman JM, et al. Bone marrow transplantation for chronic myelogenous leukemia in chronic phase: Increased risk of relapse associated with T cell depletion. Annals of Internal Medicine. 1988;108:806–814. Golemovic M, et al. AMN107, a novel aminopyrimidine inhibitor of BCR-ABL, has preclinical activity against imatinib mesylate-resistant chronic myeloid leukemia (CML). Proceedings of the American Society of Hematology; 2004. Abstract 1983. Gorre ME, et al. Clinical resistance to STI-571 cancer therapy caused by BCR-ABL gene mutation or ampliÞcation. Science. 2001;293(5531):876–880. Guilhot F, et al. Follow-up of 1047 patients with chronic myelogenous leukemia treated with interferon or interferon with cytarabine. Blood. 1997;90(suppl 1):516A. Abstract 2299. [a] Guilhot F, et al. Interferon alpha-2b combined with cytarabine versus interferon alone in chronic myelogenous leukemia. New England Journal of Medicine. 1997;337:223–229. [b] Guilhot F, et al. Interferon therapy in chronic myelogenous leukemia. Hematology/Oncology Clinics of North America. 2004;18(3):585–603. [a] Guilhot F. Sustained durability of responses plus high rates of cytogenetic responses result in long-term beneÞt for newly diagnosed chronic-phase chronic myeloid leukemia (CML-CP) treated with imatinib (IM) therapy: Update from the IRIS study. Proceedings of the American Society of Hematology; 2004. Abstract 21. [b] Hahn EA, et al. Quality of life in patients with newly diagnosed chronic phase chronic myeloid leukemia on imatinib versus interferon alpha plus low-dose cytarabine: results from the IRIS Study. Journal of Clinical Oncology. 2003;21(11):2138–2146. Hasford J, et al. For the Collaborative CML Prognostic Factors Project Group: a new prognostic score for survival of patients with chronic myeloid leukemia treated with interferon-α. Journal of the National Cancer Institute. 1998;90(11):850–858. Hegedus T, et al. Interaction of tyrosine kinase inhibitors with the human multidrug transporter proteins, MDR1 and MRP1. Biochimica et Biophysica Acta. 2002;1587:318–325.

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Hehlmann R, et al. Randomized comparison of busulfan and hydroxyurea in chronic myelogenous leukemia: prolongation of survival by hydroxyurea. The German CML Study Group. Blood. 1993;82:398–407. Hehlmann R, et al. Interferon-α before allogeneic bone marrow transplantation in chronic myelogenous leukemia does not effect outcome adversely, provided it is discontinued at least 90 days before the procedure. Blood. 1999;94:3668–3677. Hehlmann R. Genetic randomization of allogeneic BMT vs drug treatment in chronic myelogenous leukemia: the German CML Study III. Blood. 2000;96(suppl 1):abstract 605. [a] Hehlmann R. Trial of IFN or STI-571 before proceeding to allografting for CML. Leukemia. 2000;14:1560–1562. [b] Heyssel R, et al. Leukemia in Hiroshima atomic bomb survivors. Blood. 1960;15:313–331. Ho VT, et al. The history and future of T-cell depletion as graft-versus-host disease prophylaxis for allogeneic hematopoietic stem cell transplantation. Blood. 2001;98(12):3192–3204. Hughes TP, et al. Frequency of major molecular responses to imatinib or interferon alpha plus cytarabine in newly diagnosed chronic myeloid leukemia. New England Journal of Medicine. 2003;349(15):1423–1432. Issa JP, et al. Hypomethylation dynamics following decitabine therapy in chronic myelogenous leukemia. Proceedings of the American Society of Hematology; 2004. Abstract 2956. Italian Cooperative Study Group on Chronic Myeloid Leukemia. Interferon-α-2b compared with conventional chemotherapy for the treatment of chronic myeloid leukemia. New England Journal of Medicine. 1994;330:820–825. Italian Cooperative Study Group on Chronic Myeloid Leukemia. Long-term follow-up of the Italian trial of interferon-α versus conventional chemotherapy in chronic myeloid leukemia. Blood. 1998;92:1541–1548. Kalaycio ME. Chronic myelogenous leukemia: the news you have and haven’t heard. Cleveland Clinic Journal of Medicine. 2001;68(11):913–925. Kantarjian H, et al. Hematologic and cytogenetic responses to imatinib mesylate in chronic myelogenous leukemia. New England Journal of Medicine. 2002;346(9):645–652. Erratum in: New England Journal of Medicine. 2002;346(24):1923. [a] Kantarjian H, et al. Imatinib mesylate for Philadelphia chromosome-positive, chronic phase myeloid leukemia after failure of interferon alpha: follow-up results. Clinical Cancer Research. 2002;8:2177–2187. [b] Kantarjian H, Cortes J. Testing the prognostic model of Marin et al in an independent chronic myelogenous leukemia study group. Leukemia. 2004;18:650. Kantarjian HM, et al. Characteristics of accelerated disease in chronic myelogenous leukemia. Cancer. 1988;61(7):1441–1446. Kantarjian HM, et al. Treatment of chronic myelogenous leukemia: current status and investigational options. Blood. 1996;87:3069–3081. Kantarjian HM, et al. Treatment of Philadelphia chromosome-positive early chronic phase chronic myelogenous leukemia with daily doses of interferon alpha and low-dose cytosine arabinoside. Journal of Clinical Oncology. 1999;17:284–292.

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Colorectal Cancer

ETIOLOGY AND PATHOPHYSIOLOGY Terminology The term colorectal denotes both the colon and the rectum. Cancers of the colon and rectum are collectively termed colorectal cancers, despite functional, epidemiological, and anatomic distinctions and differences in treatment. Cancers of the colon and rectum are often considered together because of the difÞculties in distinguishing cancers arising in the region where the colon ends and the rectum begins (anal cancer is excluded from this study). After examining the anatomy and pathophysiology of colorectal cancer (CRC), we will look closely at the various possible causes of this cancer, covering several genes and lifestyle risk factors that have been identiÞed. Anatomy The colon and the rectum form the large intestine, which is part of the gastrointestinal (GI) tract. The purpose of the GI tract is to break down food, absorb ßuid and nutrients into the blood to be used by the body, and remove the unusable portion as waste. Early in the digestive process, food moves from the stomach into the small intestine—a 20-foot-long, narrow tube. Most of the absorption of water and nutrients is carried out in the small intestine. Undigested food residues and substances that cannot be absorbed, such as vegetable Þber, move into the large intestine at the lower right side of the abdomen. The large intestine removes Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

439

440

COLORECTAL CANCER

Stomach Transverse colon Small intestine Ascending colon

Descending colon

Cecum

Sigmoid colon

Appendix Anus

Rectum

FIGURE 1. Colorectal anatomy.

water from the remainder, passing semisolid feces into the rectum to be expelled from the body through the anus. Anatomically, the large intestine is divided into seven areas: the cecum; the ascending, transverse, descending, and sigmoid colon; the rectum; and the anus (Figure 1). As waste moves through the large intestine, it passes through the cecum, up into the ascending colon, across the transverse colon, then down the left side of the abdomen to the descending and sigmoid colon, where it is stored in the rectum. The rectum is an eight-inch section of intestine at the end of the colon. The anus is the Þnal three inches of the colon through which feces are Þnally expelled. The sphincter (the ring of muscle at the anus) allows for controlled removal of feces from the GI tract during defecation, or bowel movement. Preserving sphincter patency is the goal of surgery for rectal cancer. The wall of the large intestine comprises four structural layers: •

• • •

The innermost layer is the mucosa, which is composed of three separate sublayers: epithelium, connective tissue, and muscle. The epithelium, which contains crypts (pits or depressions), is in immediate contact with the contents of the colon. The cells deep inside the crypts have a high proliferative index and are the originating site of most CRCs. The epithelial layer provides lubrication from mucus-secreting goblet cells, which aids the passage of feces and protects the tract. The connective and muscular layers of the gastric mucosa provide both a foundation for the epithelial cells and a blood supply. The submucosa is the second layer of the large intestine and contains more connective tissue, blood vessels, lymphatic glands, and nerves. The third layer comprises muscle that controls the contractions of the colon, enabling movement of digested food through the tract. The role of the fourth and outermost layer of the large intestine, the serosa, is structural.

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441

Lymph nodes and lymph vessels permeate all layers of the GI tract and form an essential barrier to entry of the many bacteria and other pathogens to which the GI tract is exposed. However, as with other cancers, the lymphatic system is also responsible for the dissemination of CRCs to the lymph nodes, and ultimately, to distant organs. Pathophysiology Approximately 70% of all CRCs occur in the sigmoid colon and rectum. The remainder occurs in decreasing frequency in the ascending colon, the transverse colon and splenic ßexure, and the descending colon. Because adenomatous polyps and early-stage CRC cancers are usually asymptomatic and therefore difÞcult to detect, colorectal carcinomas tend to develop until they are large enough to cause symptoms, before CRC is diagnosed; the prognosis is correspondingly worse. Thus, the need for earlier diagnosis of adenomatous polyps and early-stage cancers is clear. CRC adenocarcinomas initiate in the lumen of the GI tract, then slowly invade the deeper layers of the intestinal wall. The extent of tumor invasion into the bowel correlates with both the presence of lymph node metastases and, ultimately, patient survival. CRC may spread by direct invasion of adjacent tissue and metastasize via lymphatic and hematogenous routes. Carcinomas of the cecum and ascending colon are often polyploid and may become large and bulky before presentation because of the large circumference of the right colon. Bulky mass lesions occur elsewhere in the colon as well. In the distal colon and rectum, where the bowel circumference is smaller, tumors may involve the entire circumference of the bowel to produce an annular constricting (“napkin ring”) lesion, which may obstruct the lumen. Cancers occasionally have a ßatter appearance, spreading intramurally; this feature is common in patients with irritable bowel disease. As tumors expand, they outgrow their blood supply—undergoing necrosis—and ulcerate, a common feature of larger cancers. Histologically, 90–95% of CRC carcinomas are adenocarcinomas; the remaining 5–10% are squamous cell carcinomas, undifferentiated carcinomas, rectal carcinoid tumors, or rarely, sarcomas. The adenocarcinomas are further classiÞed by grade, which is based on the degree of tumor differentiation, reßected by structural and cytological features of the specimen. Grade 1 is the most differentiated, having well-formed tubules and the least nuclear polymorphism and mitoses; grade 3 is the least differentiated, with only occasional glandular structures; and grade 2 is intermediate between grades 1 and 3. Of all CRCs, 70–90% arise from adenomatous polyps, and 10–30% arise from sessile adenomas. The larger the polyp, the greater the potential for malignancy. Polyps less than or equal to 5 mm in diameter have a negligible malignant potential, whereas polyps larger than 2 cm in diameter have a 50% chance of becoming malignant over time (Winawer SJ, 1997). Signs and Symptoms. Symptoms begin to appear once the tumor is large enough to cause bowel obstruction. The symptoms vary according to the region

442

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TABLE 1. Variation in Colorectal Cancer Symptoms According to Tumor Location Region Right colon (cecum, ascending colon, left side of transverse colon)

Left colon (sigmoid colon, descending colon, right side of transverse colon) Rectum

Symptoms Abdominal pain. Anemia (resulting from chronic blood loss). Weakness. Weight loss. Constipation alternating with diarrhea. Abdominal pain. Nausea and vomiting. Change in bowel movements. Rectal fullness. Urgency. Bleeding. Tenesmus (straining at stool).

of the colon or rectum that is affected (Table 1). Pelvic pain occurs at the later stages of the disease and usually indicates local extension of the tumor to the pelvic nerves. Staging. Although researchers have developed many staging systems for CRC, only two are commonly used: the modiÞed (combined) Astler-Coller Duke’s system and the more detailed tumor, node, metastasis (TNM) system. The former system is based on Duke’s classiÞcation system, which places patients into one of three categories (stages A, B, and C), and the Astler-Coller modiÞed system, which divides patients into Þve subdivisions. In the Astler-Coller system, A is limited to the mucosa, B1 involves the muscularis propria but does not penetrate it, B2 penetrates the muscularis propria, and C1 and C2 are counterparts of B1 and B2 with nodal metastases. The combined system has added three more stages: B3 represents involvement of adjacent structures, C3 is B3 with nodal metastasis, and D signiÞes the presence of distant metastasis. More recently, the American Joint Committee on Cancer (AJCC) and the Union Internationale Centre le Cancer (UICC) introduced the TNM staging system (Table 2). This system places patients into one of four stages (stages I–IV). The TNM system was updated in 2003 and further subdivided stage III disease according to the number of positive regional lymph nodes. Table 3 compares the two main staging systems. Prognosis. As with most cancers, the stage of CRC at the time of diagnosis is the most important prognostic factor for determining long-term outcome. In patients diagnosed with stage I disease, Þve-year survival occurs in 85–90%; by contrast, in patients diagnosed with stage IV disease, 5% or less survive Þve years. Table 4 summarizes other factors associated with prognosis of CRC. Spread of Colorectal Cancer. The pathway of metastatic spread is based largely on the lymphatic and venous drainage patterns in the area of the primary

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443

TABLE 2. TNM Categories Used for the Classification of Colorectal Cancer TNM Category Tis T1 T2 T3 T4 N0 N1 N2 M0 M1

Description Carcinoma in situ Tumor invades submucosa Tumor invades muscularis propria Tumor invades through muscularis propria Tumor invades serosa, nodes, and adjacent organs No lymph node involvement One to three positive nodes More than three positive nodes No distant metastasis Distant metastasis

M = Metastasis (absence or presence of distant metastasis). N = Node (absence or presence and extent of regional lymph node involvement). T = Tumor (size, extent, or depth of penetration of primary tumor).

TABLE 3. Comparison of the Modified Astler-Coller Duke’s and AJCC/UICC Colorectal Cancer (TNM) Staging Systems AJCC/UICC Stage Stage I Stage II Stage III Stage IV

TNM Category

Modified Astler-Coller Duke’s Stage

T1, N0, M0 T2, N0, M0 T3, N0, M0 T4, N0, M0 Any T, N1–2, M0 Any T, Any N, M1

Population Distribution (%)a

Five-Year Survival (%)

A

11

85–90

B1–B2 B3 C1–C3 D

35

60–80

26 29

30–60 0–5

a Percentages do not add to 100 because of rounding.

AJCC = American Joint Committee on Cancer. T = Tumor; N = Node; M = Metastasis. UICC = Union Internationale Contre le Cancer.

tumor. The most common sites of metastatic disease are the liver, lungs, and peritoneum, although bone and brain metastasis also can occur: •

•

Hepatic metastases are present in two-thirds of the patients who die of CRC, 40% of whom have metastases in the liver only. Most of the venous return of the colon and rectum goes into the portal vein and then into the liver, which creates a high likelihood of depositing tumor cells from the portal venous system into the liver. A minority of liver metastases are amenable to surgical resection or targeted radiotherapy. Pulmonary metastases are associated with 15–20% of CRC deaths and usually occur in patients with distal rectal cancers because the rectum’s venous drainage is directly into the inferior vena cava, bypassing the liver.

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TABLE 4. Colorectal Cancer Prognostic Factors Prognostic Factor Blood vessel (venous) invasion Duration of symptoms Eosinophilic and mast-cell infiltration Gender Histopathological tumor grade Histopathological tumor type

Location of the primary tumor Lymphatic vessel invasion

Obstruction and perforation Pattern of lymph node reaction Perineurial invasion Perioperative blood transfusion Peritumoral lymphocytic infiltration Primary tumor configuration Rectal bleeding

•

Comment The invasion of extramural veins (in pericolonic or perirectal fat) is associated with a significantly worse prognosis. Asymptomatic CRC patients have a better five-year survival rate than symptomatic patients. The presence of four or more mast cells per 30 oil immersion fields has been found to correlate with lower overall survival and to be an independent prognostic factor. Many studies report a more favorable prognosis for females than males, as with other malignancies. Higher grades are associated with worse prognosis. Some CRC subtypes—such as mucinous or colloid carcinoma, signet ring-cell carcinoma, and undifferentiated carcinoma—have a worse prognosis than typical adenocarcinoma. Five-year survival for rectal cancer at or below the peritoneal reflection is lower than for those above the reflection and for colon cancer. The incidence of lymphatic vessel invasion increases with stage and grade, as well as presence and extent of lymph node metastasis. Significantly decreases survival. Sinus histiocytosis and paracortical immunoblastic activity in the lymph nodes draining the tumor reportedly correlate with improved survival. Associated with a higher rate of recurrence and reduced survival. Associated with an increased recurrence rate and may be associated with reduced overall survival. Marked infiltration in and around the tumor is associated with a better prognosis. Polypoid and exophytic cancers have a better prognosis than flat and ulcerated lesions. Hemorrhage or rectal bleeding associated with CRC is linked to better prognosis, possibly because it leads to earlier diagnosis and intervention.

Peritoneal seeding is associated with approximately 35% of CRC deaths. Patients with intraperitoneal CRC tumors are more likely to have this type of seeding pattern than are patients with extraperitoneal tumors.

Approximately 10–20% of primary colon cancers present with synchronous metastatic cancer. With extended surgical lymphadenectomy, cure may be possible in 10–15% of these patients. Patterns of Recurrent Disease. Eighty percent of recurrences take place within two years of treatment, and CRC rarely recurs after Þve years. Patients with rectal cancer are at greater risk of local recurrence than patients with colon

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cancer because of the inherent physiological limitations on the surgeon’s ability to obtain wide radial and distal margins when removing a tumor from the rectum. Patients with a recurrence most often have metastatic disease; locoregional failure (i.e., not distant metastases) occurs in only 20% of treated patients in the absence of distant spread. Patients whose primary tumors invade nearby organs are at high risk (70%) of recurrence. Because locoregional failure is an element of 70% of recurrences, a major objective of treating recurrent disease is to restrain disease progression and control the pain, bleeding, and incontinence that result from uncontrolled pelvic disease. Etiology The exact causes of CRC are unknown. However, genetic, experimental, and epidemiological studies suggest that CRC results from complex interactions between inherited susceptibility and environmental factors. CRC incidence increases with age, and the disease is uncommon in people younger than age 40 (Figure 2). Approximately 75% of all new cases of CRC are sporadic (no apparent evidence of having inherited the disorder); the remaining 25% occur in people who have one or more predisposing factors for the disease. This high-risk group includes people with a hereditary predisposition to CRC—including those with hereditary nonpolyposis colorectal cancer (HNPCC), familial adenomatous polyps (FAP), or a family history of CRC—as well as those with a history of inßammatory bowel disease (IBD). Colorectal Tumorigenesis Pathways. Researchers have identiÞed two distinct genetic mechanisms that lead to the development of CRC: chromosome instability (sometimes referred to as loss of heterozygosity [LOH]) and microsatellite instability (MSI) (Figure 3). Chromosome instability is characterized by allelic losses of tumor suppressor genes; MSI is characterized by genetic

FIGURE 2. Incidence of colon and rectal cancer in the United States by age, 2004.

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FIGURE 3. Postulated pathways for colorectal tumorigenesis.

instability at microsatellite loci (short, repetitive DNA sequence tracts scattered throughout the genome), probably resulting from mutations in DNA mismatch repair enzymes. Fifteen percent of CRCs, including nearly all cases of HNPCC, display an MSI phenotype; the remaining 85% display LOH (Ionov Y, 1993; Lengauer C, 1997). Tumors originating from the proximal colon represent a high percentage of MSI-positive lesions and may have a better prognosis (Gervaz P, 2003). Certain clinical and biological differences between MSI and LOH tumors may ultimately affect treatment choice and prognosis. MSI tumors express the tumor marker carcinoembryonic antigen (CEA) much less frequently than LOH tumors, and they are more likely to occur in people with a family history of CRC. MSI-positive tumors have a better stage-speciÞc prognosis; however, the way in which MSI status affects tumor response to chemotherapy is unclear. Retrospective clinical studies are inconclusive and have shown that MSI-positive tumors are both more sensitive and less sensitive to cytotoxic chemotherapy (Elsaleh H, 2000; Diep C, 2003), while preclinical experiments have shown that MSI-positive cells are resistant to a variety of chemotherapeutic agents (e.g., platinum agents, antimetabolites, topoisomerase I inhibitors) (Fink D, 1998). A study reported by Brueckl and colleagues (Brueckl WM, 2003) investigated the relationship between MSI, treatment response, and survival in patients with CRC who were undergoing Þrst-line treatment with a weekly 24-hour infusion of high-dose 5-ßuorouracil (5-FU) and folinic acid. Results of immunohistochemical analyses of tumor samples showed that patients with high MSI experienced a signiÞcantly better response to treatment (72% versus 41%) and improved survival (33 months versus 19 months) than did patients whose tumors displayed microsatellite stability.

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Many more ongoing prospective clinical trials are testing the prognostic signiÞcance of the MSI-positive phenotype and whether its presence can predict response to chemotherapy. If these trials produce conclusive data, MSI status will be used to help determine whether patients should receive adjuvant chemotherapy and whether they are candidates for more aggressive therapy. Genetic Changes. Researchers have identiÞed many genes that are mutated or abnormally expressed in CRC (Table 5). The major genetic changes associated with CRC are alterations in proto-oncogenes, loss of tumor suppressor activity, and abnormalities in DNA repair genes. Risk Factors. An inherited susceptibility or predisposition often becomes apparent when cancers of the same body site or organ occur. in multiple blood relatives. Up to 10% of CRCs are associated with an identiÞable hereditary predisposition (Figure 4). Hereditary Nonpolyposis Colon Cancer. HNPCC (also known as Lynch syndrome) is the most common hereditary CRC syndrome and is inherited in an autosomal dominant manner. For a Þrst-degree relative of an HNPCC-afßicted patient, the risk of developing CRC is 50%. In this form of the disease, which is responsible for approximately 6% of CRC incidence, a preponderance of tumors arise on the right side of the colon, and about two-thirds of the cancer is located proximal to the splenic ßexure. HNPCC syndrome is more difÞcult to detect than FAP 1% HNPCC

Other hereditary predisposition factors

IBD 1%

6%

17%

75%

Sporadic FAP = Familial adenomatous polyposis. HNPCC = Hereditary nonpolyposis colorectal cancer. IBD = Inflammatory bowel disease. FIGURE 4. Incidence of colorectal cancer by risk category.

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TABLE 5. Select Genes Reportedly Involved in Colorectal Cancer Genetic Alteration Proto-oncogenes K-ras (chromosome 12)

MYC (chromosome 12) Tumor suppressor genes APC (adenomatous polyposis coli; chromosome 5)

MCC (mutated in colorectal cancer; chromosome 5) p53 (chromosome 17)

Frequency (%) 40–60

<10 >60

60

75

DCC (deleted in colorectal cancer; chromosome 18)

30–70

TGF-β type II R (chromosome 3) SMAD4/DPC4 (chromosome 18)

25–30

NFI GAP (chromosome 17)

10–15

14

DNA repair genes hMSH2 (chromosome 2)

<10

hMSH3 (chromosome 5) hMSH6 (chromosome 2) hMLH1 (chromosome 3) hPMS1 (chromosome 2) hPMS2 (chromosome 7)

<10 <10 <10 <10 <10

Metastasis genes NM23

<10

Comments Altered expression contributes to abnormal proliferation and eventual carcinogenesis. Occurs at the intermediate stage of the adenoma to carcinoma sequence. Induces cell proliferation. Originally described in association with FAP. Evidence suggests that APC defects occur early in the development of CRC because the APC gene is deleted in the adenoma, before the development of the carcinoma. APC abnormalities may disrupt normal cell adhesion through altered association with molecules called catenins and the cellular adhesion molecule E-cadherin. Unknown molecular function and onset of occurrence. Occurs late in the adenoma to carcinoma sequence. May also obstruct normal cell-to-cell adhesive interactions. Patients whose tumors have deletions of the DCC gene have a worse prognosis than those whose gene is intact. Approximately 15% of MSI cancers have mutations in both alleles. Non-sense, frameshift, missense, and deletion mutations; mutations in one allele usually accompanied by LOH of remaining allele. Mutation leads to increased active GTP-bound ras-p21. Alterations in DNA repair genes lead to DNA replication errors and increased mutation. These genes appear to play a major role in HNPCC, but similar alterations are found in approximately 15% of sporadic cancers.

Occurs very late in the adenoma to carcinoma sequence.

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TABLE 5. (continued) Genetic Alteration

Frequency (%)

Carcinogen metabolism genes CYP1A1 <10

GSTM1

<10

Comments To date, CYP1A1 variants have been evaluated in only one study of in situ CRC (Sivaraman L, 1994). The risk of CRC was increased approximately eightfold with a specific CYP1A1 mutation. Several studies have shown that GSTM1 mutations or deletions may be associated with increased risk of CRC (Sadaat I, 2001; Zhong S, 1993).

FAP = Familial adenomatous polyposis. GTP = Guanine triphosphate. LOH = Loss of heterogeneity. MSI = Microsatellite instability. TGF = Transforming growth factor. Note: Full source citations appear in ‘‘References.’’

the less common FAP because it usually lacks a clinical hallmark. Researchers have identiÞed two patterns of this familial disease—Lynch syndrome types I and II. Lynch syndrome I is a site-speciÞc syndrome associated only with adenocarcinoma of the colon; its incidence is rare compared with that of Lynch syndrome II. Lynch syndrome II—also known as the cancer family syndrome—describes a generalized, heightened risk of developing adenocarcinomas of the colon, ovary, pancreas, breast, or bile duct as well as the urologic, endometrial, or gastric systems. Both Lynch syndrome I and Lynch syndrome II are characterized by the development of multiple CRCs at an early age; the average age of onset is 40–45, 20 years younger than average age of onset for the general population. Virtually all cases of HNPCC tumors exhibit the MSI phenotype, and evidence shows that, when matched by cancer stage, HNPCC patients beneÞt from better survival rates compared with spontaneous-CRC patients (J¨arvinen H, 2000). Studies have demonstrated the cancer-related role of genes that function in the repair of DNA damage. In HNPCC, an inherited mutation in one of the mismatch repair (MMR) genes appears to be a critical factor. MMR genes normally produce proteins that identify and correct sequence mismatches that can occur during DNA replication or via DNA-damaging agents (such as ultraviolet light). A mutation that inactivates an MMR gene leads to an accumulation of cell mutations that greatly increases the likelihood of malignant transformation and cancer. MMR can also play an important role in the repair of lesions caused by chemotherapeutic agents; reducing this role could account for the possible increase in sensitivity of colon and rectal cancers to chemotherapy (Lawes DA, 2003). Familial Adenomatous Polyposis and Gardner’s Syndrome. FAP and its variant, Gardner’s syndrome, are inherited in an autosomal dominant fashion,

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with inactivation of the adenomatous polyposis coli (APC ) gene. These syndromes have served as models for studying the sequence of events in which adenoma develops into carcinoma in the large bowel. FAP causes hundreds and even thousands of polyps to form in the colon and rectum during the second and third decades of life. Although individual polyps carry little risk of malignant transformation, large numbers of polyps increase the risk of CRC occurrence to almost 100% by age 40 (Midgley R, 1999). Attenuated FAP (AFAP) is typically associated with a far less dramatic proliferation of polyps (less than 100 cumulatively) and a later age of onset than FAP (closer to age 50 years). AFAP may be difÞcult to distinguish from HNPCC because of a preponderance of polyps in the proximal (on the right side) colon, which can be accurately diagnosed only through a colonoscopy. Up to 30% of FAP cases are estimated to be caused by de novo mutations, with no family history. In Gardner’s syndrome, extracolonic manifestations occur in conjunction with polyposis. The extracolonic manifestations include epidermoid cysts (cystic tumors containing epidermal or similar tissue); osteomas of the mandible, skull, and long bones; and Þbromas. Family History. A subset of families exhibit CRC clustering but do not meet the diagnostic criteria of known genetic predisposition; they make up the largest and most difÞcult group of patients whose disease must be managed. In such families, the incomplete penetrance (i.e., some known mutation carriers do not express disease) and variable disease expression (i.e., the same germline mutation can cause different clinical symptoms/phenotypes in patients from the same or unrelated families) associated with these cancer predisposition genes suggest that additional factors modify the risk of cancer. Rare Colorectal Cancer Predisposition Syndromes. Researchers have identiÞed several other rare inherited CRC predisposition syndromes. Peutz-Jeghers syndrome is associated with distinct polyps in the colon, stomach, and small bowel resulting from mutations of the STK11 gene (Jenne D, 1998). Juvenile polyposis coli is distinguished by noncancerous polyps and is caused by defects in the tumor suppressor genes PTEN (Olschwang S, 1998) and SMAD4/DPC4. Turcot’s syndrome is characterized by multiple colorectal polyps and brain tumors. Environment and Age. The risk of CRC is lower in Asia, Africa, and South America than in North America, Europe, and Australia. When people from low-risk regions immigrate to high-risk regions, their risk of developing CRC eventually increases to that of their adopted country. In addition, when a person adopts a “Western style” diet, the risk of CRC increases. Risk prevention should therefore be tailored toward the Þnding that environmental factors and lifestyle choices may play a greater role in cancer development than genetic factors (Lichtenstein P, 2000). Table 6 summarizes lifestyle factors linked to CRC development. As mentioned earlier, age is a signiÞcant risk factor for CRC: more than 90% of CRC mortality occurs in people aged 50 or older, and the highest incidence of CRC is in people older than 70 at diagnosis.

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TABLE 6. Lifestyle Factors Linked to the Development of Colorectal Cancer Factor

Comments

Alcohol

The role of alcohol consumption in CRC is unclear. One recent clinical study found that high alcohol consumption in combination with a low micronutrient diet (retinol, thiamine, carotene, vitamins A and C) may lead to an increased risk of CRC (Jedrychowski W, 2002). Calcium-rich diets are associated with a reduction in the risk of recurrent colorectal adenomas (Baron JA, 1999). Lack of exercise is linked to an increased risk of CRC (Skibber JM, 2001). Researchers widely accept that diets high in fat and cholesterol are linked to an increased risk of CRC (Skibber JM, 2001). Historically, studies have indicated that diets high in fiber may protect against CRC (Faivre J, 1998). Later studies, however, have failed to prove that a high-fiber diet reduces the risk of the disease (Alberts DS, 2000). Although certain processed meats or sausages or meats cooked at a very high temperature (forming hetrocyclic amines) may carry some risk, data suggest that the relationship between meat consumption and CRC is weaker than the ‘‘probable’’ status given by the WCRF in 1997 (Truswell AS, 2002). Researchers postulate that yellow-green cruciferous vegetables; vitamin A-, C-, and E-rich foods; selenium; and folic acid have a general chemoprotective effect (Burnstein MJ, 1993). The role of smoking in CRC is unclear. One study in women found a specific relationship between the risk of rectal cancer and duration of smoking (with a duration of greater than 30 years posing a significant risk). The study found little evidence for altered risk of colon cancer (Terry PD, 2002). Smoking in combination with Helicobacter pylori infection, however, may pose a significant increased risk of CRC (Brenner H, 2002).

Calcium Exercise Fat Fiber

Meat

Other dietary factors

Smoking

WCRF = World Cancer Research Fund. Note: Full source citations appear in ‘‘References.’’

Inflammatory Bowel Disease. Patients with IBD—particularly ulcerative colitis or Crohn’s disease—are at an increased risk (up to 30-fold) of developing CRC. As the duration and extent of IBD increase, the patient’s risk of eventually being diagnosed with CRC rises. Fortunately, because IBD is readily apparent, this subgroup of at-risk patients is usually identiÞed long before colon cancer develops. IBD is responsible for approximately 1% of CRC incidence. CURRENT THERAPIES Treatment of colorectal cancer (CRC) is closely linked to disease stage. Treatment modalities include surgery, radiotherapy, and chemotherapy. Surgical resection of the primary tumor and regional lymph nodes is the only curative treatment for CRC and may cure up to 50% of patients.

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Pharmaceutical therapies play an important adjunct and palliative role in the treatment of most cases of stages III and IV CRC. These pharmacological agents help reduce the incidence of recurrence, prolong survival, and improve quality of life for CRC patients. A growing body of evidence also supports the use of adjuvant chemotherapy in patients with stage II CRC who undergo curative resection but have a high risk of recurrence. For rectal cancer, neoadjuvant chemoradiation therapy is used in an attempt to downstage the tumor and reduce the need for radical surgery. Investigations of direct intrahepatic administration of chemotherapeutic agents for treating CRC liver metastases (via either the hepatic artery or the portal vein) to increase drug concentration locally have revealed no clinical beneÞt. Consequently, this approach has not been widely adopted outside experimental studies. For more than four decades, the mainstay of chemotherapy for CRC has been 5-ßuorouracil (5-FU; generics), and it continues to play a major role both as a single agent and as an agent combined with irinotecan (PÞzer’s Camptosar, PÞzer/Yakult/SanoÞ-Aventis’s Campto, Daiichi’s Topotecin) or oxaliplatin (SanoÞ-Aventis’s Eloxatin/Eloxatine, Yakult’s Elplat). 5-FU is commonly used as a single agent with radiotherapy in the neoadjuvant setting for rectal cancer, and it is combined with leucovorin (LV; generics) in various regimens in the adjuvant and metastatic settings. Clinicians use many different 5-FU/LV regimens, which differ in the doses and frequency of 5-FU administration and in the duration over which 5-FU is given. However, 5-FU is commonly administered either as a short injection (bolus regimen) or continually infused over 24 or more hours (infusional regimen). The half-life of 5-FU in plasma is very short, and potentially cytotoxic concentrations are maintained for only a few hours following a bolus injection. Some research suggests that bolus 5-FU may exert its effect through misincorporation into RNA (in place of uracil) rather than by inhibition of thymidylate synthase (Heidelberger C, 1957; Duschinsky R, 1957), 5-FU’s normal method of action. Capecitabine (Roche’s Xeloda)—an oral pro-drug of 5-FU—is used increasingly in CRC owing to its advantage over intravenous 5-FU. A prospective, integrated analysis of two large randomized Phase III trials in metastatic CRC (mCRC) demonstrated that single-agent capecitabine has signiÞcantly higher tumor response rates compared with 5-FU/LV (Mayo Clinic regimen): 26% versus 17% (Van Cutsem E, 2004). Several other 5-FU pro-drugs are available for treating CRC. Floxuridine (5-ßuorodeoxyuridine [FUDR]; generics) is available in the United States and is used almost exclusively for the treatment of liver metastases in hepatic arterial infusion protocols. Doxißuridine (DFUR; Roche’s Furtulon) is an oral agent; it is available only in Japan, where it is commonly used as an adjuvant therapy and as treatment for mCRC. Two agents with different mechanisms of action, oxaliplatin and irinotecan, are used in the treatment of mCRC, as already mentioned, and oxaliplatin recently gained approval in the United Kingdom for the adjuvant treatment of stage III colon cancer. In metastatic disease, either oxaliplatin or irinotecan combined with 5-FU/LV can be considered Þrst-line treatment (although irinotecan

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is increasingly being replaced by oxaliplatin in the Þrst-line setting). Oxaliplatin or irinotecan monotherapy appears less efÞcacious than using the compounds in combination with 5-FU/LV; the combination of different mechanisms of action results in a beneÞcial synergy. Two new agents—cetuximab (ImClone/Bristol-Myers Squibb/Merck KGaA’s Erbitux) and bevacizumab (Genentech/Roche’s Avastin)—recently gained approval for the treatment of advanced mCRC and are beginning to be incorporated in mCRC therapy. Table 7 outlines the chemotherapy regimens most commonly used to treat colorectal cancer. These regimens, as well as single-agent therapies and nonpharmacological approaches for treating CRC, are described in the following sections. Roswell Park Regimen Overview. The Roswell Park regimen comprises a high-bolus dose of 500–600 mg/m2 5−ßuorouracil (5-FU; generics) administered midway through a two-hour leucovorin (LV; generics) (Figure 5) infusion, repeated once a week for up to 48 weeks. This regimen was one of the Þrst to Þnd application in the adjuvant treatment of stage III CRC, and it conÞrmed the beneÞcial role of LV in therapy. Early studies compared the Roswell Park regimen (and the Mayo Clinic regimen, discussed next) with surgery alone. A clear beneÞt was demonstrated in patients with node-positive disease: the relapse rate decreased by approximately 30–40%, and absolute survival improved by 15–20%. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

5-FU is a ßuoropyrimidine antifolate. Antifolates exert their cytotoxic effect via several mechanisms. One of the most important targets for 5-FU is thymidylate synthase (TS), a key enzyme in de novo thymidine synthesis. This pyrimidine, which requires folate cofactors, is an essential requirement for normal DNA synthesis: the inhibition of TS inhibits DNA synthesis and results in cytotoxicity. Because 5-FU is a structural analogue of uracil, it is also misincorporated into RNA in place of uracil, interfering with normal RNA function and leading to cell death.

H2N

N

HN

H N N

O

NH CH2

NH

CHO

CH2

C

CH

O

COOH

FIGURE 5. Structure of leucovorin.

CH2

COOH

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TABLE 7. Current Regimens Used for Colorectal Cancer Regimen or Class Roswell Park

Mayo Clinic

de Gramont

IFL

Regimen Components Agent

Availability

Dose

Common Toxicities

5-FU (generics)

US, F, G, I, S, UK, J

5-FU: 600 mg/m2 bolus of LV at mid-infusion.

LV (generics)

US, F, G, I, S, UK, J

5-FU (generics)

US, F, G, I, S, UK, J

LV (generics)

US, F, G, I, S, UK, J

5-FU (generics)

US, F, G, I, S, UK, J

LV (generics)

US, F, G, I, S, UK, J

5-FU (generics)

US, F, G, I, S, UK, J

LV (generics)

US, F, G, I, S, UK, J US, F, G, I, S, UK, J

LV: 500 mg/m2 given over two hours. Repeat every 7 days for 48 weeks. 5-FU: 425 mg/m2 IV Leukopenia bolus. Gastrointestinal upset (diarrhea) Nausea and vomiting Stomatitis Alopecia LV: 20 mg/m2 per day for 5 days. Repeat every 4 weeks for 6 months. 5-FU: 400 mg/m2 IV Gastrointestinal bolus followed by upset (diarrhea) 600 mg/m2 IV Nausea infusion over 22 hours. Repeat on second day; repeat cycle after 2 weeks for 6–12 months. LV: 200–400 mg/m2 IV over 2 hours prior to 5-FU. Neutropenia 5-FU: 500 mg/m2. Gastrointestinal upset (diarrhea) Vomiting LV: 20 mg/m2.

Irinotecan (Pfizer’s Camptosar, Pfizer/Yakult/ Sanofi-Aventis’s Campto, Daiichi’s Topotecan)

Irinotecan: 125 mg/m2 weekly or 350 mg/m2 IV over 90 minutes every 3 weeks. Repeat every 6 weeks.

Leukopenia Gastrointestinal upset (diarrhea) Nausea and vomiting Stomatitis Alopecia

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TABLE 7. (continued) Regimen or Class FOLFIRI

Regimen Components Agent

Availability

Irinotecan (Pfizer’s US, F, G, I, Camptosar, S, UK, J Pfizer/Yakult/ Sanofi-Aventis’s Campto, Daiichi’s Topotecan)

Dose de Gramont (as detailed above) plus irinotecan: 180 mg/m2 IV over 90 minutes on day 1, repeated every 2 weeks.

Common Toxicities

Myelosuppression Gastrointestinal upset (diarrhea) Neurotoxicity Hypersensitivity Elevated liver enzymes Alopecia Rash Pain Pulmonary toxicity Cardiac toxicity Granulocytopenia FOLFOX4 Oxaliplatin US, F, G, I, de Gramont (as Neutropenia (Sanofi-Aventis’s S, UK, J detailed above) Neurotoxicity Eloxatin/ plus oxaliplatin: Eloxatine, 85 mg/m2 on day 1 Fever of the infusion, Gastrointestinal Yakult’s Elplat) repeated every 2 upset (diarrhea) weeks. Nausea and vomiting Acne-like rash Cetuximab/ Cetuximab (Imclone US, F, G, UK Cetuximab: Diarrhea irinotecan Systems/Merck 400 mg/m2 initial dose, followed by Neutropenia KGaA’s Erbitux) 250 mg/m2 weekly. Nausea & Vomiting Irinotecan (Pfizer’s US, F, G, I, Irinotecan: Doses Camptosar, S, UK, J vary: 125 mg/m2 weekly for 4 weeks Pfizer/Yakult/ followed by 2 Sanofi-Aventis’s weeks rest; Campto, Daiichi’s 180 mg/m2 every 2 Topotecan) weeks; or 350 mg/m2 every 3 weeks. Bevacizumab/ Bevacizumab US, F, G Bevacizumab: Diarrhea 5-FU/LV (Genentech/ 5 mg/kg every 2 Leukopenia Roche’s Avastin) weeks. Stomatitis Fever Headache Rash Epistaxis Chills Thrombosis 5-FU (generics) US, F, G, I, 5-FU: 500 mg/m2 IV bolus. S, UK, J LV (generics) US, F, G, I, LV: 20 mg/m2 IV. S, UK, J

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TABLE 7. (continued) Regimen or Class

Regimen Components Agent

Cetuximab, Cetuximab single agent (ImClone Systems/Merck KGaA’s Erbitux)

Availability

US, F, G, I, UK

Irinotecan, Irinotecan (Pfizer’s US, F, G, I, single agent Camptosar, S, UK, J Pfizer/Yakult/ Sanofi-Aventis’s Campto, Daiichi’s Topotecan)

Dose Once weekly for 4 weeks, repeat every 6 weeks. Cetuximab: 400 mg/m2 initial dose, followed by 250 mg/m2 weekly until disease progression or unacceptable toxicity. Irinotecan: 300–350 mg/m2 IV 90-minute infusion on day 1, repeat every 3 weeks.

Capecitabine, Capecitabine US, F, G, I, single agent (Roche’s Xeloda) S, UK, J

Capecitabine: 1.25 g/m2 twice daily for 14 days; subsequent courses repeat every 3 weeks.

Raltitrexed, Raltitrexed single agent (AstraZeneca’s Tomudex)

F, I, S, UK

Raltitrexed: 3 mg/m2 by IV infusion over 15 minutes.

Tegafur-uracil, Tegafur-uracil single agent (Bristol-Myers Squibb’s Uftoral/UFT, Otsuka’s UFT)

F, G, I, S, UK, J

Tegafur: 300 mg/m2 with uracil: 672 mg/m2 daily in 3 divided doses for 28 days; subsequent courses repeated after 7-day interval.

Common Toxicities

Acne-like rash

Leukopenia Neutropenia Gastrointestinal upset (diarrhea) Nausea and vomiting Alopecia Leukopenia Neutropenia Thrombocytopenia Gastrointestinal upset (diarrhea) Hand-and-foot syndrome Hyperbilirubinemia Myelosuppression Gastrointestinal upset (diarrhea) Transient elevation of liver enzymes Asthenia Myelosuppression Alopecia Gastrointestinal upset (diarrhea) Elevated liver enzymes Cardiac toxicity Renal toxicity

5-FU = 5-fluorouracil. CAPOX = Capecitabine/5-fluorouracil/leucovorin. FOLFIRI and IFL = Irinotecan/5-fluorouracil/leucovorin. FOLFOX = Oxaliplatin/5-fluorouracil/leucovorin. LV = Leucovorin. US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan.

CURRENT THERAPIES

•

457

LV is a biochemical modulator of 5-FU. LV enhances the activity of 5-FU by facilitating the binding of 5-FU to the enzyme thymidylate synthase, resulting in a greater degree of activity against cancer.

Clinical Performance. The pivotal clinical study that demonstrated the beneÞt of LV-modiÞed 5-FU as adjuvant therapy for CRC was reported in 1993 (Wolmark N, 1993). This study randomized 1,081 patients with Duke’s stage B and C carcinoma of the colon to receive either lomustine (generics), vincristine (generics), and 5-FU (a regimen previously used as a chemotherapeutic treatment for CRC) or the Roswell Park regimen of leucovorin and 5-FU. The mean duration on study was 47.6 months. A signiÞcant improvement in disease-free survival (73% versus 64% disease-free survival at three years) and overall survival (84% versus 77%) was demonstrated in favor of the Roswell Park regimen. The Roswell Park regimen also conferred a 30% reduction in risk of treatment failure and a 32% reduction in mortality risk compared with the lomustine/vincristine/5-FU regimen. The beneÞts of this regimen have also been demonstrated in other studies. More recently, the Roswell Park regimen has been combined with newer agents. The addition of oxaliplatin (administered over two hours intravenously on days 1 and 15) with a modiÞed Roswell Park regimen (LV administered over two hours intravenously, and 5-FU IV bolus administered on days 1, 8, and 15, every 28 days) proved safe and effective in the Þrst-line treatment of advanced CRC (Miyata Y, 2004). According to this Japanese Phase I/II study, the oxaliplatin/modiÞed Roswell Park regimen was well tolerated in 18 patients with advanced CRC, and its efÞcacy and safety appear similar to those of the oxaliplatin/infusional 5-FU/LV combination (FOLFOX). A previous U.S. singleinstitution Phase II study of 41 patients with mCRC also reached this conclusion (Hochster HS, 2003). Mayo Clinic Regimen Overview. The Mayo Clinic regimen comprises a bolus 425 mg/m2 dose of 5-ßuorouracil (5-FU; generics) and 20 mg/m2 leucovorin (LV; generics) daily for Þve days, repeated every four weeks for six months. Early studies compared the Mayo Clinic regimen (and Roswell Park regimen) with surgery alone. A clear beneÞt of chemotherapy was demonstrated among node-positive patients: the relapse rate decreased by approximately 30–40%, and absolute survival improved by 15–20% compared with no chemotherapy. This observation established the combination of 5-FU and LV in either the Mayo Clinic or Roswell Park regimen as the mainstay adjuvant therapy in CRC. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

5-FU (Figure 6) is a ßuoropyrimidine antifolate. Antifolates exert their cytotoxic effect via several mechanisms. One of the most important targets for

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FIGURE 6. Structure of 5-fluorouracil (5-FU).

•

5-FU is thymidylate synthase (TS), a key enzyme in de novo thymidine synthesis. This pyrimidine, which requires folate cofactors, is an essential requirement for normal DNA synthesis: the inhibition of TS inhibits DNA synthesis and results in cytotoxicity. Because 5-FU is a structural analogue of uracil, it is also misincorporated into RNA in place of uracil, interfering with normal RNA function and leading to cell death. LV (Figure 5) is a biochemical modulator of 5-FU. LV enhances the activity of 5-FU by facilitating the binding of 5-FU to the enzyme thymidylate synthase, resulting in a greater degree of activity against cancer.

Clinical Performance. Data reported in 1997 (O’Connell M, 1997) demonstrated the superiority of the Mayo regimen over surgery alone as an adjuvant therapy in high-risk stage II and stage III patients. Three hundred and seventeen patients with high-risk stage II or stage III colon cancer were randomly assigned to receive either 5-FU/LV using the Mayo Clinic regimen or no further treatment. The beneÞts were seen after only six cycles of treatment: patients who had received postoperative 5-FU/LV signiÞcantly improved in their timeto-relapse rates and survival rates. Stomatitis, diarrhea, and leukopenia were the predominant toxicities. The Mayo Clinic regimen has also been widely adopted for treating mCRC; in this indication, it has shown improved activity and clinical beneÞt compared with the Roswell Park regimen. Similar objective tumor response rates of approximately 30% were observed for both treatment regimens, but the Mayo Clinic regimen produced a median survival of 10.7 months, an advantage over the median of 9.3 months’ survival seen with the Roswell Park regimen. A report by Vincent and associates (Vincent M, 2002) shows that a dose reduction was required in approximately 43% of patients receiving the Mayo Clinic regimen because of the toxicities noted. Despite this observation, the Mayo Clinic regimen remains in use because of the practical advantage of bolus over infusional dosing of 5-FU. The Mayo Clinic regimen was adopted worldwide when it demonstrated superiority over other regimens in use in the 1990s. The Mayo Clinic regimen is used both as adjuvant therapy and for the treatment of mCRC because of its conÞrmed activity and ease of administration (compared with infusional regimens) and because it avoids venous access-related complications. Results of a recent European Organization for Research and Treatment of Cancer (EORTC) randomized Phase III trial show that neither 5-FU administered

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by a 24-hour infusion in combination with LV nor 24-hour infusion of 5-FU alone prolong survival relative to bolus 5-FU/LV (the Mayo Clinic regimen) (Kohne C-H, 2005[a]). The next section provides details of this study. de Gramont Regimen Overview. The de Gramont regimen incorporates both bolus and infusional 5-FU (generics) and is sometimes referred to as a hybrid 5-FU regimen: 400 mg/m2 bolus 5-FU is administered after a 200–400 mg/m2 2−hour infusion of LV (generics); bolus 5-FU is followed by a 22-hour infusion of 5-FU at 600 mg/m2 . These treatments are given for two consecutive days, and the cycle is repeated every two weeks for 6–12 months. The de Gramont regimen improves both response rate and overall survival compared with 5-FU alone. In some studies, the de Gramont regimen has shown a superior response rate and toxicity proÞle compared with the widely used Mayo Clinic regimen. However, the de Gramont regimen requires more intense patient management. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the overall regimen activity: •

•

5-FU (Figure 6) is a ßuoropyrimidine antifolate. Antifolates exert their cytotoxic effect via several mechanisms. One of the most important targets for 5-FU is thymidylate synthase (TS), a key enzyme in de novo thymidine synthesis. This pyrimidine, which requires folate cofactors, is an essential requirement for normal DNA synthesis: the inhibition of TS inhibits DNA synthesis and results in cytotoxicity. Because 5-FU is a structural analogue of uracil, it is also misincorporated into RNA in place of uracil, interfering with normal RNA function and leading to cell death. LV (Figure 5) is a biochemical modulator of 5-FU. LV enhances the activity of 5-FU by facilitating the binding of 5-FU to the enzyme thymidylate synthase, resulting in a greater degree of activity against cancer.

Clinical Performance. In a pivotal study that compared the activity of the de Gramont regimen with the Mayo Clinic regimen in mCRC, 348 patients with measurable lesions were evaluated for response. In the de Gramont regimen–treated arm (infusional 5-FU), a signiÞcant improvement in response rate (33% versus 15%) was observed compared with the Mayo Clinic regimen–treated group (bolus 5-FU). Median progression-free survival times also signiÞcantly improved in the de Gramont arm: 27.6 weeks versus 22 weeks. However, the superior response rate did not translate to a signiÞcant improvement in survival. The de Gramont regimen was better tolerated than the Mayo Clinic regimen, with signiÞcantly lower incidence of grades 3 and 4 diarrhea and lower incidence of grades 3 and 4 nausea/vomiting and neutropenia (de Gramont A, 1997; de Gramont A, 1998). Based on recent results from the EORTC randomized Phase III trial, neither 5-FU administered by a 24-hour infusion in combination with LV nor 24-hour infusion of 5-FU alone prolong survival relative to bolus 5-FU/LV (the Mayo

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Clinic regimen) (Kohne C-H, 2005[a]). A total of 497 patients were randomized to receive the Mayo Clinic regimen (167 patients): weekly high-dose 24-hour infusional 5-FU (166 patients), or weekly high-dose 24-hour infusional 5-FU in combination with LV (164 patients). After a median follow-up of more than three years, the response rates were 12%, 10%, and 17%, respectively, in the 409 patients who were evaluable for response, but the difference was not statistically signiÞcant. Progression-free survival, however, was signiÞcantly longer for infusional 5-FU/LV: 4 months, 4.1 months, and 5.6 months, respectively. Researchers observed no difference in median survival: 11.1 months for the Mayo Clinic regimen, 13 months for infusional 5-FU, and 13.7 months for infusional 5-FU/LV. An increased incidence of grade 3 and 4 diarrhea in the infusional 5-FU/LV arm was observed compared with the other two arms, whereas stomatitis and hematologic toxicity were more frequent in patients receiving the Mayo Clinic regimen. Single-agent infusional 5-FU seems to be as effective as the Mayo Clinic regimen but with less toxicity. These results indicate that including LV is effective for prolonging the progression-free survival of weekly high-dose 24-hour infusional 5-FU (Kohne C-H, 2005[a]). In addition, results of a large Þve-arm randomized trial involving 2,135 patients with inoperable advanced CRC were presented at the American Society of Clinical Oncology (ASCO) meeting in 2005 (Maughan T, 2005). These data suggest that for these patients, the combination of high-dose LV plus bolus 5-FU followed by 5-FU infusion for 46 hours, (the modiÞed de Gramont regimen) with oxaliplatin or irinotecan appears superior to sequential 5-FU/LV followed by irinotecan. The study concluded that Þrst-line treatment with 5-FU/LV in combination with irinotecan or oxaliplatin achieves higher overall response rates. Furthermore, recent data assessing the inclusion of oxaliplatin into the de Gramont regimen (the FOLFOX regimen, discussed later) demonstrate signiÞcant improvements in response rate, time-to-progression, and overall survival in mCRC and in the adjuvant setting. IFL Regimen Overview. The IFL regimen incorporates irinotecan (PÞzer’s Camptosar, PÞzer/Yakult/SanoÞ-Aventis’s Campto, Daiichi’s Topotecin) at 125 mg/m2 , bolus 5-FU (generics) at 500 mg/m2 , and LV (generics) at 20 mg/m2 . At this time, it is used primarily for the treatment of mCRC. The treatment is given weekly for four weeks, and the cycle is repeated every six weeks. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

5-FU (Figure 6) is a ßuoropyrimidine antifolate. Antifolates exert their cytotoxic effect via several mechanisms. One of the most important targets for 5-FU is thymidylate synthase (TS), a key enzyme in de novo thymidine synthesis. This pyrimidine, which requires folate cofactors, is an essential requirement for normal DNA synthesis: the inhibition of TS inhibits DNA

CURRENT THERAPIES

•

•

461

synthesis and results in cytotoxicity. Because 5-FU is a structural analogue of uracil, it is also misincorporated into RNA in place of uracil, interfering with normal RNA function and leading to cell death. LV (Figure 5) is a biochemical modulator of 5-FU. LV enhances the activity of 5-FU by facilitating the binding of 5-FU to the enzyme thymidylate synthase, resulting in a greater degree of activity against cancer. Irinotecan (Figure 7) is a topoisomerase I inhibitor. Topoisomerase I inhibitors act as potent inhibitors of topoisomerase I, a nuclear enzyme that causes reversible, single-strand breaks in DNA during mitosis. Topoisomerase I inhibitors bind to the topoisomerase I-DNA complex and prevent re-ligation of the DNA strand, resulting in cell death.

Clinical Performance. In irinotecan’s pivotal U.S. Phase III combination therapy trial (Saltz LB, 2000), 667 previously untreated patients with mCRC received irinotecan/bolus 5-FU/LV, 5-FU/LV, or irinotecan. The triple-combination treatment elicited conÞrmed response rates of 39%, compared with 21% for 5-FU/LV alone and 18% for irinotecan alone. Median survival favored the IFL combination: patients assigned to receive irinotecan/5-FU/LV had a median survival of 14.8 months compared with 12.6 months for patients assigned to receive 5-FU/LV, and 12 months for patients assigned to receive irinotecan alone. In addition, time-to-tumor progression was signiÞcantly prolonged with the combination (7 months versus 4.3 months). The toxicity proÞle also favored the IFL regimen, with the exception of the incidence of grades 3 and 4 diarrhea, which almost doubled to 23% compared with irinotecan alone. Similar results were reported from a European Phase III trial, in which researchers observed a signiÞcant increase in median survival: 17.4 months versus 14.1 months (Douillard JY, 2000[a]). The FDA approved irinotecan in 2000 as Þrst-line treatment when used in combination with 5-FU/LV. Combination therapy with bolus 5-FU is associated with more severe toxicity than infusional 5-FU, so the latter approach has been more widely adopted.

H3C O

O

O N

N N O N

HO H3C

FIGURE 7. Structure of irinotecan.

O

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Researchers investigated adding irinotecan to 5-FU/LV postsurgery in the treatment of stage III colon cancer in an attempt to improve outcomes as observed in advanced disease. Results of an intergroup adjuvant trial presented at the 2004 ASCO meeting found that adding irinotecan to the 5-FU/LV regimen does not improve survival compared with using 5-FU/LV alone in the treatment of postsurgery stage III colon cancer (Saltz LB, 2004[a]). The trial, which was halted prematurely because of increased toxicity and no additional clinical beneÞt in the irinotecan combination arm, included 1,264 patients with a median follow-up of 2.6 years. The addition of irinotecan to 5-FU/LV did not produce any further beneÞt over 5-FU/LV alone, and irinotecan/5-FU/LV was associated with significantly more severe side effects including neutropenia, febrile neutropenia, and treatment-related deaths, compared with 5-FU/LV alone. The study concluded that weekly irinotecan/bolus 5-FU/LV should not be used in the management of stage III colon cancer (Saltz LB, 2004[a]).

FOLFIRI Regimen Overview. The FOLFIRI regimen comprises the de Gramont regimen of 5-FU (generics) and LV (generics) plus irinotecan (PÞzer’s Camptosar, PÞzer/Yakult/SanoÞ-Aventis’s Campto, Daiichi’s Topotecin) at 180 mg/m2 given on the Þrst day of treatment and repeated every two weeks. The combination of these three agents has become the standard of care in many cancer centers as Þrst-line treatment for mCRC. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

•

•

5-FU (Figure 6) is a ßuoropyrimidine antifolate. Antifolates exert their cytotoxic effect via several mechanisms. One of the most important targets for 5-FU is thymidylate synthase (TS), a key enzyme in de novo thymidine synthesis. This pyrimidine, which requires folate cofactors, is an essential requirement for normal DNA synthesis: the inhibition of TS inhibits DNA synthesis and results in cytotoxicity. Because 5-FU is a structural analogue of uracil, it is also misincorporated into RNA in place of uracil, interfering with normal RNA function and leading to cell death. LV (Figure 5) is a biochemical modulator of 5-FU. LV enhances the activity of 5-FU by facilitating the binding of 5-FU to the enzyme thymidylate synthase, resulting in a greater degree of activity against cancer. Irinotecan (Figure 7) is a topoisomerase I inhibitor. Topoisomerase I inhibitors act as potent inhibitors of topoisomerase I, a nuclear enzyme that causes reversible single-strand breaks in DNA during mitosis. Topoisomerase I inhibitors bind to the topoisomerase I-DNA complex and prevent re-ligation of the DNA strand, resulting in cell death.

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Clinical Performance. A recent randomized EORTC Phase III trial comparing weekly high-dose infusional 5-FU/LV with or without irinotecan demonstrated that adding irinotecan to 5-FU/LV increases progression-free survival and response rate in patients with untreated mCRC compared with 5-FU/LV alone (Kohne C-H, 2005[b]). Four hundred and thirty patients were randomized to weekly high-dose infusional FU/LV (216 patients) or weekly high-dose infusional 5-FU/LV with irinotecan (214 patients). After a median follow-up of 28 months, patients in the irinotecan combination arm achieved a median progression-free survival of 8.5 months compared with 6.4 months in patients who received the infusional 5-FU/LV alone. Response rate and duration of response were also signiÞcantly higher in the group: 62% versus 34% and 10 months and 9 months, for those in the irinotecan combination arm. The addition of irinotecan to infusional 5-FU/LV, however, showed a nonsigniÞcant trend toward improved overall survival: 20 months for the irinotecan combination arm compared with 17 months for patients treated with infusional 5-FU/LV alone. Side effects in this trial were generally manageable, with grades 3 and 4 diarrhea occurring in 21% and 24% of patients and febrile neutropenia in 1% and 2% of patients receiving the infusional 5-FU/LV and infusional 5-FU/LV/irinotecan combination, respectively. This study supports the idea that infusional 5-FU, as in the FOLFIRI regimen, may be the optimal way to administer 5-FU when combined with either irinotecan or oxaliplatin because of the lower toxicity and high efÞcacy (Kohne C-H, 2005[b]). In addition, an Italian group (Gruppo Oncologico Dell’Italia Meridionale) recently found that the primary difference between FOLFIRI and FOLFOX4 is their toxicity proÞle. The group performed a randomized, multicenter Phase III trial comparing FOLFIRI with FOLFOX4 for the treatment of advanced CRC. This study found no statistically signiÞcant difference in overall response rate, time-to-progression, or overall survival between the two regimens (Colucci G, 2005). In the 336 chemotherapy-naive patients randomized in the trial (164 to FOLFIRI and 172 to FOLFOX4), both regimens were equally effective as Þrstline treatment; overall response rates were not statistically different (31% versus 34%). After a median follow-up of 31 months, the median time-to-progression, duration of response, and overall survival were similar in the two arms: 7 months versus 7 months, 9 months versus 10 months, and 14 months versus 15 months, respectively (Colucci G, 2005). The one-year survival rate was 55% for FOLFIRI and 62% for FOLFOX4. The toxicity proÞle was manageable in both regimens. A high incidence of grade 1 and 2 alopecia and gastrointestinal disturbances were found in the FOLFIRI arm, whereas grade 1 and 2 thrombocytopenia, neurotoxicity, and hypersensitivity reactions were more common among patients who received FOLFOX4. Researchers are also investigating the use of 5-FU/LV in combination with both irinotecan and oxaliplatin to learn whether additional clinical beneÞt will be achieved. Results of a Phase II Italian study of a 48-hour continuous infusion of 5-FU, and LV combined with both irinotecan and oxaliplatin (the FOLFOXIRI regimen) show promising activity and efÞcacy in the Þrst-line treatment of

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mCRC, and the regimen offers a manageable toxicity proÞle (Masi G, 2004). In the 32 patients with unresectable mCRC, median progression-free survival and median survival observed were 10.8 and 28.4 months, respectively, after a median follow-up of 18 months. FOLFOX Regimen Overview. The intravenously delivered, third-generation platinum complex oxaliplatin (SanoÞ-Aventis’s Eloxatin/Eloxatine, Yakult’s Elplat) is approved in the United States and Europe as Þrst- or second-line therapy in combination with 5-FU/LV for patients with mCRC as the FOLFOX regimen. Oxaliplatin is also approved for the adjuvant treatment of stage III colon cancer after complete resection of a primary tumor. In March 2005, oxaliplatin received approval in Japan for the treatment of advanced-stage or recurrent CRC (incurable and unresectable disease). Single-agent oxaliplatin has achieved response rates in mCRC of almost 10% in previously treated patients and 20–24% in chemotherapy-naive patients. The FOLFOX regimen incorporates oxaliplatin therapy with 5-FU and LV. Oxaliplatin is the Þrst platinum agent to have demonstrated meaningful activity in the treatment of CRC. To date, seven different FOLFOX regimens have been studied in an attempt to optimize dose levels and schedule and to account for the numerous 5-FU/LV regimens in current use; all are administered every two weeks. The oxaliplatin dose is either 85 mg/m2 (FOLFOX3 and 4), 100 mg/m2 (FOLFOX2 and 6), or 130 mg/m2 (FOLFOX1 [every two treatment cycles] and FOLFOX7). The 5-FU dose is given either by infusion only for 22–24 hours at 1.5–2.0 g/m2 (FOLFOX1, 2, and 3) or as a 400 mg/m2 bolus followed by infusion, 600 mg/m2 for 22 hours (FOLFOX4) or 2.4 g/m2 over 46 hours (FOLFOX6 and 7). LV is given at 200 mg/m2 (FOLFOX4), 400 mg/m2 (FOLFOX6 and 7), or 500 mg/m2 (FOLFOX1, 2, and 3), typically as a two-hour infusion. FOLFOX4 is equivalent to the de Gramont 5-FU/LV regimen plus oxaliplatin. Based on convincing efÞcacy data, FOLFOX4 is increasingly being used for the treatment of advanced disease. Some physicians in Europe also administer FOLFOX6 in the metastatic setting to compensate for the greater dose intensity of oxaliplatin. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

5-FU (Figure 6) is a ßuoropyrimidine antifolate. Antifolates exert their cytotoxic effect via several mechanisms. One of the most important targets for 5-FU is thymidylate synthase (TS), a key enzyme in de novo thymidine synthesis. This pyrimidine, which requires folate cofactors, is an essential requirement for normal DNA synthesis: the inhibition of TS inhibits DNA synthesis and results in cytotoxicity. Because 5-FU is a structural analogue of uracil, it is also misincorporated into RNA in place of uracil, interfering with normal RNA function and leading to cell death.

CURRENT THERAPIES

NH2

O

O

O

O

465

Pt NH2

FIGURE 8. Structure of oxaliplatin.

•

•

LV (Figure 5) is a biochemical modulator of 5-FU. LV enhances the activity of 5-FU by facilitating the binding of 5-FU to the enzyme thymidylate synthase, resulting in a greater degree of activity against cancer. Oxaliplatin (Figure 8) is a platinum agent. Platinum agents inhibit tumor-cell replication by creating intra- and interstrand DNA cross-links.

Clinical Performance. Updated results of a large randomized Phase III trial involving 795 previously untreated mCRC patients show signiÞcant improvements in response rate, time to progression, and overall survival with the FOLFOX regimen compared with both an irinotecan/bolus 5-FU/LV combination (the IFL regimen) and an irinotecan/oxaliplatin combination (Goldberg RM, 2004). Patients receiving FOLFOX experienced a signiÞcantly higher response rate and longer time-to-progression than patients receiving either of the other regimens: response rates were 45%, 31%, and 35% for FOLFOX, IFL, and oxaliplatin/irinotecan, respectively; median times to tumor progression were 8.7 months, 6.9 months, and 6.5 months. Patients treated with FOLFOX also had an improved median survival (19.5 months, compared with 15.0 months and 17.4 months, respectively, for the IFL regimen and the irinotecan/oxaliplatin combination). FOLFOX was found to be more active and better tolerated than both the IFL and irinotecan/oxaliplatin combinations. These data formed the basis of the FDA approval on January 2004 of oxaliplatin combined with infusional 5-FU/LV for the Þrst-line treatment of mCRC. Final data from the Multicenter International Study of Oxaliplatin/5-FU/LV in the Adjuvant Treatment of Colon Cancer (MOSAIC) trial comparing adjuvant FOLFOX4 with an established 5-FU/LV regimen (the de Gramont regimen) have been reported for the overall population (Andre T, 2004). After a median follow-up of 48.6 months, with 1,123 patients in each arm, updated efÞcacy data presented at the 2005 ASCO meeting revealed a statistically signiÞcant 24% reduction in risk of recurrence in the FOLFOX4 arm compared with the 5-FU/LV-alone arm. For stage III patients who received the FOLFOX4 regimen, disease-free survival at four years was 70% compared with 61% with 5-FU/LV alone. The overall survival rate was 84% in the FOLFOX4 arm and 82.7% in the 5-FU/LV arm (de Gramont A, 2005). Follow-up is ongoing for a minimum of Þve years. This pivotal Phase III MOSAIC study led to FDA approval in November 2004 of FOLFOX4 as an adjuvant chemotherapy option for patients with stage III colon cancer who have undergone complete resection of the primary tumor. In both Þrst- and second-line treatment of mCRC, infusional 5-FU/LV in combination with oxaliplatin (FOLFOX) proved more effective than 5-FU/LV alone

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(de Gramont A, 2000; Giacchetti S, 2000; Grothey A, 2002; Rothenberg ML, 2003). Although a large amount of data now support the use of infusional 5-FU in combination with other agents, recent Þndings also show that bolus 5-FU/LV can produce a high degree of efÞcacy when combined with agents such as oxaliplatin (Wolmark N, 2005). Data from a randomized Phase III trial presented at the 2005 ASCO meeting showed that the addition of oxaliplatin to weekly bolus 5-FU/LV (FLOX) signiÞcantly improves three-year, disease-free survival in stages II and III colon cancer patients, compared with bolus 5-FU/LV alone (Wolmark N, 2005). These results indicate that oxaliplatin’s beneÞt does not depend on the schedule of 5-FU administration. Data from this trial conÞrmed and extended the results of the MOSAIC trial discussed previously (Wolmark N, 2005). Oxaliplatin has also been studied in combination with 5-FU/LV for the neoadjuvant treatment of hepatic metastases, rendering a proportion of patients with previously unresectable liver tumors suitable for surgery by downstaging their tumors. In a clinical trial of 151 patients with liver metastases unsuitable for resection because of tumor size, number, or location (Giacchetti S, 1999), 77 patients (51%) receiving oxaliplatin became amenable to potentially curative hepatic surgery. The median survival in this group of patients was 48 months, compared with 15.5 months in the 74 patients unable to have surgery. The Þveyear survival rate for the patients undergoing surgery is estimated to be 50%; no Þve-year survivors in the patient group had not had surgery. Hepatic resection following neoadjuvant therapy with oxaliplatin/5-FU/LV appears to provide survival equivalent to that of patients with initially resectable metastases. Recent research has shown that the FOLFOX4 regimen can also be combined with gemcitabine (Eli Lilly’s Gemzar) for the treatment of refractory mCRC patients with liver involvement. In a Phase II trial, the treatment was well tolerated and achieved a response rate of approximately 69% of treated patients (Correale P, 2005). An ongoing EORTC Phase III trial is also comparing surgery for metastatic liver disease with and without neoadjuvant FOLFOX4 in patients with resectable CRC liver metastases. Interim safety data from the Þrst 346 patients demonstrated that preoperative chemotherapy with FOLFOX4 can be safely administered. Survival results are expected in 2006, not yet available at this time (Nordlinger B, 2005). Cetuximab/Irinotecan Regimens Cetuximab (ImClone/Bristol-Myers Squibb/Merck KGaA’s Erbitux) is a chimeric monoclonal antibody (MAb) that blocks the epidermal growth factor receptor (EGFR) and supposedly causes receptor internalization, thereby irreversibly blocking EGFR activation. Cetuximab was Þrst approved in Switzerland in December 2003 for the second-line treatment of advanced mCRC. In February 2004, cetuximab received approval in the United States for use in combination with irinotecan or alone (if patients cannot tolerate irinotecan). Cetuximab as a single agent is discussed in a later section. In June 2004, cetuximab in

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combination with irinotecan was granted approval in Europe for patients with EGFR-expressing mCRC after failure of irinotecan-based chemotherapy. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

•

Cetuximab is a recombinant, chimeric MAb that binds to the extracellular domain of the human EGFR, thereby blocking EGFR activation. Approximately 70% of CRC patients are EGFR-positive (Saltz LB, 2001). The precise role of EGFR inhibition is not fully understood because some EGFR inhibitors are active in EGFR-negative patients as well as in EGFR-positive patients. Irinotecan (Figure 7) is a topoisomerase I inhibitor. Topoisomerase I inhibitors act as potent inhibitors of topoisomerase I, a nuclear enzyme that causes reversible single-strand breaks in DNA during mitosis. Topoisomerase I inhibitors bind to the topoisomerase I-DNA complex and prevent religation of the DNA strand, resulting in cell death.

Clinical Performance. The U.S. and European approval of cetuximab, either alone or in combination with irinotecan, was based primarily on the results of a multicenter, randomized, controlled clinical trial conducted in 329 patients who had progressed after treatment with irinotecan-based chemotherapy (Cunningham D, 2003). Supporting data were derived from an open-label, single-arm trial (138 patients) of cetuximab in combination with irinotecan and an open-label, singlearm trial (57 patients) of cetuximab as a single agent (discussed later under “Cetuximab, Single Agent”). The multicenter, randomized, controlled Phase II trial presented at the 2003 ASCO meeting found that patients treated with the combination regimen had a signiÞcantly higher response rate and longer median time to disease progression than patients treated with single-agent cetuximab (Cunningham D, 2003). The trial randomized 329 patients to receive either cetuximab/irinotecan (218 patients) or cetuximab alone (111 patients), with the option to switch to the O CH3 HN

O F

N O H3C

HO

O

N

OH

FIGURE 9. Structure of capecitabine.

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cetuximab/irinotecan combination after failure with cetuximab as a single agent. Cetuximab was administered as a 400 mg/m2 initial dose, followed by 250 mg/m2 weekly in both arms of the study until disease progression or the occurrence of unacceptable toxicity. Patients in the combination arm received irinotecan at the same dose and schedule at which they had been progressing. The primary endpoint was to determine the objective response rates of the cetuximab/irinotecan combination and cetuximab as a single agent in patients with EGFR-positive, irinotecan-refractory mCRC. Secondary end points included time to progression, duration of response, overall survival time, and the incidence of adverse effects. All patients entered into the study were evaluable for efÞcacy. According to an independent review committee assessment, the overall response was 22.9% in the combination arm, median duration of response was 5.7 months, and median time to progression was 4.1 months; the overall response rate in the cetuximab-alone arm was 10.8%, median duration of response was 4.2 months, and median time to progression was 1.5 months (Cunningham D, 2004). Median survival times were 8.6 months in the combination arm and 6.9 months in the single-agent arm. In the combination arm, the overall survival rates at 6 and 12 months were 66% and 29%, respectively. In the cetuximabalone arm, the overall survival rates at 6 and 12 months were 58% and 32%, respectively. An acne-like rash, attributed to cetuximab, occurred in approximately 80% of patients in each treatment arm, and grade 3 or 4 toxic effects on the skin were observed in 9.4% and 5.2% in the combination and cetuximabalone groups, respectively. Diarrhea and neutropenia were more frequent in the combination arm, but their frequency was similar to the known safety proÞle of irinotecan alone. No treatment-related deaths were reported. In this trial, cetuximab-based therapy was similarly effective among patients who had previously received oxaliplatin in addition to irinotecan. The number of previous treatment regimens and the prior use of oxaliplatin did not affect the efÞcacy of cetuximab, either alone or in combination with irinotecan. Researchers concluded that cetuximab has clinically signiÞcant activity when given alone or in combination with irinotecan to mCRC patients refractory to irinotecan-based chemotherapy (Cunningham D, 2004). Cetuximab was also studied in combination with irinotecan in an open-label, single-arm trial involving 138 patients with EGFR-positive mCRC that was refractory to irinotecan-based chemotherapy. Patients received a 20 mg test dose of cetuximab on day 1, followed by a 400 mg/m2 initial dose, and 250 mg/m2 weekly until disease progression or unacceptable toxicity occurred. Patients received the same dosing and schedule for irinotecan on which they had previously failed. The overall response rate was 15% for all patients and 12% for the irinotecan-failed patients. The median durations of response were 6.5 and 6.7 months, respectively (Saltz LB, 2002; Saltz L, 2004[b]). Researchers are also investigating cetuximab in the Þrst-line setting for mCRC. A Phase II European trial evaluated the safety and efÞcacy of cetuximab in combination with 5-FU continuous infusion/LV/irinotecan (the FOLFIRI regimen) as Þrst-line treatment in 23 mCRC patients expressing EGFR (Van Laethem JL,

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2003). Updated trial results were presented at the 2004 ASCO meeting. Of the 22 patients evaluable for efÞcacy, 10 patients achieved a partial response, 9 patients had stable disease, and 3 patients had progressive disease (Rougier P, 2004). The median time to progression was 10.9 months. A further 29 patients were enrolled in the trial to allow researchers to better assess safety and efÞcacy for a follow-on Phase III trial. Of the 42 patients evaluable for safety, the most frequent grade 3 and 4 adverse events were diarrhea (14.3%), neutropenia (11.9%), rash (11.9%), and nausea and vomiting (9.5%). Grade 1 and 2 acne-like rash were more frequent than grade 3 and 4. Researchers concluded that the combination of cetuximab and FOLFIRI is effective, safe, and feasible in patients with EGFR-expressing mCRC. Recent results of an international Phase II trial of cetuximab in combination with FOLFOX4 reported at the 2005 ASCO meeting showed that the regimen has an acceptable safety proÞle with demonstrable advantages in response rate in the Þrst-line treatment of patients with nonresectable, EGFR-positive mCRC who have not received any prior chemotherapy (Diaz Rubio E, 2005). Of the 43 patients enrolled in the study, all were evaluable for efÞcacy and safety. The overall response rate was 72% (conÞrmed responses), including 9% complete responses and 63% partial responses. Twenty-three percent of patients achieved stable disease and 8 patients are still receiving treatment. Progression-free survival at the time of presentation was 10.2 months. The major adverse events included grade 3 and 4 acne-like rash (30.2%), diarrhea (25.6%), neurotoxicity (25.6%), neutropenia (20.9%), and stomatitis/mucositis (16.3%). A total of ten patients have so far undergone surgery for their metastases (Diaz Rubio E, 2005). A Phase III trial has evaluated the FOLFOX4 regimen with or without cetuximab in EGFR-positive mCRC patients previously treated with irinotecan. (Badarinath S, 2004). Safety of cetuximab treatment has been assessed in the Þrst 38 patients. Grades 3 and 4 toxicities included asthenia (8%), diarrhea (5%), nausea (3%), vomiting (5%), thromboembolism (3%), and skin toxicity (3%) (Badarinath S, 2004). Although the trial was designed to enroll 1,100 patients, accrual has stopped owing to recent changes in clinical practice resulting in the replacement of irinotecan with oxaliplatin in the Þrst-line setting (Jennis A, 2005). A total of 43 patients were treated with cetuximab followed by FOLFOX4, and 47 patients were treated with FOLFOX4 alone. According to interim data for the patients treated with FOLFOX4 plus cetuximab, 9 had a partial response, 20 patients had stable disease, and 6 patients had progressive disease. At this time, it is too early to determine a response in the remaining 8 patients. Of the 42 patients evaluable in the FOLFOX4 group, 4 had partial response, 26 had stable disease, and 9 had progressive disease. It is also too early to determine a response in the remaining 3 patients. The combination of FOLFOX4 and cetuximab is effective in patients with mCRC previously treated with irinotecan (Jennis A, 2005). Bevacizumab/Fluorouracil/Leucovorin Overview. Bevacizumab (Genentech/Roche’s Avastin) is a recombinant, humanized vascular endothelial growth factor (VEGF) MAb. Bevacizumab used

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in combination with intravenous 5-FU–based chemotherapy gained U.S. approval in early 2004 for the Þrst-line treatment of patients with mCRC. In early 2005, bevacizumab launched in several European countries, including Germany and the United Kingdom, for the Þrst-line treatment of patients with mCRC. Bevacizumab is being investigated in several different settings in CRC and in various combinations: •

• • •

In mCRC, in combination with capecitabine, oxaliplatin and capecitabine (CAPOX regimen), and with cetuximab, irinotecan, and erlotinib (OSI/ Genentech/Roche’s Tarceva); In the adjuvant setting, in combination with oxaliplatin, 5-FU (generics), and LV (FOLFOX4); As a neoadjuvant treatment for patients with unresectable liver metastases; and In combination with radiotherapy or chemoradiotherapy for patients with primary rectal cancer.

Bevacizumab has been shown to improve survival in several studies when combined with chemotherapy in Þrst-line therapy for mCRC. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

•

•

Bevacizumab is an inhibitor of VEGF. Bevacizumab binds to VEGF and prevents the interaction of VEGF with its receptors on the surface of endothelial cells. VEGF, a multifunctional cytokine and potent permeability factor secreted in response to hypoxia, has a major angiogenesis-promoting effect. 5-FU (Figure 6) is a ßuoropyrimidine antifolate. Antifolates exert their cytotoxic effect via several mechanisms. One of the most important targets for 5-FU is thymidylate synthase (TS), a key enzyme in de novo thymidine synthesis. This pyrimidine, which requires folate cofactors, is an essential requirement for normal DNA synthesis: the inhibition of TS inhibits DNA synthesis and results in cytotoxicity. Because 5-FU is a structural analogue of uracil, it is also misincorporated into RNA in place of uracil, interfering with normal RNA function and leading to cell death. LV (Figure 5) is a biochemical modulator of 5-FU. LV enhances the activity of 5-FU by facilitating the binding of 5-FU to the enzyme thymidylate synthase, resulting in a greater degree of activity against cancer.

Clinical Performance. The efÞcacy and safety of bevacizumab were studied in two randomized, controlled clinical trials in combination with IV 5-FU–based chemotherapy. These trials, which were the basis of bevacizumab’s approval, are discussed in the following paragraphs. In the Þrst trial, a Phase III multicenter, randomized, double-blind trial of 813 patients with previously untreated mCRC, 402 patients were randomized to

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receive irinotecan, bolus 5-FU, and LV (the IFL regimen) with bevacizumab; 411 patients were to receive IFL plus placebo (Hurwitz H, 2004). The bolus-IFL regimen comprises irinotecan 125 mg/m2 IV, 5-FU 500 mg/m2 IV, and LV 20 mg/m2 IV given once weekly for four weeks of every six-week cycle; the bevacizumab dose was 5 mg/kg every two weeks. The primary endpoint was overall survival; secondary end points included progression-free survival, response rate, duration of response, safety, and quality of life. The IFL plus bevacizumab regimen increased the median overall survival of patients from 15.6 to 20.3 months, and the median duration of progressionfree survival from 6.2 months to 10.6 months compared with IFL alone. The one-year survival rate improved from 63.4% for the IFL-alone group to 74.3% when bevacizumab was added to IFL. Overall response rates for patients who had bevacizumab added to the IFL regimen improved from 35% to 45%, and duration of response increased from 7.1 months to 10.4 months. The combination was well tolerated; grade 3 hypertension was the only adverse event that signiÞcantly increased in the IFL plus bevacizumab arm, occurring in 11% of patients versus 2.3% in the IFL alone group (Hurwitz H, 2004). The aforementioned study initially contained a third treatment arm (bevacizumab in combination with 5-FU/LV), undertaken because no safety data existed on the IFL/bevacizumab combination at the time the trial was designed. Once analysis revealed an acceptable safety proÞle for the IFL/bevacizumab combination, the 5-FU/LV and bevacizumab combination was discontinued to allow full enrollment in the two main arms (IFL and IFL with bevacizumab). Results of this third group consisting of 110 patients show that bevacizumab in combination with the 5-FU/LV regimen is active and generally well tolerated in the Þrst-line treatment of patients with mCRC. The median overall survival was 18.3 months, median progression-free survival was 8.8 months, overall response rate was 40%, and median duration of response was 8.5 months (Hurwitz H, 2005). The toxicity proÞle observed was consistent with the expected toxicities for bolus 5-FU/LV alone and for bevacizumab. These further data demonstrate that the combination of 5-FU/LV with bevacizumab without irinotecan is an active regimen and is as effective as irinotecan with 5-FU/LV for the Þrst-line treatment of mCRC. This regimen is a potentially useful therapeutic option for patients who cannot tolerate more-aggressive chemotherapy (Hurwitz H, 2005). A second randomized, controlled Phase II trial evaluated two doses of bevacizumab in combination with 5-FU/LV versus 5-FU/LV alone as Þrst-line treatment of mCRC (Kabbinavar F, 2003). A total of 104 previously untreated patients were randomized to three treatment arms: 36 to 5-FU/LV (5-FU 500 mg/m2 , leucovorin 500 mg/m2 weekly for six weeks of each eight-week cycle), 35 to 5-FU/LV plus bevacizumab (5 mg/kg every two weeks), and 33 to 5-FU/LV plus bevacizumab (10 mg/kg every two weeks). 5-FU/LV was administered according to the Roswell Park regimen, and patients were treated until disease progression. The primary end points of the trial were objective response rate and progression-free survival. Treatment with bevacizumab (at both dose levels) plus 5-FU/LV resulted in higher response rates compared with FU/LV alone:

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the 5-FU/LV arm had a 17% response rate; the low-dose arm, a 40% response rate; and the high-dose arm, a 24% response rate. The median survival was longer in the bevacizumab combination arms: 13.8 months in patients receiving 5-FU/LV, 21.5 months in the low-dose bevacizumab arm, and 16.1 months in the high-dose bevacizumab arm. The median time to disease progression was longer when bevacizumab was added to 5-FU/LV compared with 5-FU/LV alone: 5-FU/LV, 5.2 months; low-dose arm, 9.0 months; high-dose arm, 7.2 months. After crossover from the 5-FU/LV arm, 2 of 22 patients had a partial response to bevacizumab alone at 10 mg/kg and 7 patients had stable disease. The median duration of bevacizumab therapy was two months, as was the median time to progression. Researchers reported 50 deaths in the study. The incidence and severity of 5-FU/LV’s side effects (diarrhea, leukopenia, and stomatitis) did not increase when bevacizumab was added to the regimen. Fever, headache, rash, epistaxis, and chills were associated with bevacizumab therapy but were generally mild to moderate in severity; proteinuria and hypertension were manageable. In the bevacizumab arms, an increased incidence of bleeding, hypertension, and thrombosis were reported. Thrombosis was the most signiÞcant adverse event, occurring more frequently with bevacizumab than with chemotherapy alone, and it was fatal in one patient, resulting in bevacizumab being discontinued in three additional patients. These preliminary results suggest that bevacizumab, in combination with 5-FU/LV, increases response rate, prolongs time to progression, and prolongs survival compared with 5-FU/LV alone in patients with mCRC (Kabbinavar F, 2003). Bevacizumab has also been evaluated in combination with cetuximab and irinotecan in irinotecan-refractory CRC patients. Interim data from a Phase II trial comparing bevacizumab/cetuximab/irinotecan with bevacizumab/cetuximab in 74 irinotecan-refractory, bevacizumab-naive CRC patients show that adding bevacizumab increases the efÞcacy of cetuximab and cetuximab/irinotecan therapies (Saltz LB, 2005). Of the 35 patients who received bevacizumab/cetuximab, 8 had a partial response, with a median time to progression of 6.9 months. Of the 39 patients who received bevacizumab/cetuximab/irinotecan, 5 had a partial response, with a median time to progression of 8.5 months. The incidence of irinotecan-related grades 3 and 4 toxicities included neutropenia in 7 patients, diarrhea in 10 patients, and fatigue in 4 patients. In the bevacizumab/cetuximab/irinotecan arm, 7 patients experienced grade 3 acne-like rash related to cetuximab exposure. Similarly, 6 patients in the bevacizumab/cetuximab arm experienced grade 3 acne-like rash (Saltz LB, 2005). Researchers concluded that concurrent administration of bevacizumab and cetuximab is feasible and that it does not produce any unexpected toxicities. Bevacizumab has been studied in other Phase III trials. A Phase III Eastern Cooperative Oncology Group (ECOG) trial presented at the 2005 ASCO meeting found that high-dose bevacizumab (10 mg/kg IV every two weeks) combined with FOLFOX4 (oxaliplatin/5-FU/LV) is well tolerated and improves overall survival and progression-free survival in advanced-CRC patients previously treated with a 5-FU and irinotecan–based regimen (Giantonio BJ, 2005[a]). A total 828

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patients were randomized to receive bevacizumab alone (243 patients), FOLFOX4 alone (289 patients), or bevacizumab plus FOLFOX4 (290 patients). The end points were overall survival, progression-free survival, and response rate. The median overall survival was 12.9 months (bevacizumab/FOLFOX4), 10.8 months (FOLFOX4), and 10.2 months (bevacizumab alone). The median progression-free survival was 7.2 months, 4.8 months, and 2.7 months respectively (Giantonio BJ, 2005[a]). Data from the two main arms—FOLFOX4 alone or bevacizumab/ FOLFOX4—show that the median overall survival for patients in the bevacizumab/FOLFOX4 arm improved by 16% compared with FOLFOX4 alone. Patients who received bevacizumab in combination with FOLFOX4 had a signiÞcantly higher incidence of grade 3 or 4 hypertension (6% versus 3%) and sensory neuropathy (16% versus 10%) compared with patients who received FOLFOX4 alone (Giantonio BJ, 2005[a]; Giantonio BJ, 2005[b]). The incidence of grade 3 or 4 hemorrhage, nausea, and vomiting was also higher in the bevacizumab/FOLFOX4 arm. Nonetheless, researchers concluded that highdose bevacizumab in combination with FOLFOX4 is well tolerated and increases survival in advanced CRC patients previously treated with a 5-FU and irinotecanbased regimen. Results from another Phase III trial (Kabbinavar FF, 2005) demonstrate that bevacizumab provides signiÞcant clinical beneÞt when combined with Þrstline chemotherapy for the treatment of mCRC. A total of 209 previously untreated mCRC patients who were not candidates for Þrst-line irinotecan-based therapy were randomized to receive either 5-FU/LV/placebo (105 patients) or 5-FU/LV/bevacizumab (104 patients). 5-FU/LV was administered according to the Roswell Park regimen. The primary endpoint was overall survival. The median overall survival was found to be longer in the 5-FU/LV/bevacizumab group (16.6 months) than in the FU/LV/placebo group (12.9 months). Median progression-free survival was 9.2 months for the 5-FU/LV/bevacizumab group and 5.5 months for the 5-FU/LV/placebo group. Bevacizumab in addition to 5-FU/LV was also associated with increases in the response rate (26.0% versus 15.2%) and median duration of response (9.2 versus 6.8 months). Grade 3 hypertension was more common with bevacizumab treatment (16% versus 3%) but was managed with oral medication and did not cause treatment discontinuation (Kabbinavar FF, 2005). Many trials have identiÞed hemorrhage, thromboembolism, proteinuria, and hypertension as possible bevacizumab-associated toxicities, but this recent Phase III trial found no increases in venous thrombosis, grade 3 bleeding, or clinically signiÞcant (grade 3) proteinuria. A total of 2,632 patients will be accrued within 2.5 years into a National Cancer Institute adjuvant chemotherapy Phase III trial of 5-FU/LV/oxaliplatin with or without bevacizumab in patients with resected stage II or III colon cancer. The study plans to follow up patients every 3 months for 2 years, every 6 months for 3 years, and then annually thereafter. The primary endpoint is disease-free survival, and the secondary endpoint is overall survival. Preclinical data have

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already shown bevacizumab’s promising activity in early-stage tumors (Gerber H-P, 2005). Cetuximab, Single Agent Overview. Cetuximab as a single agent is indicated for the treatment of EGFRpositive mCRC in patients who cannot tolerate irinotecan-based chemotherapy. Mechanism of Action. Cetuximab is a recombinant, chimeric MAb that binds to the extracellular domain of the human EGFR, thereby blocking EGFR activation. Approximately 70% of CRC patients are EGFR-positive (Saltz LB, 2001). The precise role of EGFR inhibition is not fully understood because some EGFR inhibitors are active in EGFR-negative patients as well as in EGFR-positive patients. Clinical Performance. At the ASCO meeting in 2004, researchers reported on results of a Phase II trial of 329 patients (see “Cetuximab/Irinotecan Regimen,” above, for details). The study found that cetuximab has clinically signiÞcant activity when given alone or in combination with irinotecan in mCRC patients who are refractory to irinotecan-based chemotherapy (Cunningham D, 2004). Single-agent cetuximab has demonstrated a good side-effect proÞle and may therefore be an option for patients who are not considered candidates for further treatment with irinotecan-based chemotherapy or who do not want to receive such treatment (Cunningham D, 2004). In a multicenter, open-label, single-arm clinical trial, cetuximab was evaluated as a single agent in 57 patients (median age 56) with EGFR-expressing mCRC whose disease progressed following irinotecan-based chemotherapy (Saltz LB, 2004[b]). Patients received a 20 mg test dose of cetuximab on day 1, followed by a 400 mg/m2 initial dose, and 250 mg/m2 weekly dose until toxicity necessitated interruptions. The primary endpoint was to determine the response rate. Secondary endpoints included duration of response, time to progression, survival duration, and toxicity. According to an independent response assessment committee, 5 patients (9%) had a partial response rate (Saltz LB, 2004[b]). Twenty-one additional patients had stable disease and minor responses. The median time to progression was 1.4 months, with a median duration of response of 4.2 months. The median survival for all patients was 6.4 months. The most commonly observed grade 3 and 4 adverse events were an acne-like skin rash, which is characteristic of cetuximab toxicity (86% with any grade, 18% with grade 3), and a composite of asthenia, fatigue, malaise, or lethargy (56% with any grade, 9% with grade 3). Two patients (3.5%) experienced grade 3 and 4 allergic reactions, leading them to discontinue therapy. Researchers found that patients with skin rash of any grade had superior survival compared with patients who did not experience skin rash, with a trend toward improved survival correlating with increasing grade of rash (Saltz LB, 2004[b]). Results of cetuximab therapy in patients with EGFR-positive mCRC presented at the 2005 ASCO meeting found the drug to be well tolerated among heavily

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pretreated patients—those refractory to a ßuoropyrimidine, irinotecan, and oxaliplatin (Lenz H, 2005). A total of 346 patients were enrolled and treated in the study. According to an independent review committee, an objective response rate was achieved in 12% of patients, with a median duration of response of 4.2 months. A further 32% of patients had stable disease; the median survival was 6.6 months with 27% of patients alive at one year. The most common adverse events were acne-like skin rash (90% any grade, 6% grade 3) and asthenia (45% any grade, 9% grade 3) (Lenz H, 2005). Irinotecan, Single Agent Overview. The topoisomerase I inhibitor irinotecan (PÞzer’s Camptosar, PÞzer/Yakult/SanoÞ-Aventis’s Campto) is approved as monotherapy for patients who have relapsed following 5-FU chemotherapy. Irinotecan is administered intravenously at doses of 125 mg/m2 IV over 90 minutes, for 4 weeks every 6 weeks or 350 mg/m2 IV over4 90 min, once every 3 weeks. Mechanism of Action. Irinotecan (Figure 7) is a topoisomerase I inhibitor. Topoisomerase I inhibitors act as potent inhibitors of topoisomerase I, a nuclear enzyme that causes reversible single-strand breaks in DNA during mitosis. Topoisomerase I inhibitors bind to the topoisomerase I-DNA complex and prevent re-ligation of the DNA strand, resulting in cell death. Clinical Performance. As a second-line therapy for mCRC, single-agent irinotecan has been shown to produce response rates of 11–23% and to significantly improve survival compared with best-supportive-care median survival: 9.2 months versus 6.5 months. Single-agent irinotecan also increases median survival over infusional 5-FU: 10.8 versus 8.5 months (Cunningham D, 1998; Rougier P, 1998). These data demonstrate clinical beneÞts for patients whose disease has progressed while on standard 5-FU/LV treatment. Irinotecan’s most signiÞcant clinical toxicity is diarrhea, and treating physicians are likely to avoid irinotecan in patients prone to chemotherapy-related diarrhea. First-line treatment with single-agent irinotecan produces response rates of 18–29%. The observation of single-agent activity in mCRC, both as Þrst-line and second-line treatment, led to the development of irinotecan-containing regimens (see “IFL Regimen” and “FOLFIRI Regimen,” above) in order to improve response rates and survival over 5-FU/LV or single-agent irinotecan. Generally, combination treatments are now preferred and are under investigation as adjuvant treatments. Before oxaliplatin was introduced, irinotecan was considered a standard part of Þrst-line therapy in many cancer centers. Irinotecan has been competing with oxaliplatin since oxaliplatin’s approval as a Þrst-line therapy. Irinotecan is now used as a Þrst-line treatment in a minority of advanced-CRC patients (Blanke CD, 2005). However, it continues to play a key role in combination regimens, notably with cetuximab, as discussed later.

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Capecitabine, Single Agent Overview. Capecitabine (Roche’s Xeloda) (Figure 9) is an oral pro-drug of 5-FU. Capecitabine is indicated as Þrst-line treatment in patients with mCRC when treatment with ßuoropyrimidine therapy alone is preferred. Capecitabine elicits better response rates and has a different, more tolerable toxicity proÞle than bolus 5-FU. And, unlike 5-FU, it does not require coadministration of LV. However, no improvement in survival has been noted when capecitabine is compared with 5-FU. Capecitabine has been approved in the United States and Europe as a Þrst-line treatment for mCRC and is in Phase II clinical trials for mCRC in Japan. Clinical data have shown no beneÞt for adding LV to capecitabine therapy. Single-agent capecitabine has recently been approved in the United States and Europe for the adjuvant treatment of patients with Dukes’ C colon cancer on the basis of a large international Phase III trial—Xeloda Adjuvant Colon Cancer Trial (X-ACT). Capecitabine combinations in place of 5-FU/LV have also been investigated in several clinical trials in the United States and Europe. Mechanism of Action. Capecitabine is a ßuoropyrimidine. Fluoropyrimidines, which are structural analogues of naturally occurring metabolic products (generally folates, purines, and pyrimidines) needed for synthesis of nucleic acids, act by inhibiting thymidylate synthase (TS), a key enzyme in DNA synthesis. Clinical Performance. Capecitabine’s approval was based on the results of two large, multicenter Phase III trials involving more than 1,200 patients at hospitals and centers worldwide. Both trials compared capecitabine (2,500 mg/m2 per day in two divided doses, administered for 14 days every three weeks) with the Mayo Clinic regimen in patients with mCRC (Van Cutsem E, 2001; Hoff PM, 2001). Capecitabine yielded an overall response rate of 18.9% and 24.8% in the two trials, compared with 15% and 15.5% for 5-FU/LV. Duration of response, progression-free survival, and overall survival were similar in both arms in both trials. A higher incidence of grade 3 and 4 hand-and-foot syndrome occurred in patients receiving capecitabine (16%) compared with patients receiving 5-FU/LV (less than 1%). However, other grade 3 and 4 toxicities were more common among patients receiving 5-FU/LV than in patients receiving capecitabine. Grade 3 and 4 stomatitis occurred in 13–16% of 5-FU/LV patients and in 1–3% of capecitabine patients; grade 3 and 4 neutropenia occurred in 20–27% of 5-FU/LV patients and in 2–3% of capecitabine patients. Single-agent capecitabine has recently gained approval in the United States and Europe for the adjuvant treatment of patients with Dukes’ C colon cancer who have undergone complete resection of the primary tumor when treatment with ßuoropyrimidine therapy alone is preferred. Capecitabine’s approval was based on data from X-ACT, a large international Phase III trial whose results were presented at the 2005 ASCO meeting (Twelves C, 2005). A total of 1,987 patients with resected Dukes C colon cancer were randomized to receive capecitabine (1,250 mg/m2 twice daily days 1–14 every 3 weeks) or intravenous 5-FU/LV

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(Mayo Clinic regimen, a bolus 5-FU regimen: LV 20 mg/m2 plus 5-FU 425 mg/m2 days l–5, every 4 weeks) for 24 weeks. The study met its primary endpoint: capecitabine was at least equivalent to 5-FU/LV in terms of disease-free survival after a median follow-up of 4.3 years. A trend toward superior disease-free survival and overall survival for capecitabine was also observed. The three-year, disease-free survival rate was 66% for patients treated with capecitabine compared with 62.9% for patients treated with 5-FU/LV. Patients treated with capecitabine experienced less (all grades) diarrhea, nausea/vomiting, stomatitis, alopecia, and neutropenia, and less grade 3 and 4 neutropenia, stomatitis, and neutropenic fever/sepsis compared with 5-FU/LV patients. Non-life-threatening hand-and-foot syndrome and hyperbilirubinemia were more common with capecitabine than with 5-FU/LV. Researchers concluded that capecitabine should replace 5-FU/LV as adjuvant therapy for colon cancer (Twelves C, 2005; Cassidy J, 2004). Another study showed single-agent capecitabine to be effective and well tolerated as Þrst-line therapy in 51 elderly patients with advanced CRC (aged 70 years and older) who were considered ineligible for combination chemotherapy (Feliu J, 2005). Their overall response rate was 24%, including two complete responses and ten partial responses. The median time to progression and overall survival were 7 and 11 months, respectively. Oral administration of single-agent capecitabine averts the discomfort and inconvenience of IV therapy and is an important therapeutic option for elderly patients ineligible for combination chemotherapy. Capecitabine combinations are being investigated in many clinical trials. Several Phase II studies have reported response rates of slightly more than 50% for capecitabine combined with either irinotecan or oxaliplatin in the metastatic setting (Kerr D, 2002) and have conÞrmed equivalence with the corresponding 5-FU/LV-containing regimens. The capecitabine plus oxaliplatin (XELOX, or CAPOX) combination is a highly effective Þrst-line treatment for mCRC. Although the response rates, time to progression, and overall survival were similar to those seen with FOLFOX regimens, XELOX is more convenient and requires only one clinic visit per three-week cycle for a two-hour infusion of oxaliplatin (Cassidy J, 2004). Of the 96 patients in the XELOX study, 53 achieved an objective response, and 30 had stable disease maintained for at least three months. The median time to progression was 7.7 months, and median overall survival was 19.5 months after a minimum follow-up of 24 months. The one-year survival rate was 70%, dropping to 30% at two years; objective response rate was 45%. The safety data for XELOX were similar to data for the FOLFOX4 regimen, with a low incidence of myelosuppression. Evidence from this trial suggests that capecitabine has a strong potential to replace 5-FU/LV as the standard combination partner for oxaliplatin (Cassidy J, 2004). Capecitabine and oxaliplatin (the CAPOX regimen) in combination with cetuximab has shown signiÞcant clinical activity in 15 patients with EGFR-positive mCRC refractory to standard chemotherapy (Grothe W, 2005). Partial responses

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were observed in 27% of patients, 27% had stable disease, and 46% had progressive disease. The progression-free survival time was 2.5 months, and median overall survival was greater than 5.5 months. Researchers are also investigating the combination of capecitabine, oxaliplatin, and bevacizumab (XELOXA) as a more convenient and active regimen for untreated patients with mCRC. Preliminary results of a Phase II trial involving 30 patients, presented at the 2005 ASCO meeting, demonstrated that the XELOXA regimen is highly active (Fernando N, 2005). Overall, XELOX regimens offer a novel, active, well-tolerated, and active backbone around which innovative targeted agents can be incorporated. Raltitrexed, Single Agent Overview. After it became known that 5-FU exerts its primary cytotoxic effect in CRC by inhibiting thymidylate synthase (TS), several pharmaceutical companies developed selective TS inhibitors. The most established of these agents is raltitrexed (AstraZeneca’s Tomudex), which has limited application in treatment of mCRC. Raltitrexed is an oral inhibitor of TS with efÞcacy similar to that of 5-FU/LV. The antifolate is marketed in France, Italy, Spain, and the United Kingdom as a Þrst-line treatment for mCRC in patients who are unsuitable for treatment with 5-FU. In the United Kingdom, however, the National Institute for Clinical Excellence (NICE) has recommended that raltitrexed be administered only in the context of a well-designed clinical trial. AstraZeneca is no longer developing raltitrexed in the United States or Japan, and the company halted an adjuvant Phase III trial after an excess of deaths in the raltitrexed arm. Mechanism of Action. Raltitrexed (Figure 10) is a folate analogue. Folate analogues act by selectively inhibiting TS, a key enzyme in DNA synthesis. The inhibition of TS leads to DNA fragmentation and cell death. Clinical Performance. A meta-analysis has shown that patients treated with raltitrexed have a survival rate similar to that of patients treated with 5-FU/LV. The analysis included four Phase III trials that cumulatively involved 2,261 patients with untreated mCRC. The median survival time across the four trials was 11 months for 5-FU/LV compared with 10 months for raltitrexed (Cunningham D, 2002[b]). Analysis revealed no signiÞcant difference in response rates,

O

O HN H3C

N N

S

COOH HN

CH3 FIGURE 10. Structure of raltitrexed.

COOH

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but in three of the four trials, the median time to progression was longer for 5-FU/LV than for raltitrexed (six versus Þve months). Patients who received raltitrexed experienced signiÞcantly lower incidences of grade 3 or 4 mucositis and leukopenia. Researchers anticipated that the lower incidence of severe mucositis and leukopenia associated with raltitrexed, combined with its convenient dosing schedule, would give the compound a clinical advantage. In a trial that compared raltitrexed with bolus 5-FU regimens, quality of life was similar for both treatments except at the two-week point, when quality of life in the raltitrexed arm was superior; this exception corresponds to the time when 5-FU–induced mucositis is greatest (Cocconi G, 1998). In a trial that compared raltitrexed with infusional 5-FU regimens, however, patients taking raltitrexed experienced an inferior quality of life (Maughan TS, 2002). Trials have shown that the treatment-related death rate is higher with raltitrexed than with 5-FU/LV. An overview of randomized raltitrexed trials found that the level of raltitrexed’s causally related mortality was slightly higher than that of 5-FU/LV: 3.8% versus 2.6% (Zalcberg J, 1997). In the trial reported by Maughan and colleagues, 18 deaths were attributed to raltitrexed compared with a total of 3 attributed to two infusional 5-FU/LV regimens (Maughan TS, 2002). The authors of these studies note that clinicians should carefully monitor patients receiving raltitrexed and that appropriate dose modiÞcations should always be made. No advantage has been observed for replacing 5-FU/LV–containing regimens with raltitrexed as Þrst-line treatment of mCRC in patients who can tolerate 5-FU/LV therapy; thus, raltitrexed is likely to be reserved for second-line or subsequent treatment. Raltitrexed has been evaluated in combination with both oxaliplatin and irinotecan. The combination of raltitrexed/oxaliplatin achieved an overall response rate of 57%—signiÞcantly greater than either raltitrexed alone (20% response rate) or oxaliplatin alone (33% response rate). Median survival time exceeded 13 months, which was longer than the median survival time for either raltitrexed or oxaliplatin alone (Douillard JY, 2000[b]). Raltitrexed combined with irinotecan elicited responses in 38% of 62 previously untreated mCRC patients (Aparicio J, 2002). The combination was associated with some grade 3 and 4 toxicities—23% suffered diarrhea and 17% neutropenia—a proÞle that severely limits the utility of a raltitrexed/irinotecan combination in metastatic patients. In 2003, the combination of raltitrexed with irinotecan produced a response rate of 27% in 48 patients with untreated advanced CRC (Chiara S, 2003). The combination of raltitrexed and irinotecan is an attractive regimen but has a signiÞcant incidence of side effects. Raltitrexed has several properties that confer a theoretical advantage over 5-FU. It undergoes extensive intracellular polyglutamation, which allows it to be retained within cells for prolonged periods, and the metabolites of this polyglutamation are 100 times more potent than the parent compound for inhibiting thymidylate synthase. Raltitrexed consequently boasts long-lasting action that allows infrequent administration. Raltitrexed is administered by

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15-minute infusion once every three weeks; standard 5-FU/LV regimens can require Þve consecutive days of drug treatment or 48-hour 5-FU infusions. In a Phase II study of patients with stages II and III rectal cancer, neoadjuvant treatment with oxaliplatin, raltitrexed, and radiotherapy achieved signiÞcant downstaging in 76% of patients (25 of 33 patients) and complete response in 6 patients (18%) (Casado S, 2004). The regimen was found to be feasible, with a low toxicity proÞle. Based on recent Phase II data presented at the 2005 ASCO meeting, TOMOX (raltitrexed 3 mg/m2 plus oxaliplatin 130 mg/m2 , both on day one every three weeks) shows promising activity, with a response rate of 44% (Gravalos C, 2005). One hundred and ninety-one patients who had not received any prior therapy for advanced CRC were randomized to receive either FOLFOX4 or TOMOX. Of the 156 evaluable patients, the response rate was 44% in the TOMOX arm and 39% in the FOLFOX4 arm. The median time to progression was 7.4 months for the TOMOX arm versus 8.8 months for patients who received FOLFOX4. Median overall survival was 14.9 months and 13.8 months, respectively. Grade 3 and 4 neutropenia were more common in the FOLFOX4 group (36% versus 3.7%), with transaminitis more frequent in the in TOMOX group (1.3% versus 18.5%). This Phase II study reveals that FOLFOX4 and TOMOX demonstrate similarly high activity in terms of response rate, time to progression, and overall survival as Þrst-line treatment of advanced CRC (Gravalos C, 2005). Tegafur-Uracil, Single Agent Overview. The oral ßuoropyrimidine tegafur-uracil, also known as UFT (Bristol-Myers Squibb’s Uftoral/UFT, Otsuka’s UFT), is a combination of tegafur (a cytotoxic oral pyrimidine ßuoride) and uracil (a pyrimidine). Bristol-Myers Squibb’s Orzel is a co-packaged product containing oral UFT and oral leucovorin. The UFT/LV combination demonstrates antitumor activity equivalent to that of intravenously infused 5-FU. Notably, UFT/LV does not provoke the toxic reactions typical of many other ßuoropyrimidine-based therapies, such as handand-foot syndrome, neurological toxicity, myelosuppression, and cardiotoxicity. Developed in Japan by Taiho Pharmaceuticals, UFT had been marketed in that country for treating various cancers since 1984. The clinical impact of UFT alone (as well as in combination with LV) for the treatment of CRC was not realized until UFT was reevaluated outside Japan—after Bristol-Myers Squibb obtained development and marketing rights to the agent in 1995. In March 2001, the FDA issued a “not approvable” letter for UFT/LV because of concerns about the contribution of uracil to the efÞcacy of UFT and lack of evidence of equivalent activity with the Mayo Clinic regimen. The FDA’s stance was surprising, given the Oncologic Drug Advisory Committee’s (ODAC’s) unanimous recommendation for approval. Although Bristol-Myers Squibb expressed its intention to continue pursuing U.S. approval, no further updates are available at this time. The agent was approved in Europe in October 2000. Mechanism of Action. As noted, tegafur-uracil is a ßuoropyrimidine. Fluoropyrimidines, which are structural analogues of naturally occurring metabolic

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products (generally folates, purines, and pyrimidines) needed for synthesis of nucleic acids, act by inhibiting TS, a key enzyme in DNA synthesis. Clinical Performance. The ODAC recommendation and European approval of tegafur-uracil were based on two large Phase III studies of UFT/oral LV versus IV 5-FU/IV LV (Pazdur R, 1999; Carmichael J, 1999). The studies found the two regimens to have similar efÞcacy, although the UFT/LV regimen was better tolerated. In both studies, the UFT/LV regimen comprised UFT 300 mg/m2 per day and 75 or 90 mg/m2 of LV on days 1–28, every Þve weeks. Pazdur and colleagues compared UFT/LV with the standard Mayo Clinic regimen in 816 patients with untreated mCRC. Although the response rate was lower in the UFT/LV arm (12% versus 15%), the difference was not signiÞcant. The median times to progression were worse for patients receiving UFT/LV (3.5 months versus 3.8 months for the Mayo Clinic regimen), but overall survival was similar in the two arms (12 months versus 13 months) (Vernook A, 2001). The UFT/LV regimen caused signiÞcantly less mucositis (1% versus 20% for the Mayo Clinic regimen) and myelosuppression (1% versus 56%); rates of grade 3 or 4 diarrhea were similar for both regimens. The results of the second trial were similar. Several Phase I/II trials are evaluating the tolerability of UFT/LV in combination with other agents—such as oxaliplatin (Douillard JY, 2004; Garcia-Gir´on C, 2000; Rosati G, 2004,), raltitrexed (Mel JR, 2000), raltitrexed and oxaliplatin (Garcia-Alfonso P, 2002), and irinotecan (Douillard JY, 2004; Hill M, 2000)—in Þrst-line treatment of advanced CRC. Based on preliminary results, researchers report, these combinations exhibit antitumor activity comparable to that of 5-FU combinations. The agents are well tolerated and show no unexpected toxicity. A Phase III randomized trial of UFT as adjuvant therapy following surgery, conducted in several centers in Japan, indicated that UFT is effective at lowering the risk of local recurrence following surgery, particularly in rectal cancer patients. A multicenter Phase III U.S. trial has compared UFT/LV with IV 5-FU/LV as adjuvant therapy following resection of stage II and III colon cancer. The trial, whose end points were overall and disease-free survival, enrolled 1,608 patients. Preliminary results presented at the 2004 ASCO meeting demonstrated no signiÞcant differences in disease-free survival or overall survival between the two treatment arms over a 64-month follow-up period, with equal tolerability in patients with stages II-III colon cancer (Wolmark N, 2004). These Þndings are of major signiÞcance because they reinforce the growing importance of oral chemotherapy in the treatment of CRC. Nonpharmacological Approaches Surgery is the primary treatment approach for patients with either colon or rectal cancer and is employed with both curative intent (in early-stage disease) and palliative intent (to reduce symptoms, such as pain from obstruction). Overall, surgery will cure approximately 50% of CRC patients and is used alone in stage I patients and low-risk stage II patients. Optimally, the tumor is entirely resected,

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with ample proximal, distal, and radial margins, along with regional lymph nodes, and the procedure has a minimal effect on the bowel. Because of anatomic differences, the tumor and the tissue surrounding the tumor generally can be more fully resected in colon cancer than in rectal cancer. Consequently, local recurrence rates are higher for rectal cancer than for colon cancer. When metastatic disease is limited to isolated sites in the liver or lungs, surgical management is possible because the potential for cure exists. Usually, however, only patients with up to four isolated metastases—approximately 10% of all patients—are eligible for hepatic resection because the operation does not improve the chance of survival unless all known disease is removed. Fewer than one-third of these patients are cured by surgery alone. Radiotherapy is widely employed to prevent local recurrence in high-risk rectal cancer patients, to treat locally recurrent disease, to make locally advanced disease resectable, and to control pain. Although radiotherapy for stage II and III disease reduces local recurrence rates by 16–50%, its effect on overall survival is less clear. Two meta-analyses have reported conßicting results, one suggesting a small survival advantage with radiotherapy (Camma C, 2000) and the other suggesting no difference between patients who received radiotherapy and patients who did not (Colorectal Cancer Collaborative Group, 2001). Preoperative and postoperative radiotherapy are two alternative strategies. In terms of overall survival, preoperative radiotherapy seems to offer some advantages, according to Hay and colleagues, though it has the disadvantage that postoperative staging is more reliable than preoperative tumor assessment (Hay J, 2003). Evidence also suggests that preoperative radiotherapy reduces toxicity and rates of local recurrence, but not mortality or distant recurrence (i.e., not overall survival) compared with postoperative radiotherapy for the treatment of locally advanced rectal cancer (Barclay L, 2004; Sauer R, 2004). Results on radiotherapy for rectal cancer remain conßicting. Preoperative therapy delays surgery by eight to ten weeks: four to Þve weeks for the treatment and four to Þve weeks for tissue repair. But preoperative radiotherapy offers the following advantages over postoperative radiotherapy: • •

•

•

Doses of adjuvant (postoperative) chemotherapy are not compromised by the additional toxicity of concurrent radiation therapy. The opportunity to perform sphincter-sparing procedures is improved because tissues at the periphery of the tumor are sterilized against cancer before surgery. Overall treatment morbidity is reduced (e.g., wound healing is less affected by preoperative radiotherapy than by postoperative therapy). A greater proportion of patients receive the scheduled course of radiotherapy (some patients scheduled to receive postoperative therapy do not receive it because of complications following surgery). Radiation therapy is less effective for managing recurrent disease than in preventing recurrence. Recurrent tumors are often encased in a Þbrotic matrix as a result of surgical manipulation of the original tumor and so are less susceptible to radiation. Nonetheless, for patients who received no prior

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483

radiation, localized pelvic recurrence can be managed in many cases by a combination of aggressive surgery and radiotherapy. For patients with prior irradiation, further use of radiation therapy when disease recurs is extremely limited by the sensitivity of surrounding normal tissue. By itself, radiotherapy offers only short-term pain relief by reducing the size of the tumor.

EMERGING THERAPIES A wide range of emerging therapeutics that target colorectal cancer (CRC) are undergoing clinical evaluation, from drugs designed to enhance the efÞcacy of existing regimens to immunostimulant vaccines and gene therapies. Emerging therapies must improve survival and address the issue of resistance to traditional chemotherapies. Since 2003, overall survival of metastatic CRC (mCRC) has improved signiÞcantly owing to the approval of two monoclonal antibodies (MAbs): bevacizumab (Genentech/Roche’s Avastin) and cetuximab (ImClone/Bristol-Myers Squibb/Merck KGaA’s Erbitux). Bevacizumab is well tolerated and—in combination with the IFL chemotherapy regimen (irinotecan [PÞzer’s Camptosar, PÞzer/Yakult/SanoÞ-Aventis’s Campto, Daiichi’s Topotecin], bolus 5-ßuorouracil [5-FU, generics], and leucovorin [LV, generics])—signiÞcantly improves the median overall survival in Þrst-line treatment of patients with mCRC by a signiÞcant 4.7 months. Cetuximab also produces clinically signiÞcant activity when given alone or in combination with irinotecan in the second-line treatment of patients with epidermal growth factor receptor (EGFR)-positive, irinotecan-refractory mCRC. These new treatments represent a major step forward and give patients with advanced CRC additional options. For nonmetastatic disease, surgery remains the primary curative treatment for CRC patients; however, with new approvals, more intensive drug regimens proven in the metastatic setting are being incorporated into adjuvant drug therapy. Despite improvements with cytotoxic agents, major unmet needs remain in late-stage CRC, and metastatic patients ultimately die of their disease. Given this need, novel mechanisms of action and efÞcacies that are synergistic with conventional chemotherapies are crucial requirements for new agents to treat CRC because such agents will allow the combination of therapies for maximum effect. Of the large number of agents in development for CRC, Table 8 lists the most promising emerging therapies. Epidermal Growth Factor Receptor Inhibitors Overview. A vast amount of R&D is ongoing in the area of EGFR inhibition, driven by the success of cetuximab. Cetuximab, a chimeric (mouse) MAb, effectively inhibits the proliferation of a variety of EGFR-expressing cancer cells. Other notable agents include Merck KGaA/Takeda’s humanized EGFR-targeting

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TABLE 8. Emerging Therapies in Development for Colorectal Cancer Compound

Development Phase

Epidermal growth factor receptor inhibitors Erlotinib (Tarceva) United States II Europe — Japan I

Marketing Company

Genentech/OSI Roche Chugai

Gefitinib (Iressa) United States Europe Japan

D — D

AstraZeneca — AstraZeneca

Panitumumab United States Europe Japan

III III —

Abgenix/Amgen Abgenix/Amgen —

Vascular endothelial growth factor inhibitors Vatalanib United States III Europe III Japan — Farnesyl protein transferase inhibitors Tipifarnib United States III Europe III Japan — Immunotherapies OncoVax United States Europe Japan

Novartis/Schering AG Novartis/Schering AG —

Johnson & Johnson/Janssen Johnson & Johnson/Janssen —

— — —

Intracel — —

IGN101 United States Europe Japan

— II —

— Aphton —

TroVax United States Europe Japan

— II —

Oxford BioMedica

Theratope United States Europe Japan

II — —

Biomira — —

dSLIM United States Europe Japan

— II —

— Mologen/Sanofi-Aventis —

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TABLE 8. (continued) Compound Cytotoxic agents Pemetrexed (Alimta) United States Europe Japan

Development Phase

Marketing Company

II II —

Eli Lilly Eli Lilly —

Edotecarin United States Europe Japan

D — II

Pfizer — Banyu

Aroplatin (Platar) United States Europe Japan

II — —

Antigenics — —

Gimatecan (ST-1481, LBQ-707) United States Europe Japan

— II —

— Novartis/Sigma-Tau —

II II I

Daiichi Pharmaceutical Daiichi Pharmaceutical Daiichi Pharmaceutical

II — I

Abbott Laboratories — Eisai

S S —

Pfizer Pfizer —

Microtubule modulators DJ-927 United States Europe Japan ABT-751 United States Europe Japan COX-2 inhibitors Celecoxib (Celebrex) United States Europe Japan COX-2 = Cyclooxygenase-2. D = Discontinued. S = Suspended.

MAb matuzumab, which is undergoing Phase II trials for solid tumors including CRC, and Genentech/Roche’s trastuzumab (Herceptin), a HER-2 inhibitor marketed as a breast cancer therapy that is being investigated in Phase I/II trials for CRC. (HER-2 is a member of the EGFR family of tyrosine kinase receptors.) In September 2005, Genzyme announced the launch of its genetic test, called EGFR Mutation Assay, for patients with non-small-cell lung cancer (NSCLC). The test will be used to detect EGFR mutations in NSCLC patients. The availability of this test will help physicians identify patients who are likely to respond to treatment with targeted therapies, thereby improving survival. Two EGFR targeted therapies are under investigation for the treatment of CRC.

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Mechanism of Action. The EGFR family is a group of four structurally similar growth factor receptors with tyrosine-kinase activity: HER-1/erb-B1, HER-2/erbB2, HER-3/erb-B3, and HER-4/erb-B4. They are composed of an extracellular ligand-binding domain, a transmembrane domain, and an intracellular tyrosine kinase domain (Pao W, 2005; Sridhar SS, 2003). These receptors are widely expressed on the cell surface and play a critical role in cancer development. Binding to the receptor results in tyrosine kinase activation and receptor autophosphorylation, which then initiate signal transduction cascades implicated in cell proliferation. Drug intervention, either at the EGFR or the intracellular tyrosine kinase, blocks cell signaling pathways, thereby preventing proliferation and subsequent tumor growth. Erlotinib. Erlotinib (Tarceva), is a small-molecule EGFR tyrosine kinase inhibitor under development by OSI Pharmaceuticals in alliance with Genentech, Roche, and Chugai. Erlotinib inhibits the intracellular tyrosine kinase domain of the EGFR, thereby blocking receptor activity. This orally active quinazoline derivative is in Phase II trials in the United States and Phase I trials in Japan for CRC. Erlotinib has been approved in the United States and Switzerland for the treatment of patients with locally advanced or metastatic NSCLC after failure of at least one prior chemotherapy regimen. However, erlotinib has failed to demonstrate a survival advantage when incorporated into a Þrst-line NSCLC regimen in a Phase III randomized trial disclosed in mid-2003. An ongoing Phase II trial presented at the 2005 American Society of Clinical Oncology (ASCO) meeting assessed the antitumor activity of single-agent erlotinib in second- and third-line treatment of 41 patients with mCRC who had previously received one (13 patients) or two (28 patients) 5-FU–based chemotherapy regimens, including irinotecan and/or oxaliplatin (Keilholz U, 2005). Patients received erlotinib at a daily dose of 150 mg until disease progression. Singleagent erlotinib was generally well tolerated. The toxicity was mild and included skin rash. Of the 24 patients evaluable for efÞcacy, a partial response was observed in 2 patients (8%) and stable disease in 33% of patients. Previous Phase II trials were presented at the 2003 ASCO meeting. In one, 30 patients with mCRC received erlotinib at a daily dose of 150 mg (Oza AM, 2003). Of 25 evaluable patients, 8 had stable disease for at least 98 days; 12 patients had progressive disease. The most common adverse events of any grade were rash in 25 patients, diarrhea in 11 patients, and lymphopenia in 13 patients. Researchers also observed grade 4 adverse events (2 patients experienced constipation) and grade 3 adverse events (24 incidences in all; the most common events were rash in 4 patients, diarrhea in 3 patients, nausea in 2 patients, and vomiting in 2 patients). Interestingly, EGFR expression, as indicated by staining techniques, did not correlate with response. Data from another Phase II trial presented at the ASCO meeting in 2003 showed the feasibility and tolerability of combining erlotinib with the FOLFOX4 regimen in solid cancers, including CRC (Hanauske AR, 2003). All 24 patients

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enrolled in the trial were evaluable for response. Partial response was achieved in 6 patients (5 with CRC), 11 patients had stable disease (6 with CRC), and 7 patients had progressive disease (4 with CRC). The most common side effects were rash and diarrhea (Hanauske AR, 2003). Erlotinib in combination with oxaliplatin and capecitabine has been evaluated in a Phase II study of previously treated patients with mCRC (Meyerhardt JA, 2004). Of the 22 patients enrolled in the trial, all were evaluable for toxicity and 20 were evaluable for response. The partial response rate was 20%, and stable disease was observed in 64% of patients. Gastrointestinal (GI) toxicities were more common with this combination (Meyerhardt JA, 2004). Gefitinib. GeÞtinib (AstraZeneca’s Iressa) has now been discontinued. This was another small molecule synthesized chemically and therefore much less expensive to manufacture than biologic agents such as cetuximab. GeÞtinib had U.S. FDA, Japanese, and Swiss approval as a once-daily oral monotherapy for advanced NSCLC in patients who had failed previous chemotherapy regimens. However, geÞtinib failed to demonstrate an overall survival advantage in the follow-up Phase III Iressa Survival Evaluation in Lung (ISEL) cancer trial that compared geÞtinib with placebo in patients with advanced, recurrent NSCLC (AstraZeneca, press release, December 2004). Following a discussion with the FDA’s Oncology Drugs Advisory Committee (ODAC), AstraZeneca announced in June 2005 that it is making a labeling change to geÞtinib. Based on the revised label, Iressa should be used only in patients who are beneÞting or have beneÞted from Iressa. Iressa’s new label is effective as of September 15, 2005. After this date, new patients have not been allowed access to Iressa unless they are being enrolled in a clinical trial approved by an Institutional Review Board (IRB) prior to June 17, 2005, or they are part of a clinical trial that is being conducted under an investigational new drug application (IND) (AstraZeneca, press release, June 2005). GeÞtinib inhibited the intracellular tyrosine kinase domain of the EGFR, blocking receptor activity. Unlike MAbs, geÞtinib did not cause receptor internalization, which is a potential disadvantage because receptor blockage could be reversed. Updated preliminary results of Phase II data from a U.S. trial evaluating geÞtinib in combination with 5-FU/LV/oxaliplatin (the FOLFOX4 regimen) in mCRC patients were disclosed at the 2004 ASCO meeting (Fisher GA, 2004). In the trial, patients stratiÞed to group A (no prior therapy for metastatic disease [32 patients]) or group B (prior therapy for metastatic disease [24 patients]) continued on treatment until progression or until they reached unacceptable levels of toxicity. All 56 patients were evaluable for toxicity, and 49 were evaluable for response. Two patients withdrew prior to completing four treatment cycles because of unacceptable toxicity. Grade 3 and 4 toxicities included diarrhea (49%), nausea (28%), neutropenia (53%), and vomiting (21%). One treatmentrelated death (E. coli sepsis) was observed. Response rates were encouraging: in group A, 21 of 27 patients (78%) achieved a partial response, while 8 of 22

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patients (36%) in group B achieved a partial response. Nine patients in group A were amenable for resection of their liver metastases, of whom only four had been considered to have resectable disease at the beginning of the study. GeÞtinib was also investigated as a single agent in the treatment of CRC. An Eastern Cooperative Oncology Group (ECOG) randomized Phase II trial of single-agent geÞtinib in 115 patients with mCRC failed to demonstrate any activity (Rothenberg ML, 2004). Patients who had progressed to irinotecan/5FU/LV were randomized to two dose levels: 250 mg and 500 mg per day of geÞtinib. The primary end point was progression-free survival at four months. Eighty mCRC patients (70%) agreed to undergo tumor biopsies. Researchers detected no signiÞcant changes in biological indicators of EGFRpathway activation from serum or tumor biopsies obtained before and one week after treatment with geÞtinib (Rothenberg ML, 2004). Among the 110 patients evaluable for efÞcacy, there were no complete responses, and only one patient in the 500 mg dose group had a partial response, yielding an overall response rate of 1%. Eighteen percent of patients achieved stable disease in the trial. Fourmonth, progression-free survival was 13%, with a median survival of 6.3 months for both groups. The most common toxicities were skin rash and diarrhea, which were slightly more common at the 500 mg dose level. Eventually, researchers concluded that geÞtinib is inactive as a single agent in patients with progressive mCRC. As mentioned above, in July 2005, AstraZeneca discontinued clinical development of geÞtinib for treatment of CRC. Panitumumab. Following in the footsteps of cetuximab, the EGFR-directed MAb Amgen/Abgenix’s panitumumab (formerly ABX-EGF) is under clinical evaluation for the treatment of CRC. Amgen (formerly Immunex) holds a license for worldwide codevelopment and copromotion of the antibody with Abgenix. Panitumumab targets EGFR that is overexpressed in a variety of cancers, including CRC. Because this agent is fully humanized, it could potentially cause fewer unwanted immunologic responses than chimeric MAbs such as cetuximab. Updated Phase II interim data for panitumumab were presented at the 2004 ASCO meeting (Hecht JR, 2004). Patients with mCRC who had failed therapy with a ßuoropyrimidine and either irinotecan or oxaliplatin (or both) received single-agent panitumumab. A total of 148 patients were enrolled, with all patients evaluable after eight weeks of treatment. Fifteen patients (10%) had conÞrmed partial responses, and 54 (36.5%) had stable disease. Skin rash, the most frequent side effect, occurred in more than 90% of patients and was grade 3 in 3.4% of patients. Other grade 3 and grade 4 adverse events occurring in more than 3 patients were fatigue (9 patients) and anemia (4 patients). Grade 3 infusionrelated reaction was observed in 1 patient. Panitumumab was well tolerated and demonstrated encouraging single-agent antitumor activity in heavily pretreated mCRC patients, including patients with lower EGFR staining intensity (Hecht JR, 2004). At the 2005 ASCO meeting, matured data from the above Phase II trial were presented (Malik I, 2005). Based on the recent results, the median progressionfree survival time for all patients was 13.6 weeks, and the median overall survival

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was 37.6 weeks after a median follow-up time of 30.9 weeks. According to a central review, the objective response rate at the end of treatment was 9%, and median duration of response was 18.1 weeks. Grade 3 and 4 adverse events were rash (5 patients), fatigue (4 patients), vomiting (2 patients), nausea (1 patient), and pruritus (1 patient). Nearly all patients had skin rash (141 patients [95%]), with 5% at grade 3. These recent data conÞrm the safety and response Þndings, and provide encouraging survival data for patients with mCRC who have failed multiple lines of standard chemotherapy (Malik I, 2005). In April 2005, Amgen and Abgenix initiated a randomized, multicenter, openlabel Phase III trial called Panitumumab Advanced Colorectal Cancer Evaluation (PACCE) for the Þrst-line treatment of mCRC. The study evaluated the addition of panitumumab to bevacizumab and either oxaliplatin or irinotecan every other week, with endpoints of progression-free survival, overall survival, and response rate. Enrollment of approximately 1,000 patients was noted (Abgenix, press release, April 26, 2005). This pivotal Phase III study will further explored the potential beneÞts of panitumumab in the Þrst-line treatment of mCRC. Back in early 2004, pivotal Phase III trials evaluating single-agent panitumumab as a third-line therapy in patients with advanced CRC were also initiated. In August 2005, the FDA awarded panitumumab fast-track designation for mCRC. In January 2006, Amgen and Abgenix submitted a biologics license application (BLA) to the FDA for panitumumab in patients who have failed prior standard chemotherapy, including irinotecan and oxaliplatin. The anticipation is that the drug would be approved in the second half of 2006. At this time, no other information is available. Vascular Endothelial Growth Factor Inhibitors Overview. Angiogenesis, the formation of new blood vessels, plays a major role in many normal physiological processes. It also plays a central role in several pathological conditions, including solid tumor growth and metastasis, and therefore represents an area of considerable commercial, and clinical, interest. VEGF is considered one of the most potent and important pathways of tumor angiogenesis, and its role has been validated by the clinical success of the VEGF inhibitor bevacizumab for the treatment of CRC. Bevacizumab acts by binding to VEGF and preventing its interaction with VEGF receptors. Novartis/Schering’s vatalanib is a small-molecule, oral VEGF receptor tyrosine kinase inhibitor in a more advanced stage of development than the many other angiogenesis inhibitors in development for CRC. Other interesting antiangiogenic compounds within this class include ImClone’s IMC-1121B, a fully human anti-VEGF receptor (anti-VEGFR2) antibody that blocks Kdr-VEGFR2 interaction; MedImmune’s α-v/β-3 integrin antagonist; Bayer/Onyx Pharmaceuticals’ BAY-439006 (Sorafenib); and OXiGENE’s combretastatin A4 pro-drug. These agents are in early-phase clinical testing for the treatment of CRC. Mechanism of Action. VEGF is a multifunctional cytokine and potent permeability factor that is secreted in response to hypoxia (reduced oxygen). The

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biological effects of VEGF are mediated by the binding of VEGF to one of three endothelial surface receptors: VEGF-R1 (ßt-1), VEGF-R2 (ßk-1/kdr), or VEGF-R3; binding to the coreceptor neurophilin enhances signaling. VEGF has a major angiogenesis-promoting effect. The inhibition of VEGF receptors, either by interaction at the extracellular domain or by inhibition of intracellular receptortyrosine kinases, will, in the best-case scenario, kill the solid tumor that depends on new blood vessel formulation for its survival, or at least slow tumor growth by preventing new vessel formation and thereby limiting the supply of blood to the tumor. Vatalanib. Vatalanib, under development by Novartis and Schering, is an orally administered, small-molecule angiogenesis inhibitor that targets the VEGFR-1, 2, and -3 receptors; targeting more than one VEGFR subtype could provide better anti-angiogenic action. The lead indication for vatalanib is CRC, for which the compound is undergoing two Phase III trials in the United States and Europe, both initiated in the Þrst quarter of 2003. The Þrst trial, Colorectal Oral Novel Therapy for the Inhibition of Angiogenesis and Retarding of Metastases (CONFIRM) 1, is evaluating the potential progression-free survival and overall survival beneÞt of once-daily vatalanib in combination with FOLFOX4, compared with FOLFOX4 alone in previously untreated mCRC patients. A total of 1,168 patients were randomized to receive FOLFOX4 plus vatalanib (585 patients) or FOLFOX4 alone (583 patients). Preliminary results were disclosed at the 2005 ASCO meeting (Hecht JR, 2005). Based on central review, vatalanib in combination with FOLFOX4 showed no improvement over FOLFOX4 alone: progression-free survival was 7.7 months for FOLFOX4 with vatalanib and 7.6 months for FOLFOX4 alone. Treatment with vatalanib in combination with FOLFOX4 was generally well tolerated. Side effects attributed to vatalanib were similar to those of other VEGF inhibitors. Interestingly, an exploratory analysis suggests that patients with high lactate dehydrogenase (LDH) levels (indicative of liver involvement) beneÞted the most from vatalanib treatment in terms of progression-free survival (Hecht JR, 2005). This subgroup of patients will likely be investigated further. Another ongoing Phase III trial, CONFIRM 2, is comparing vatalanib (once daily) in combination with FOLFOX4 and FOLFOX4 alone in patients with mCRC whose disease has progressed after irinotecan-based Þrst-line chemotherapy. According to an independent Data Safety Monitoring Board (DSMB) assessment, interim analysis from the CONFIRM 2 trial shows that vatalanib is unlikely to extend survival in second-line therapy for mCRC (Novartis and Schering, press releases, July 28, 2005). Patients with high LDH levels in the vatalanib treatment group had an improved progression-free survival. This information is consistent with earlier Þndings in the CONFIRM 1 trial, suggesting that LDH levels may predict response to vatalanib. Investigators are still hopeful that vatalanib could extend overall survival in the Þnal analysis. The investigators of the CONFIRM 2 trial will decide whether patients should continue to receive treatment; all patients in the study will be followed up for overall survival. The CONFIRM 1 trial has proceeded as planned

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with overall survival data expected in the second half of 2006. Even if the results are positive, vatalanib could still Þnd it difÞcult to compete with bevacizumab, given the latter agent’s signiÞcant improvement of overall survival in mCRC. Farnesyl Transferase Inhibitors Mechanism of Action. Protein farnesylation is a modulatory process that permits the addition of several cellular proteins to cell membranes. Farnesylated proteins—including ras, rac, rho, and other small G-proteins—are involved in cell transformation and proliferation. For this reason, farnesyl transferase inhibitors (FTIs) have emerged as a new class of antineoplastic agent. Mutated ras oncogenes, in particular, play a causative role in malignant cell transformation and are frequently detected in human tumor cells. Inhibition of ras oncoprotein farnesylation is considered an attractive target for development of antineoplastic agents. However, even though many farnesyl transferase inhibitors have undergone clinical trials for use in CRC, results with these agents have been disappointing (Cunningham D, 2002 (a). Tipifarnib. Tipifarnib (R-115777; Zarnestra), an oral farnesyltransferase inhibitor, is being developed by Janssen and its parent company, Johnson & Johnson. Phase III trials in advanced CRC patients have been carried out in the United States and Europe. Preliminary Phase III trial results presented at the 2002 ASCO meeting were disappointing. The international randomized trial compared tipifarnib with placebo in patients with advanced refractory CRC who had failed two prior chemotherapy regimens for metastatic disease. Failure was deÞned as disease progression during treatment or within three months thereafter. The study was designed to detect a 50% increase in overall survival, the primary endpoint. The primary endpoint was not reached; the median overall survival of patients, all receiving tipifarnib, was 5.7 months versus 6.1 months for patients receiving placebo (Cunningham D, 2002). Updated results of this Phase III trial remain disappointing. Only one patient had a partial response in the tipifarnib group. Stable disease lasting more than 3 months was observed in 24.3% of patients receiving tipifarnib compared with 12.8% of patients in the placebo group. However, this result did not translate into a statistically signiÞcant increase in progression-free survival or improved quality of life (Rao S, 2004). Tipifarnib monotherapy did not improve overall survival compared with best supportive care in patients with refractory advanced CRC. Negative Phase II data of another trial were presented at the ASCO 2003 meeting (Whitehead RP, 2003). Of the 51 eligible patients, one patient (2%) had a partial response. Two additional patients had unconÞrmed partial responses, for an overall response rate of 6%. The estimated median overall survival was 7.9 months, and the time to treatment failure was 1.7 months. Even if investigations into tipifarnib’s potential in combination with chemotherapy are still warranted in earlier stages of CRC, the lack of efÞcacy of

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tipifarnib monotherapy will make it difÞcult to embark on further studies of this agent in CRC. Immunotherapies Overview. Numerous immunotherapies are being developed for use in CRC treatment and/or prophylaxis. We have collectively grouped all major immunotherapies in this section. Other noteworthy compounds that do not receive full discussion here include anti-CEA immunotherapies from Therion Biologics and SanoÞ-Aventis and the vaccines CTP-37 (AVI Biopharma’s Avicine) and Antigenics’ Oncophage (vitespen, formerly HSPPC-96). No signiÞcant positive data have been made available on these agents. For CRC vaccines, the requirement for a strong immune response for effecting recovery has yet to be empirically proved, because no randomized, large-scale trial data are available. Mechanism of Action. Immunotherapy is a general term that encompasses a wide variety of mechanisms of action, but the fundamental mechanism is the production of an immune response in order to treat or prevent disease. Major anticancer immunotherapeutic subcategories include MAbs, anti-idiotypic antibodies, tumor antigens, cellular vaccines, gene therapy, immunostimulatory drugs, and other biologic drugs. The concept of utilizing the immune system for treatment of cancer has been tried for some time without any direct breakthrough success. So far, “passive” immunotherapy (that which does not require the immune system to take an active role in the agent’s anticancer activity) has experienced the greatest success, with antibody and cytokine therapies leading the way. OncoVAX. OncoVAX, developed by Intracel, is a vaccine being investigated as a postsurgical treatment for stage II colon cancer. OncoVAX is an autologous tumor cell/bacillus Calmette Guerin (BCG) vaccine that is prepared for each patient using the patient’s own surgically removed tumor. The vaccine is in Phase III development in the United States for colon cancer. Recent results of a randomized, multicenter, Phase III trial conducted in the Netherlands in 254 colon cancer patients—including 157 in stage II and 84 in stage III—in the adjuvant setting showed a statistically signiÞcant beneÞt of OncoVAX for all end points, including overall survival, recurrence-free interval, and recurrence-free survival in patients with stage II colon cancer (Uyl-de Groot CA, 2005). Patients were randomized to receive either OncoVAX therapy or no treatment after surgical resection of the primary tumor. Treatment with the vaccine was safe and well tolerated. After a median follow-up of 5.8 years, OncoVAX produced a signiÞcant prognostic beneÞt, positive clinical outcomes, and was cost-effective in patients with stage II colon cancer. No statistically prognostic beneÞts occurred in the patients with stage III colon cancer (Uyl-de Groot CA, 2005).

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However, an ECOG randomized, multicenter Phase III trial of OncoVAX in 412 patients with stage II and III colon cancer did not reach statistical significance in the intent-to-treat population. After a median follow-up of 7.6 years, no statistically signiÞcant differences were found in clinical outcomes between the groups who received vaccine and those who underwent surgery alone. However, trends favoring the vaccine with regard to disease-free survival and overall survival were seen in 262 evaluable patients (Harris JE, 2000). Although OncoVAX is already marketed in the Netherlands for the treatment of stage II colon cancer, caution about cancer vaccines continues. Stage II CRC is still considered largely curable by optimal surgery and well-tolerated, low-cost, 5-FU-based adjuvant therapy. IGN101. Aphton’s IGN101 is an antiepithelial cell adhesion molecule (antiEpCAM) vaccine under development for the treatment of epithelial cancers. Aphton acquired this agent when it acquired the drug’s developer, Igeneon, in March 2005. IGN101 contains 0.5 mg alum-absorbed murine MAb edrecolomab that structurally mimics EpCAM, a cell-surface glycoprotein expressed on both tumor and normal tissues. Positive Phase II data in 239 evaluable patients with stages III or IV epithelial cancers, including CRC (163 [95 stage IV]), upper-GI tract cancer (32), NSCLC (38), and liver cancer (6), were presented at the 2005 ASCO meeting (Himmler G, 2005). Results of this randomized, double-blind, placebo-controlled trial demonstrated a trend toward prolonged survival in the overall stage IV CRC group during the Þrst year of treatment: overall survival at 12 months was 48.8% for placebo compared with 60% for IGN101. Also, a statistically signiÞcant survival prolongation was observed in 53 patients with stage IV rectal cancer: median survival was 415 days for IGN101-treated patients compared with 253 days for the placebo group. The one-year survival rate more than doubled—from 29.5% for patients receiving placebo to 62% for patients receiving IGN101. Use of concomitant chemotherapy had no effect on survival. Vaccinations with IGN101 were well tolerated. Almost all patients in the IGN101 group mounted an antibody response to the vaccine antigen (Himmler G, 2005). Another Phase II multicenter, open-label, single-arm, uncontrolled trial was presented at ASCO 2005 (Samonigg H, 2005). Circulating tumor cells in peripheral blood were analyzed in patients on days 1, 29, 57, and 71. In the 16 patients who completed the study, the median number of circulating tumor cells decreased from 10.5 at day 1 to 5 cells per blood sample after four vaccinations with IGN101 at day 71. The difference, however, was not statistically signiÞcant. Researchers concluded that in several patients with CRC, ovarian cancer, or breast cancer, a reduction of circulating tumor cells was observed after approximately one-third of the study had been completed. In metastatic breast cancer patients, an elevated number of circulating tumor cells has been shown to predict short survival (Samonigg H, 2005). TroVax. Oxford BioMedica’s TroVax, a gene-based tumor vaccine, is in Phase II development in patients with mCRC. TroVax delivers a tumor-associated antigen

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(5T4) using a pox virus vector. 5T4 is widely expressed on the surface of a wide range of solid cancer cells, including CRC; its presence is correlated with poor prognosis. Two open-label, Phase II trials are ongoing in which TroVax is administered alongside chemotherapy with either irinotecan/5-FU/LV (the IFL regimen) or 5-FU/LV/oxaliplatin (the FOLFOX regimen). TroVax is administered in up to six doses in both trials: two prior to, two during, and two post-chemotherapy. The primary endpoint is to assess the immune response to TroVax; the secondary endpoint is the safety proÞle of TroVax in combination with chemotherapy. Interim results for both studies were presented at the 2005 ASCO meeting (Harrop R, 2005). Both endpoints—immunologic responses and safety—were achieved. In the irinotecan trial, of the 14 patients who received more than or equal to four TroVax injections, 9 patients had positive proliferative responses to 5T4 protein, and 7 mounted antibody responses. In the oxaliplatin trial, of the 7 patients who received more than or equal to four TroVax injections, 2 patients showed positive proliferative responses to 5T4 protein, and 4 mounted 5T4-speciÞc antibody responses. Injection site reactions were the main side effect observed. TroVax was safe and well tolerated when it was administered in conjunction with the IFL or FOLFOX regimens (Harrop R, 2005). More-advanced data are required before the effectiveness of TroVax in CRC can be assessed. Theratope. Biomira’s entry in the CRC vaccine arena is Theratope (STn-KLH), a synthetic Sialyl-Tn antigen linked to the keyhole limpet hemocyanin (KLH) protein carrier and administered with Corixa’s Detox-B adjuvant. The working hypothesis for this vaccine is that it stimulates an immune response to the tumorassociated STn epitope, which leads to an anticancer effect. Merck KGaA’s U.S. afÞliate, EMD Pharmaceuticals, had been codeveloping Theratope in the United States; however, in June 2004, all rights were returned to Biomira. Given Theratope’s failure in breast cancer trials, additional trials were likely to be required to support its registration; the agent no longer met Merck KGaA’s future product launch timetable. A Phase II study investigating Theratope in mCRC is ongoing in the United States. Preliminary Phase II data of a single-arm pilot trial were presented at the 2003 ASCO meeting (Butts CA, 2003). Data showed that patients exhibit a response to vaccination while on concurrent chemotherapy, though no time-to-progression or survival data are available. Results of a Phase II single-center, double-blind trial that randomized 45 mCRC patients by dose were disclosed at the ASCO meeting in 2001. Survival was analyzed in a subgroup of 35 colon cancer patients, 77% of whom had previous treatment for metastases. Median survival was 14.2 months from the commencement of treatment with Theratope. Previous trials have also shown a signiÞcantly beneÞcial survival effect (Tempero MA, 2001). All 45 patients experienced transient, delayed-type, hypersensitivity-like symptoms at injection sites. Systemic toxicity was minimal; 15 patients noted mild myalgia, and a smaller number reported other ßulike symptoms. Phase III data that further examine the observed survival beneÞt are warranted. The drug’s tolerability also requires further examination.

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dSLIM. Mologen and codeveloper SanoÞ-Aventis are investigating the potential of double stem-loop immunomodulating oligodeoxyribonucleotides (dSLIM) as a potential treatment for CRC. The technology uses Mologen’s proprietary Minimalistic Immunogenically DeÞned Gene Expression (MIDGE) vectors to deliver the oligonucleotides; Mologen hopes that these vectors will be safer than conventional vectors because they contain only the genetic message that is to be introduced into the patient, not the additional DNA material contained in conventional vectors. In July 2004, Mologen licensed rights to dSLIM to the Chinese biotechnology corporation Starvax for development in East Asian countries. A Phase II German trial of dSLIM in CRC was completed in July 2003. The Phase II trial compared the safety and efÞcacy of chemotherapy with or without dSLIM in 17 mCRC patients. Of the 9 patients treated with dSLIM, 4 patients showed total remission of disease and 4 exhibited disease stabilization. One patient experienced disease progression during dSLIM treatment. The efÞcacy of dSLIM is being evaluated in an ongoing analysis of the trial’s immunological data (Mologen, press release, July 4, 2003). No additional clinical data have been reported. Although the Phase II data are positive, only nine patients received dSLIM in this trial; large-scale, randomized trial data are required for a clearer picture of the commercial and therapeutic potentials of this immunotherapy. Cytotoxic Agents Overview. Cytotoxic anticancer drugs are collectively the largest and most established chemotherapy group. The main candidates in the cytotoxic agents class are discussed in the following sections. Because SuperGen’s rubitecan was found to be clinically inactive in a Phase II trial in mCRC patients who had not been pretreated (Schoffski P, 2002), we do not discuss this agent herein. R&D strategies are focusing on new generations and/or formulations of standard therapies that provide advantageous efÞcacies or toxicity proÞles. Drugs with new, improved methods of administration may yield commercial success, but a major barrier will be higher costs compared with their unmodiÞed counterparts. Mechanism of Action. The fundamental aim of cytotoxic agents is to exert selective toxicity toward cancer cells, but they do so via a variety of pathways. Several mechanisms of action have been investigated, but none have generated strikingly positive results. The only signiÞcant pathway in terms of commercial potential appears to be the classical pathway of thymidylate synthesis (TS) inhibition. By inhibiting TS, these agents block normal DNA synthesis, thereby stopping cell replication. The newer agents directed toward this pathway have not yet demonstrated any signiÞcant advantages over available TS inhibitors, however. Pemetrexed. Eli Lilly is developing the multitargeted antifolate/antimetabolite compound pemetrexed (Alimta) (Figure 11) for the treatment of several cancers, including CRC. Phase II trials are ongoing in the United States and Europe. The

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FIGURE 11. Structure of pemetrexed.

compound exerts its antitumor activity by inhibiting at least three enzymes in the folate pathway, including the classic target of inhibition, TS. Pemetrexed as a single agent is approved in the United States and Europe for the treatment of locally advanced or metastatic NSCLC in patients who have failed at least one other chemotherapy regimen. Pemetrexed is also approved in combination with cisplatin for Þrst-line treatment of unresectable mesothelioma. Phase II data were presented at the ASCO meeting in 2003. One trial evaluated pemetrexed in combination with oxaliplatin as a Þrst-line mCRC treatment (Atkins JN, 2003). Of the 54 patients evaluable for toxicity, 23% developed grade 3 or 4 neutropenia, and 3.7% experienced grade 3 febrile neutropenia. Grade 4 thrombocytopenia was seen in 1.9%. No episodes of grade 3 or 4 diarrhea were observed. Grade 4 nonhematologic toxicity was evident in 5.6% of patients. Early deaths thought to be disease-related occurred in 3.7% of patients, but a relationship to treatment could not be entirely excluded. Of the 47 patients evaluable for response, 23% achieved an objective response: 1 complete response and 10 partial responses. Another trial evaluated pemetrexed plus carboplatin and pemetrexed plus oxaliplatin as Þrst-line treatments in 80 mCRC patients (Scagliotti GV, 2003). Both regimens achieved objective response rates in the 30% range, and median survival was close to 10 months for both regimens. Hematologic grade 4 toxicities were greater in the pemetrexed/carboplatin regimen: neutropenia, thrombocytopenia, and anemia occurred in 26%, 18%, and 8% of patients, respectively, compared with 7%, 2%, and 2%, respectively, of patients treated with the pemetrexed/oxaliplatin regimen. A different range of nonhematologic grade 4 toxicities was evident with each regimen, but these toxicities were generally rare. An ongoing Phase II study is evaluating pemetrexed, alone and in combination with irinotecan, as a second-line therapy in CRC patients with 5-FU–refractory disease (de Gramont A, 2002). A Phase I dose-escalation study showed that this combination is well tolerated up to a dose of 500 mg/m2 pemetrexed and 300 mg/m2 irinotecan (Kroening H, 2003). No further update has been reported for pemetrexed in CRC. Pemetrexed’s efÞcacy is similar to that observed for other available, intravenously administered single agents. It does not appear to offer any advantages over such marketed agents as AstraZeneca’s raltitrexed (Tomudex), an established TS inhibitor.

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Edotecarin. Banyu and PÞzer were codeveloping edotecarin, a topoisomerase I inhibitor, following in the footsteps of the already established agent within this class, irinotecan. In contrast to irinotecan, however, edotecarin is a noncamptothecin, cell-cycle-independent topoisomerase I inhibitor that shows an extended in vitro/in vivospectrum of activity, therapeutic index, and pharmacodynamic half-life compared with irinotecan. In April 2005, PÞzer discontinued studies of edotecarin and returned all rights to Banyu, owing to the agent’s insufÞcient activity as demonstrated in the different cancers studied. These new data, however, have not been reported. Earlier Phase II data were disclosed at the ASCO meeting in 2003 (Nahum K, 2003). The study was performed to determine the activity and safety of singleagent edotecarin in patients with irinotecan-naive mCRC. Twenty-four patients received 134 cycles of edotecarin, with a median of 4 cycles; 83% of the patients had previously received ßuoropyrimidine-based therapy. Three patients (13%) had a partial response, and 13 patients (57%) had stable disease. Median time to tumor progression was 7.1 months. Survival data are not yet available. The only grade 3 or 4 edotecarin-related adverse event was febrile neutropenia, occurring in one patient (4.2%); no edotecarin-related deaths were reported. One patient discontinued edotecarin owing to toxicity. Less-convincing Phase II data on edotecarin in irinotecan-naive or -refractory mCRC patients were presented at the previous year’s ASCO meeting (Perez RP, 2002). In 32 treated patients, neutropenia was the predominant adverse effect (grade 3 or 4 in 8 patients). Other, infrequent adverse effects included abdominal pain (grade 3 or 4 in 3 patients) and grade 3 diarrhea (2 patients), nausea (1 patient), vomiting (1 patient), and fatigue (1 patient). Although no irinotecan-refractory patients met the criteria for partial response, 5 of 18 patients experienced tumor shrinkage, and the estimated median time to progression (six months) appeared to be prolonged. Of 15 irinotecan-naive patients, 2 experienced a sustained, greater than 75% reduction in tumor mass, and 2 patients had a sustained, greater than 50% decrease in carcinoembryonic antigen (CEA, a CRC marker). Given the low incidence of grade 3 and 4 vomiting and neutropenia and the absence of grade 4 diarrhea, edotecarin may offer an improved therapeutic index over irinotecan, but large-scale randomized data are required before edotecarin’s efÞcacy and side-effect proÞle can be properly assessed. Aroplatin. Antigenics’ aroplatin (Platar) is a third-generation, platinumcontaining compound similar to the established agent oxaliplatin but formulated in a liposomal particulate carrier. A Phase II trial is ongoing in the United States. Antigenics recently developed an improved formulation of aroplatin intended to produce better pharmacological activity. The new formulation has entered a Phase I clinical trial by the end of 2005 in solid tumors and b-cell lymphoma. Aroplatin may employ a different mechanism of action than current platinumbased chemotherapeutics and therefore may be useful in cancers that are already resistant to platinum agents. In addition, liposome encapsulation has been shown

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to increase a drug’s bioavailability, which can extend the treatment’s effect. In some cases, liposomal drugs have been shown to accumulate at the site of a tumor, delivering higher concentrations of the drug to a disease target. The liposomal delivery system can also reduce drugs’ damaging effects on healthy tissues. Preliminary Phase II data were disclosed at the 2004 Gastrointestinal Cancers Symposium (Dragovich T, 2004). This single-arm, open-label trial evaluated aroplatin in patients whose disease was not responding standard treatments. One out of 15 patients showed a partial response, and two achieved stable disease. Even if this compound demonstrates advantageous efÞcacy over oxaliplatin, its higher cost will be an obstacle to its launch. Gimatecan. Novartis and Sigma-Tau’s gimatecan (ST-1481, LBQ-707) is an oral, camptothecin-derivative topoisomerase I inhibitor in development for the potential treatment of tumors, including CRC. Researchers from Italy and Switzerland are investigating gimatecan in Phase II trials. Results of this Phase II study were revealed at the 2004 ASCO meeting (Boni C, 2004). Gimatecan was administered at two dose levels to 18 mCRC patients, with 9 patients in each arm: 5.6 mg/m2 or 5.0 mg/m2 , orally. Of the six patients evaluable for response, 1 achieved a partial response, 3 had stable disease, and 2 had progressive disease (Boni C, 2004). Myelosuppression was the dose-limiting toxicity in previous Phase I clinical studies of gimatecan. The Phase II study is ongoing to evaluate the potential antitumor activity of gimatecan. Microtubule Modulators Overview. Microtubules, major structural components in cells, are the target of a large and diverse group of anticancer drugs, including taxanes and epothilones. Taxanes are potent antitumor agents; members of this class are marketed for numerous forms of cancer, but to date are unproven in CRC. PG-TXL (Xyotax, a poly-[L-glutamic acid]-conjugate of paclitaxel) is being developed by Cell Therapeutics, under license from the M.D. Anderson Cancer Center in Houston and in collaboration with Chugai. However, data are scarce for this agent in CRC. Epothilones are also being investigated for the treatment of CRC. Owing to lack of data, we do not discuss any of these agents in the following sections. Novartis’s epothilone B (Patupilone [EPO 906]) is in a Phase II trial for solid tumors, including CRC. In June 2004, Kosan and Roche discontinued a Phase II CRC study with epothilone D (KOS-862) because of unacceptable toxicities, so we do not discuss this agent in detail. Two agents, DJ-927, an oral taxane derivative, and ABT-751, a tubulin antagonist, are detailed in the following section. Mechanism of Action. Microtubules are protein polymers that are responsible for various aspects of cellular shape and movement. The major component of microtubules is the protein polymer tubulin. Microtubule modulators, like taxanes and epothilones, promote microtubule polymerization and inhibit tubulin depolymerization, arresting mitotic cell division at the metaphase/anaphase transition and inducing cell death.

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Considerable evidence indicates that at lower concentrations, agents in this class have a common mechanism of action: they suppress the dynamics of microtubules without appreciably changing the mass of microtubules in the cell. The drugs bind to diverse sites on the tubulin and at different positions within the microtubule, and they have diverse effects on microtubule dynamics. However, by their common mechanism of suppressing microtubule dynamics, they all block mitosis at the metaphase/anaphase transition and induce cell death. DJ-927. Daiichi Pharmaceutical’s DJ-927 is an oral taxane derivative under development for the potential treatment of cancer. DJ-927 is a tubulin depolymerization inhibitor that causes apoptosis and DNA cell-division arrest. Phase II trials are ongoing in the United States and Europe and Phase I trials in Japan. In preclinical studies, DJ-927 was found to be more potent than either paclitaxel or docetaxel (Syed SK, 2004). Preliminary results of a Phase II trial of DJ-927 as a second-line treatment for CRC were disclosed at the 2005 ASCO meeting (Rhee JM, 2005). DJ927 was administered as a single oral dose on day 1 of a 21-day cycle. Of the 15 patients enrolled with advanced CRC who had failed oxaliplatin- or irinotecan-based chemotherapy, 8 were evaluable for response, and all patients were evaluable for toxicity. DJ-927 demonstrated partial responses in 2 patients (25%), and 3 patients had stable disease (37.5%). Neutropenia was the main toxicity observed in this study, with grade 3 or 4 occurring in 7 of the 15 patients (47%). Preliminary results indicate that DJ-927 has some activity as second-line treatment for advanced CRC in patients refractory to oxaliplatin- or irinotecan-based chemotherapy (Rhee JM, 2005). ABT-751. ABT-751, an oral antimitotic tubulin antagonist, is in a Phase I trial conducted by Eisai in Japan and in a Phase II trial by Abbott Laboratories in the United States for the potential treatment of solid tumors. ABT-751 inhibits the polymerization of microtubules. This disruption of microtubule dynamics blocks cell division, resulting in apoptosis. Despite promising preclinical and Phase I data on single-agent ABT-751 presented at the 2003 ASCO meeting (Sprague E, 2003), the agent was not active in a Phase II trial. ABT-751 failed to show antitumor activity in a recent multicenter, Phase II study in patients with multiple refractory mCRC. Results of the Phase II study in patients with refractory mCRC were presented at the 2005 ASCO meeting (Benson AB, 2005). Patients received 200 mg/day oral ABT-751 for 21 days followed by 7 days of rest. The study evaluated the safety and efÞcacy of ABT751 in 49 mCRC patients refractory to irinotecan and oxaliplatin. ABT-751 was well tolerated in CRC patients. After a median duration of two cycles (range 1–8), median time to progression was 2.1 months, and no complete or partial responses were observed at the time of the presentation (Benson AB, 2005). Cyclooxygenase-2 Inhibitors Overview. Cyclooxygenase-2 (COX-2) inhibition is a well-established chemopreventive pathway, most notably in CRC; celecoxib (PÞzer’s Celebrex) has

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already been approved in the United States (and is awaiting European approval) as an adjuvant to surgery for treating familial adenomatous polyposis (FAP), a precursor to CRC. FAP is a rare condition, however, so celecoxib is infrequently used as a preventive measure, and few patients are routinely identiÞed as being eligible for chemoprevention with drugs from this class. Because of the recent scrutiny of these agents in terms of cardiovascular events, and their subsequent withdrawal for many indications, the established chemopreventive role of COX-2 inhibition and relative inexpensiveness of COX-2 inhibitors will likely no longer be sufÞcient to enable their widespread use in a healthy population. The COX-2 inhibitor class came under intense scrutiny when these agents were found to be associated with increased cardiovascular events in a CRC study. Rofecoxib (Merck & Co.’s Vioxx) was found to increase the risk of cardiovascular events in patients with a history of colorectal adenomas. In September 2004, Merck announced a worldwide withdrawal of rofecoxib, and all studies were suspended. In addition, a study investigating PÞzer’s COX-2 inhibitor celecoxib (Celebrex) in cancer patients was halted because of data suggesting that celecoxib increased patients’ risk of myocardial infarction and stroke. Although celecoxib remains on the market, in August 2005, the FDA added warnings to its packaging. Although not used for CRC, valdecoxib (PÞzer’s Bextra) is another COX-2 inhibitor that was also withdrawn after an FDA review concluded that this agent poses unacceptable cardiovascular risks and the risk of potentially life-threatening, sometimes fatal skin reactions but offers “no demonstrated advantage compared with other NSAIDs [nonsteroidal anti-inßammatory drugs].” These side-effect issues have led to considerable uncertainty about the future of COX-2 inhibitors. We include them herein because recent trial data in mCRC were presented at the 2005 ASCO meeting. Mechanism of Action. The mechanism of anticancer activity of COX-2 inhibitors is not fully understood. COX-2 is expressed in most CRCs. Thus, high levels of COX-2 may promote tumorigenesis through enhanced angiogenesis, increased tumor invasiveness, resistance to apoptosis, and lowered host-immune surveillance. Celecoxib. Celecoxib (PÞzer’s Celebrex) (Figure 12) received FDA approval as an adjuvant treatment to surgery for FAP in December 1999. No randomized data are available that unequivocally demonstrate celecoxib’s efÞcacy in CRC treatment. Inconclusive Phase II results presented at the 2003 ASCO meeting (Pan C, 2003) were generated from a trial that examined the combination of the IFL regimen plus celecoxib as a Þrst-line therapy for mCRC. Of the 41 patients treated, 38 were evaluable for toxicity. No episodes of grade 4 diarrhea were recorded, but 16 patients (42.1%) had grade 3 diarrhea; 9 patients (23.7%) had grade 3 granulocytopenia; and 4 patients (10.5%) had grade 4 granulocytopenia. Dose modiÞcations were required for 27 of 38 patients. Two treatment-related deaths (GI bleeding/aspiration; possible pulmonary embolism) were reported. Of the 31 patients evaluable for response, 2 experienced complete responses, 11 had

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O

H2N S O

N

N CF3

H3C FIGURE 12. Structure of celecoxib (R1 = NH2 , R2 = CH3 ).

partial responses, 9 had stable disease, and 9 had progressive disease (overall response rate = 41.9%). The 12-month overall survival rate was 57%. Because other agents in its class (i.e., rofecoxib, valdecoxib) have been withdrawn from the market, experts are increasingly concerned about the risks associated with long-term celecoxib treatment. Although celecoxib remains on the market, in April 2005, the FDA requested that all prescription NSAIDs marketed in the United States, including celecoxib, add a boxed warning to their labels that highlights the potentially increased risk of adverse cardiovascular events and GI bleeding. In August 2005, the FDA added the expected warnings to the celecoxib label. Data from the Adenoma Prevention with Celecoxib (APC) study provide evidence for an increased cardiovascular risk (Solomon SD, 2005). The APC study was a large, randomized, placebo-controlled, double-blind, multicenter trial that assessed the efÞcacy of celecoxib for the prevention of adenomatous polyps. The study compared the efÞcacy and safety of celecoxib at 200 mg twice daily and at 400 mg twice daily with placebo for reducing the occurrence of adenomatous polyps in the colon and rectum one year and three years after endoscopic polypectomy in 2,035 patients. After follow-up of at least 2.8 years of surviving patients, the study found that treatment with celecoxib (200 or 400 mg twice daily) to prevent colorectal adenomas instead led to a dose-related increase in the risk of serious cardiovascular events, including death, myocardial infarction, stroke, and heart failure (Solomon SD, 2005). Furthermore, a European Organization for Research and Treatment for Cancer (EORTC) randomized, double-blind, multicenter, four-arm, Phase III study assessing the addition of COX-2 inhibitors to chemotherapy in patients with advanced CRC was suspended after eight fatal events that did not result from progressive disease (Kohne C, 2005). All these results buttress the Þndings that COX-2 inhibitors are associated with increased cardiovascular risks. However, data from a small U.S. Phase II trial presented at the 2005 Gastrointestinal Cancers Symposium showed promising activity for capecitabine/irinotecan in combination with celecoxib in 32 previously untreated (apart from adjuvant therapy) patients with mCRC. On an intent-to-treat basis, objective response and stable disease were observed in 41% and 31% of patients,

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respectively (El-Rayes BF, 2005). The median progression-free survival was 7.1 months, and the median duration of response was 6.5 months. No disease progression was seen in 19 patients, and no grade 4 toxicities or treatmentrelated deaths were observed. The only grade 3 toxicities noted were diarrhea (32%), hand-and-foot syndrome (6%), nausea (16%), vomiting (13%), and neutropenia (9%). Researchers concluded that celecoxib can safely be administered in combination with irinotecan and capecitabine. Based on the progression-free survival and response rates observed in this study, the regimen was considered to have promising activity (El-Rayes BF, 2005). Even so, a large number of studies involving COX-2 inhibitors have either been terminated or put on hold, making their role in CRC prevention highly uncertain. At the time of composing this reference, it appears that celecoxib is still being investigated, but primarily in advanced disease, rather than for early disease or as a preventive. Rofecoxib. At the time of composing this reference, all studies involving Rofecoxib (vioxx) appear to have been terminated.

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Malignant Melanoma

ETIOLOGY AND PATHOPHYSIOLOGY Melanin is the pigment that gives skin its color and protects it against the damaging effects of ultraviolet radiation. Melanoma is an invasive neoplasm of the melanocytes—melanin-producing cells. Melanocytes are derived from the neural crest and migrate to various parts of the body during early embryonic life. Although most commonly located in the basal layer of the epidermis, melanocytes are also found in the eyes, respiratory tract, gastrointestinal tract, and the mucous membranes of the genitalia and mouth. Most melanomas occur in the skin, but they may occur at any site to which melanocytes have migrated. In women, melanoma occurs most frequently on the extremities; in men, melanoma is most common on the trunk, head, and neck. Melanoma may originate in normal skin or from a pre-existing nevus (mole)—see the section “Common Nevi” below. Approximately 90% of melanoma cases are sporadic; about 10% are familial. Ultraviolet radiation (UVA and UVB) is the only known causative agent for the development of melanoma. Risk factors include high intermittent sun exposure (particularly risky for pale-skinned Caucasians, especially freckled redheads); a high number of common nevi (more than 100); a high number of atypical nevi (more than ten); blond hair and blue eyes; use of artiÞcial tanning equipment; residence at equatorial latitudes; having a relative with malignant melanoma (MM); previous malignant melanoma; and previous organ or marrow transplantation. The latter risk factor is related to the immunosuppression associated with these procedures. Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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Common Nevi Common nevi are benign clusters of melanocytes that may be present at birth or appear during childhood or early adulthood. Nevi on the skin are commonly called moles. The number of nevi normally reaches a maximum in the third decade of life (the average Caucasian person has approximately 15) and then gradually diminishes. Estimates for the annual transformation rate of any single mole into melanoma range from 0.0005% (fewer than 1 in 200,000) for both men and women younger than 40 years to 0.003% (approximately 1 in 33,000) for men and women older than 60 years (Tsao H, 2003[b]). The different types of nevi are distinguished by features that may have a bearing on the pathogenesis of melanoma: •

•

•

Junctional nevi are small, ßat, brown lesions that occur at the junction of the dermis and epidermis. Malignant transformation of junctional nevi is possible but rare. Compound nevi are larger than junctional nevi; they are usually raised and contain coarse hair. They result from the proliferation of junctional nevi and their penetration into the dermis. The frequency of malignant transformation of compound nevi is difÞcult to ascertain, since data on the prevalence of nevi are not captured in databases. Intradermal nevi result from continued multiplication of the deep dermal cells of compound nevi. These nevi—also known as acquired common nevi —generally have little pigmentation and are raised. They have no known chromosomal aberrations. The frequency of malignant transformation of intradermal nevi is unknown.

Atypical and Dysplastic Nevi An atypical nevus is deÞned clinically as a ßat macule at least 5 mm in diameter that has at least two of the following features: asymmetric outline, variable color, and indistinct borders. Atypical nevi are re-classiÞed as dysplastic nevi when histological examination reveals cytogenetic changes. Dysplastic nevi can develop from acquired common nevi. Most dysplastic nevi are stable, or even regressive, and do not progress to melanoma. Those with dysplastic nevi have a risk of developing melanoma that is 27 times the risk of the general population (Kraemer KH, 1985). Disease Progression Histological analysis has deÞned Þve steps in the development and progression of melanoma. The Þrst step is the presence of acquired or congenital common nevi with normal melanocytes. Nevi development is thought to occur when melanocyte-keratinocyte communication is disrupted, allowing melanocytes to escape from keratinocyte regulatory control (in unmarked skin, the presence of keratinocytes prevents melanocytes from expressing melanoma-associated antigens).

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The second and third steps are the progression to atypical and dysplastic nevi and then to radial growth. These events are believed to result from poorly understood genetic mutations. A local immune response that probably inßuences disease outcome is present during both the dysplastic and the radial growth phases. The radial growth phase is characterized by tumor cells in the epidermis producing an irregular surface on the skin. Although the melanoma at this step may be invasive within the dermis, it is unlikely to metastasize to either the regional lymph nodes or beyond. The fourth step is progression to vertical growth. Melanomas in the vertical growth phase penetrate deep into the dermis, increase their vasculature, and elicit a diminished host immune response. During this step, the melanoma has great potential to metastasize. The Þfth and Þnal step is the development of metastases. Figure 1 illustrates the progression of melanoma. Adhesion Receptors As melanoma progresses through the Þve stages, the pattern of adhesion receptor expression changes (Table 1). Some adhesion receptors increase expression (e.g., N-cadherin, and intercellular adhesion molecule [ICAM]-1), and some decrease expression (e.g., E-cadherin, vascular cellular adhesion molecule [VCAM]-1). These changes affect the melanoma cells’ ability to adhere to and communicate with surrounding cells. For example, melanocytes require E-cadherin to establish gap junctions with keratinocytes that regulate melanocyte growth. Melanocytes that express low levels of E-cadherin cannot adhere to keratinocytes; because of the high-level expression of N-cadherin, however, they can establish gap junctions with Þbroblasts and endothelial cells, which may result in increased tumor vasculature (McGary EC, 2002). Growth Factors and Cytokines The expression of growth factors (proteins that direct cells to proliferate) and cytokines (proteins that coordinate the immune response) increases as melanoma progresses. A small number of these proteins increase their expression after the cells’ transition to nevus cells, but the most signiÞcant increases occur after the radial growth phase. The names and functions of the key growth factors upregulated in melanoma cells are shown in Table 2. The cytokines interleukin-8 (IL-8) and monocyte chemoattractant protein1 (MCP-1) are produced by melanoma cells and elicit an immune response, attracting neutrophils and monocytes, respectively. Moderate production of IL-8 and MCP-1 can facilitate tumor growth by stimulating production of angiogenic cytokines (e.g., tumor necrosis factor-alpha [TNF-α]) or by stimulating stroma formation; high production can elicit an immune response strong enough to kill the malignant cells. Melanoma Subtypes Several subtypes of melanoma have been classiÞed according to their distinct presentations, patterns of progression, and prognoses. The three major subtypes,

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FIGURE 1. Progression of malignant melanoma.

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TABLE 1. Change in Expression of Adhesion Receptors During Melanoma Progression Adhesion Receptors Increasing

Adhesion Receptors Decreasing

α2(β 1) α4(β 1) (αv)β 3/αIIbβ 3 Mel-CAM ICAM-1 N-cadherin CSPG

E-cadherin (αv)β 5 α 6(β 1) VCAM-1

Adhesion Receptors Unchanged α5β 1 CD44

CSPG = Chondroitin sulfate proteoglycans. ICAM = Intercellular adhesion molecule. VCAM = Vascular cell adhesion molecule.

which account for more than 90% of all melanomas, are superÞcial spreading melanoma, nodular melanoma, and lentigo maligna melanoma. Features of these and less common subtypes are shown in Table 3. Melanoma-Associated Antigens Melanoma is considered the most immunogenic of the cancers. The identiÞcation of tumor-associated antigens speciÞc to melanoma, recognized by T lymphocytes (T cells), has opened the door to a new era in the immunotherapy of this disease, as well as the possibility of developing melanoma vaccines that incorporate melanoma-associated antigens. These vaccines are the subject of widespread investigation. A melanoma-associated antigen can be classiÞed into one of four types: • • • •

Unique tumor antigens—antigens speciÞc to individual patients. Differentiation antigens—expressed in melanoma and in normal melanocytes. Shared tumor antigens (also known as cancer testis antigens)—expressed in several tumors and the testes. Gangliosides (acidic glycoproteins)—the most extensively expressed antigens on the melanoma cell surface, also found in other tissues, including nervous system tissue and the spleen.

Examples of these antigens are shown in Table 4. The Genetics of Melanoma The genetics of melanoma are poorly understood. One gene with an established role in familial cases is CDKN2A, located on chromosome 9p21. The protein product of the CDKN2A gene, p16(INK4a), downregulates the pathway that controls entry of the cell into the S phase of the cell cycle by binding to CDK4 and cyclin D complexes, thus blocking kinase activity (Sherr CJ, 1999; Rutter JL, 2003). Germ-line CDKN2A mutations are present in approximately 20–25% of melanoma-prone families; these mutations are comparatively rare in families with only two or three cases. Individuals and families from Scotland and France

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TABLE 2. Growth Factors and Cytokines Upregulated in Malignant Melanoma Growth Factors

Role

Comments

Basic fibroblast growth factor (bFGF)

A survival factor, growth stimulator, and motility factor. Binds to matrix proteins and can stimulate fibroblasts and endothelial cells. Upregulates serine proteinases and metalloproteinases. Stimulates mitosis in tumor-infiltrating fibroblasts. Induces production of matrix proteins (e.g., collagen) that provide a structure for adhesion by melanoma cells. A family of at least three closely related proteins (TGFβ 1−3). Involved in tissue remodeling, wound repair, and angiogenesis, together with inhibition of T-cell proliferation. Stimulates angiogenesis: a gateway to rapid proliferation and metastasis. An exogenous growth factor produced by activated fibroblasts. Highly mitogenic. Also stimulates survival by stabilization of beta-catenin.

An autocrine growth factor. Blocking bFGF with antisense oligonucleotides has been shown to halt tumor growth.a

Platelet-derived growth factor

Transforming growth factor-beta (TGFβ)

Vascular endothelial growth factor (VEGF) Insulin-like growth factor-1 (IGF-1)

Cytokines Interleukin-8 (IL-8)

Monocyte chemoattractant protein-1 (MCP-1)

Attracts monocytes.

Attracts monocytes.

The most significant fibroblast-stimulating factor.

Raised levels of one of these proteins (TGFβ 2) are seen in widespread metastatic disease.

Presence of monocytes can stimulate tumor growth by producing angiogenic cytokines or triggering stroma formation, but high production of IL-8 can induce enough monocytes to kill tumor. Presence of monocytes can stimulate tumor growth by producing angiogenic cytokines or triggering stroma formation, but high production of MCP-1 can induce enough monocytes to kill tumor.

a Yang Y, Becker D. Antisense targeting of basic fibroblast growth factor and fibroblast growth factor

receptor-1 in human melanomas blocks intratumoral angiogenesis and tumor growth. Nature Medicine. 1997;3:887–893.

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TABLE 3. Characteristics of Malignant Melanoma Subtypes Morphotype Superficial spreading

Nodular

Lentigo maligna melanomas Acral lentiginous

Desmoplastic melanoma

Clinical Features at Presentation Usually asymptomatic, larger lesions 2–3 cm in diameter. Flat or slightly raised plaque. Color variation (black/brown) and areas of depigmentation (often a pale central area) are common. Raised nodule on the skin. Often bleeds and is friable or ulcerated. Has almost always deeply penetrated the dermis by time of diagnosis. Similar to superficial spreading melanoma. Flat pigmented area on the skin.

No distinctive clinical features.

Common Sites

General Comments

Among men, the trunk is the most common site; among women, the feet or lower legs are. Can occur at any site.

The most common type, accounting for 60–70% of malignant melanoma cases. Chiefly occurs in middle-aged people. Typically spreads slowly (several years) over the superficial skin layer before penetrating deeper layers.

Areas of skin not exposed to light often affected.

Associated with early metastasis.

Face

Usually occurs in the elderly.

Non-hairy skin: palms, soles, and beneath nails.

More common in nonwhites. Grows slowly over many years (radial phase). Invasive vertical growth occurs later as nodules within the lesion. Diagnosis is often delayed, worsening prognosis. A rare and locally aggressive form. Tumor may be amelanotic. Histological features include aggregates of lymphocytes and stromal myxoid change.

Head and neck.

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TABLE 4. Malignant Melanoma-Associated Antigens Antigen Unique Differentiation Shared (cancer testis antigens) Gangliosides

Example CDK4, β-catenin, Mum-1b44, CDC27 Tyrosinase, gp100, Melan-A/MART-1, TRP-1 MAGE, BAGE, GAGE families, NY-ESO-1 GM2, GD2, GD3

are most likely to have germ-line CDKN2A mutations, and individuals from the United States and Australia are least likely (Pollock PM, 2000 and 2002). Other genes implicated as inheritable determinants of familial melanoma are the CDK4 gene, which has a lower penetrance than CDKN2A (Goldstein AM, 2000) but also interacts with p16(INK4a), and the CMM1 gene, where conßicting data exist for its role in families at high risk of melanoma. The understanding of the genetic basis of sporadic (nonfamilial) melanoma was advanced in 2002 by the discovery at the Human Cancer Genome Centre (Cambridge, United Kingdom) of mutations in the Braf gene (Davies H, 2002). Braf mutations were identiÞed in 66% of the melanoma samples studied by the group. BRAF, the protein kinase product of Braf, mediates a pathway of Ras signaling. Evidence supports a biochemical pathway in which Ras activates BRAF, which in turn activates the mitogen-activated protein kinases The authors showed that mutations in Braf lead to increased kinase activity, leaving the “growth” signal always turned on. Correction of this aberrant pathway is the subject of ongoing research. Cytogenetic studies and loss of heterozygosity (LOH) studies have identiÞed six chromosomal regions likely to contain genes involved in melanoma development: 1p, 6p, 7p, 9p, 10q, and 11q. Two other genes are known to participate in the development of sporadic melanoma: CDKN2A, mapping to chromosome 9p21, and PTEN, mapping to 10q23 (Casula M, 2003). Mutations in CDKN2A have been observed in only 8% of uncultured tumor samples; the percentage is higher in cell lines. A PTEN mutation is present in one-third of cultured melanoma cell lines but only 5% of uncultured tumors and can result in abnormal or loss of PTEN protein expression. The PTEN protein exhibits lipid phosphatase and protein phosphatase activity and is believed to affect cell survival and proliferation by regulating response to growth factor stimulation. PTEN may also help modulate cell migration and invasion by interacting with focal adhesion kinase to regulate the signals generated at focal adhesions (Pollock PM, 2000; Tsao H, 2003[b] Ongoing genome-wide scans at the National Human Genome Research Institute may lead to identiÞcation of other genes that play a role in predisposition to melanoma and in the development of sporadic melanoma. Staging of Melanoma Several systems are currently used to stage melanomas, each focusing on one or more key prognostic factors: Breslow’s ClassiÞcation, which measures only tumor thickness (Table 5); Clark’s ClassiÞcation, which measures only degree

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TABLE 5. Breslow’s Classification of Malignant Melanoma Breslow’s Classification (thickness) ≤0.75 mm or less >0.75 mm to 1.50 mm >1.50 mm to 4.0 mm >4.0 mm

TABLE 6. Clark’s Classification of Malignant Melanoma Clark’s Classification (level of invasion) Level I Level II Level III Level IV Level V

Characteristics Lesions involve only the epidermis (in situmelanoma); not an invasive lesion Invasion of the papillary dermis but not the papillary-reticular dermal interface Invasion fills and expands the papillary dermis but does not extend into the subcutaneous tissue Invasion of the reticular dermis but not the subcutaneous tissue Invasion through the reticular dermis into the subcutaneous tissue

of invasion (Table 6); and the American Joint Committee on Cancer (AJCC) scheme, which uses the tumor node metastases (TNM) system to assess nodal and metastatic involvement. The AJCC classiÞcation also accounts for tumor thickness, level of invasion, and ulceration (Table 7) and groups the TNM categories into four stages (Table 8). A revised staging system was approved by the AJCC in 2001 and became ofÞcial with publication of the 2002 (6th ) edition of the AJCC Cancer Staging Manual (Balch CM, 2001[b]), with the recommendation that all patients diagnosed starting in January 2002 should be classiÞed within the new system. Please note that at the time of composing this reference, this remains the most up-to-date edition. The new staging system was developed by the AJCC Melanoma Staging Committee and was validated using data from a cohort of 17,600 patients (Balch CM, 2001[a]). This analysis revealed that, among patients without nodal or metastatic involvement, tumor thickness and ulceration were the most important prognostic factors: an ulcerated tumor behaves like a biologically aggressive tumor. Among patients with nodal involvement, three independent prognostic factors were evident: the number of positive nodes, whether nodal involvement was micrometastatic or clinically apparent, and the presence or absence of ulceration. Among patients with distant metastases, the site of involvement was the most signiÞcant prognostic factor. Because of these discoveries, the AJCC made the following key changes to its 1997 staging system: •

Primary tumor classiÞcation regards only thickness and ulceration; level of invasion is considered only in the case of thin (T1) melanomas.

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TABLE 7. AJCC Malignant Melanoma TNM 2002a Classification TNM Definitions Tis T1

Melanoma in situ (Clark’s level I) ≤1.0 mm a: without ulceration and Clark’s level II/III b: with ulceration or Clark’s level IV/V 1.01–2.0 mm a: without ulceration b: with ulceration 2.01–4.0 mm a: without ulceration b: with ulceration >4.0 mm a: without ulceration b: with ulceration No regional lymph node metastasis 1 node a: micrometastasesb b: macrometastasesc 2–3 nodes a: micrometastases b: macrometastases c: in transit metastases/satellite(s) without metastatic nodes 4 or more metastatic nodes, or matted nodes, or in transit metastases/ satellite(s) with metastatic nodes(s) No distant metastasis a: distant skin, subcutaneous, or nodal metastases, serum LDH normal b: lung metastases, serum LDH normal c: all other visceral metastases, LDH normal; any distant metastasis, elevated LDH

T2

T3

T4

N0 N1

N2

N3 M0 M1

a The AJCC Executive Committee has approved the final version of the melanoma staging system. It became

official with the publication of the sixth edition of the AJCC Cancer Staging Manual in 2002. b Micrometastases are diagnosed after sentinel or elective lymphadenectomy. c Macrometastases are defined as clinically detectable nodal metastases confirmed by therapeutic lym-

phadenectomy or when nodal metastasis exhibits gross extracapsular extension. AJCC = American Joint Committee on Cancer. LDH = Lactate dehydrogenase. TNM = Tumor, node, metastasis

• • • •

Nodal classiÞcation considers the number of involved nodes rather than their dimensions. Nodal classiÞcation distinguishes between microscopic and macroscopic nodal involvement. Metastasis classiÞcation considers the site of metastases and the presence of serum lactate dehydrogenase. All patients with ulceration are upstaged

The revised staging system will probably lead eventually to changes in the treatment of current stage III patients (nodal involvement or primary tumor more than 4 mm thick and/or satellites within 2 cm of the primary). Stage III patients have generally been considered a high-risk group that requires aggressive treatment,

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TABLE 8. Revised AJCC Groupings for Cutaneous Melanoma Stage Grouping 0 IA IB IIA IIB IIC III IIIA

Clinicala Tis, N0, M0 T1a, N0, M0 T1b, N0, M0 T2a, N0, M0 T2b, N0, M0 T3a, N0, M0 T3b, N0, M0 T4a, N0, M0 T4b, N0, M0 Any T, N1-3, M0

IIIB

IIIC

IV

Any T, any N, M1

Pathologicalb Tis, N0, M0 T1a, N0, M0 T1b, N0, M0 T2a, N0, M0 T2b, N0, M0 T3a, N0, M0 T3b, N0, M0 T4a, N0, M0 T4b, N0, M0 T1-4a, N1a, M0 T1-4a, N2a, M0 T1-4b, N1a, M0 T1-4b, N2a, M0 T1-4a, N1b, M0 T1-4a, N2b, M0 T1-4a/b, N2c, M0 T1-4b, N1b, M0 T1-4b, N2b, M0 Any T, N3, M0 Any T, any N, M1

a Clinical staging includes microstaging of the primary melanoma and

clinical/radiological evaluation for metastases. By convention, it should be used after complete excision of the primary melanoma with clinical assessment for regional and distant metastases. b Pathological staging includes microstaging of the primary melanoma and pathological information about the regional lymph nodes after partial or complete lymphadenectomy. Pathological stage 0 or stage IA patients are the exception; they do not require pathological evaluation of their lymph nodes.

but the recent AJCC analysis revealed that these patients are heterogeneous in terms of prognosis. The study found Þvefold differences in Þve-year survival, depending on the presence or absence of ulceration and the presence of clinically apparent occult metastases: patients with nonulcerated tumors with a single, clinically occult metastasis had a Þve-year survival rate of 69%, compared with 13% for patients with ulcerated melanomas and four or more clinically apparent nodal metastases. On the basis of these Þndings, AJCC investigators recommend that patients with nodal metastases should not be considered a single group and that clinical trials take into account that some stage III patients have a good prognosis. Figure 1 illustrates the progression of melanoma. Prognosis The prognosis of melanoma varies widely, depending on the stage of the disease. Most patients with tumors less than 2.0 mm in thickness, conÞned to the skin, are cured following surgical removal of the tumor (ten-year survival is more than

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TABLE 9. Five-Year Overall Survival of Patients with Cutaneous Melanoma Grouped by American Joint Committee on Cancer (AJCC) Stage Following Surgery Alone AJCC Stage Stage IA IB IIA IIB IIC IIIA IIIB IIIC

Surgery Alone (baseline prognosis) 95% 90% 77% 65% 45% 67% 53% 26%

85%). Five-year survival for patients with thicker tumors (greater than 2 mm) conÞned to the skin, with no ulceration, is approximately 65%. Patients with local lymph node involvement have a Þve-year survival rate of approximately 45%, depending on the extent of local disease (Table 9). The outlook for patients with metastatic disease is poor; median survival time is merely 6–12 months, with a Þve-year survival rate of approximately 10% depending on extent and location of metastatic disease. CURRENT THERAPIES Surgery is the mainstay of treatment for malignant melanoma (MM). Adjuvant drug therapy, when administered, is almost exclusively interferon-alpha-2 (IFNα-2). The treatment of metastatic malignant melanoma (mMM) is based largely on dacarbazine (Bayer’s DTIC-Dome, generics), and this agent is increasingly being combined, in biochemotherapy regimens, with biological response modiÞers, such as interleukin-2 (IL-2) and IFN-α-2, in preference to combining it with other cytotoxic agents. These and other current therapies for MM, however, do not adequately address this disease, and trial results for many of them are poor or inconsistent. IFN-α-2b (Schering-Plough’s Intron A) is the only therapy approved for MM patients who do not have metastases. It is used interchangeably with IFN-α-2a (Roche/Chugai’s Roferon A; Takeda’s Canferon A) in markets where both are available. High-dose IFN-α-2b is approved in the United States and Europe as adjuvant therapy for resected MM patients at high risk of relapse. Its application in the adjuvant setting remains controversial. Data from a meta-analysis of ten trials and approximately 3,700 patients conÞrm that IFN can delay time to disease progression (Wheatley K, 2001), but data demonstrating a convincing long-term survival beneÞt over observation are lacking. IFN use may improve quality of life (by extending disease-free survival); however, its lack of overall survival beneÞt needs to be weighed against both its considerable toxicity and its Þnancial cost.

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Chemotherapy is generally reserved for the treatment of stage IV (metastatic) disease. Response rates to single agents are modest (less than 25%), and the duration of response is short. In an attempt to improve outcomes, clinicians have combined chemotherapy (most notably dacarbazine) with biological agents (IFNα-2 and/or IL-2), a combination known as biochemotherapy. IL-2 is also approved for single-agent use in mMM. Overall, the efÞcacy of such combinations has not been encouraging, and data from individual trials are conßicting (Eigentler TK, 2003). The observed increase in median survival—from 6–9 months to 12 months in some studies—needs to be conÞrmed in large randomized studies if this approach is to be widely accepted. Further, the addition of biological agents to chemotherapy signiÞcantly increases toxicity, and this toxicity needs to be considered alongside any improvements in response rate and survival. Optimal treatment doses and schedules for these regimens need to be identiÞed, and these Þndings could increase the attractiveness of biochemotherapy. Like dacarbazine, fotemustine (Servier’s Muphoran) is an alkylating agent approved to treat mMM in France and Spain, where it may be used in place of dacarbazine. Fotemustine, like temozolomide, has the ability to cross the blood-brain barrier and therefore has activity against brain metastases of MM. Approximately 25% of MMs have demonstrable estrogen receptors, and this fact has provided the rationale for investigating the role of tamoxifen (AstraZeneca’s Nolvadex and generics) in the treatment of MM. After tamoxifen failed to demonstrate convincing activity as a single agent (Rumpke P, 1992), clinicians combined it with standard chemotherapy. However, despite early reports from small clinical trials of improved response rates and median survival (Cocconi G, 1992), larger randomized studies failed to prove the value of adding tamoxifen, and its regular use in MM is now largely obsolete (Rusthoven JJ, 1996; Rusthoven JJ, 1998). The disappointing Þndings with tamoxifen, however, may result from poor trial design and failure to select patients with estrogen-expressing tumors. Taxanes have been investigated for the treatment of MM. EfÞcacy in patients treated with single-agent paclitaxel (Bristol-Myers Squibb’s Taxol) was disappointing, with response rates of between 12% and 18% (Legha SS, 1990). A Phase II study with docetaxel (Aventis’s Taxotere) also produced low response rates, although the 2 (of 37) patients who responded positively maintained the response for more than two years (Einzig AI, 1996). Overall, the severe additional toxicity that taxanes cause in MM patients—particularly neutropenia—has not justiÞed their routine use; thus, despite modest use of these agents as second- or subsequent-line therapy in mMM, we do not discuss them in this reference. Three different treatment regimens—intralesional interferon-beta, DAVferon, and DacTam—are used exclusively in Japan. Because these regimens are not widespread and the incidence of MM is low in Japan, we do not discuss the clinical performance of these regimens. Isolated limb perfusion (ILP) has been applied in eligible patients; this procedure is used almost exclusively for patients with in-transit disease conÞned to a single extremity, a situation that is uncommon. ILP is a means of delivering

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chemotherapy to a local area at doses signiÞcantly higher than would be tolerable systemically. Melphalan (GlaxoSmithKline’s Alkeran, an alkylating agent) is the most commonly used agent and achieves response rates of approximately 50% (complete response) and 75% (partial response) (Rossi CR, 2004). Melphalan is delivered via canulation of the external iliac artery and vein (for perfusion of the lower extremity) or via the axillary vessels (upper extremity). Melphalan is delivered with hyperthermia (38.8–40.0◦ C) as a strategy to augment tissue concentrations of the drug. The optimal duration of therapy is unknown, but duration of approximately one to two hours is standard. Tumor necrosis factor (TNF) may also be used in combination with melphalan to improve response rates. Only two small, prospective trials, some still ongoing, seem to have evaluated the efÞcacy of ILP. Among some signiÞcant results achieved: 58% recurrence among stage III control patients versus 12.5% recurrence among treated patients (Ghussen F, 1986). These results have not been replicated, however, and are controversial. The second trial achieved an improved disease-free survival time (median time of 17 months in the treated group versus 10 months in the control group) but no overall survival beneÞt (Hafstrom L, 1991). Owing to the inadequate treatment of MM by current therapies and conßicting study data showing no apparent relationship between response rate and overall survival, many companies are investigating new approaches. In addition, prospective, randomized Phase III studies that evaluate existing regimens are urgently needed to clarify the optimal currently available treatment; randomized clinical trials, ideally trials that incorporate molecular measures of response, seem to be the best treatment option for MM patients at this time. Table 10 summarizes the current treatment regimens commonly used to treat MM, including their doses and toxicities. High-Dose Interferon-Alpha-2 Overview. High-dose interferon-alpha-2 (IFN-α-2) regimens are administered at doses of 10–20 MU/m2 . IFN-α-2a (Roche/Chugai’s Roferon A, Takeda’s Canferon A) and IFN-α-2b (Schering-Plough’s Intron A) tend to be used interchangeably in those markets where both are available, although IFN-α-2b is the only IFN with approval for this indication. We present the trial results for IFNα-2b because this agent is marketed for adjuvant use in high-risk stage II and stage III MM. Mechanism of Action. The IFNs are a family of naturally occurring cytokines that exhibit antiviral, antitumor, and immunomodulatory activities. The antitumor effect of IFNs derives from their ability to inhibit cell-cycle progression, induce a reduction in tumor-cell protein synthesis, and inhibit the formation of colonies of late progenitor cells. Their immunomodulatory role involves inducing the expression of major histocompatability antigens and modulating the expression and function of T cells, monocytes, and natural killer cells (large granular lymphocytes that bind to and kill cells by releasing cytotoxins).

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TABLE 10. Current Regimens/Classes Used for Malignant Melanoma Regimen or Class

Regimen Components Agent

Availability

Dose

Common Toxicities

Interferon-alpha-2 US, F, G, I, 20 MU/m2 IV (or IM) Granulocytopenia 5 days/week for 4 Leukopenia (Schering-Plough’s S, UK, J Fatigue weeks followed Intron A, by 10 MU/m2 SC Liver toxicity Roche/Chugai’s 3 times weekly Neurological Roferon A, for 48 weeks effects Takeda’s Canferon Infection A) Psychological disturbances Intermediate- Interferon-alpha-2 US, F, G, I, 10 MU SC Granulocytopenia dose (Schering-Plough’s S, UK, J 5 days/week for 4 Leukopenia interferonIntron A, weeks followed Fatigue alpha-2 Roche/Chugai’s by 5 MU SC 3 Liver toxicity Roferon A, times weekly for Neurological Takeda’s Canferon 24 months effects A) Infection Low-dose Interferon-alpha-2 US, F, G, I, 3 MU SC 3 times Granulocytopenia interferon(Schering-Plough’s S, UK, J weekly for Leukopenia alpha-2 Intron A, 6–36 months Fatigue Roche/Chugai’s Liver toxicity Roferon A, Neurological Takeda’s Canferon effects A) Infection Aldesleukin Aldesleukin US, F, G, I, 600,000–720,000 Hypotension (recombinant S, UK, J IU/kg IV every Fever interleukin-2) 8 hours for 14 Diarrhea (Chiron’s Proleukin, doses over Vomiting Shionogi’s 5 days (as Thrombocytopenia Imunace) tolerated) Malaise Dacarbazine Dacarbazine (Bayer’s US, F, G, I, 850–1,000 mg/m2I V Neutropenia day 1 or 250 IV Thrombocytopenia DTIC-Dome, S, UK, J mg/m2 days 1–5; Anemia generics) repeated every Nausea/vomiting 3–4 weeks Cisplatin/ Cisplatin US, F, G, I, 100 mg/m2 IV day Leukopenia 1; repeated every Neutropenia with/ vinca (Bristol-Myers S, UK, J 4 weeks for 3 without fever alkaloid/ Squibb’s Platinol cycles Thrombocytopenia dacarbazine AQ/Platinex, Nausea/vomiting regimen Bristol/Nihon Diarrhea Kayaku’s Neuropathy Briplatin/Randa, Renal toxicity generics) Vinblastine (Eli US, F, G, I, 3 mg/m2 IV days 1, 8, and 15; Lilly/EG S, UK, J repeated every 4 Labo/Crinos/ weeks for 3 Clonmel’s Velbe, cycles Eli Lilly/Shionogi’s Exal, generics) High-dose interferonalpha-2

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TABLE 10. (continued) Regimen or Class

Regimen Components Agent

Availability

Dose

Common Toxicities

Vindesine (EG F, G, I, S, 3 mg/m2 IV days1, Labo/Cell 8, and 15; UK, J Pharm/Crinos/ repeated every 4 Clonmel’s Eldisine, weeks for 3 Ciclum Farma’s cycles Enison, Shionogi’s Fildesin) Dacarbazine (Bayer’s US, F, G, I, 250 IV mg/m2 days 1–5; repeated DTIC-Dome, S, UK, J every 4 weeks for generics) 3 cycles Dartmouth Dacarbazine (Bayer’s US, F, G, I, 220 IV mg/m2 days Leukopenia 1–3; repeated Neutropenia regimen DTIC-Dome, S, UK, J every 3 weeks Thrombocytopenia generics) Nausea/vomiting Fatigue Dyspnea Cisplatin US, F, G, I, 25 mg/m2 IV days 1–3; repeated (Bristol-Myers S, UK, J every 3 weeks Squibb’s Platinol AQ/Platinex, Bristol/Nihon Kayaku’s Briplatin/Randa, generics) Carmustine US, F, G, S, 150 mg/m2 IV day 1; repeated every (Bristol-Myers UK odd 3-week cycle Squibb’s BICNU/ Carmubris) Tamoxifen US, F, G, I, 20–40 mg PO (AstraZeneca’s S, UK, J starting 1 week Nolvadex, before generics) chemotherapy, continuing as long as patient is on treatment Leukopenia Cisplatin/ Cisplatin US, F, G, I, 25 mg/m2 /day IV days 1–3; repeat Neutropenia vinca (Bristol-Myers S, UK, J cycle every Thrombocytopenia alkaloid/ Squibb’s Platinol 28 days Nausea/vomiting dacarbazine/ AQ/Platinex, Fatigue aldesleukin/ Bristol/Nihon Fever interferonKayaku’s alpha-2 Briplatin/Randa, regimen generics)

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TABLE 10. (continued) Regimen or Class

Regimen Components Agent

Availability

Dose

Common Toxicities

Vinblastine (Lilly/EG US, F, G, I, 3 mg/m2 IV days 1, Labo/Crinos/ 8, and 15; S, UK, J Clonmel’s Velbe, repeated every 4 Eli Lilly/Shionogi’s weeks for 3 Exal, generics) cycles Dacarbazine (Bayer’s US, F, G, I, 250 mg/m2 /day IV days 1–3; repeat DTIC-Dome, S, UK, J cycle every generics) 28 days Interferon-alpha-2 US, F, G, I, 5 MU/m2 /day SC days 6, 8, 10, 13, (Schering-Plough’s S, UK, J and 15; repeat Intron A, cycle every Roche/Chugai’s 28 days Roferon A, Takeda’s Canferon A) Aldesleukin US, F, G, I, 18 MU/m2 /day days 6–10 and 13–15; (recombinant S, UK, J repeat cycle interleukin-2) every 28 days (Chiron’s Proleukin, Shionogi’s Imunace) IV = Intravenous; SC = Subcutaneous; PO = Oral; IM = Intramuscular. US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan.

Clinical Performance. Numerous controlled trials have been conducted in the United States to evaluate the effect of adjuvant high-dose IFN-α-2b on relapsefree survival and overall survival in high-risk patients (stages II and III). High-dose IFN-α-2b received approval in 1995 on the basis of the Eastern Cooperative Oncology Group (ECOG) trial ECOG 1684 (Kirkwood JM, 1996). Based on a seven-year follow-up, this trial of 280 patients with stage IIB and stage III MM (using a high-dose regimen of 20 MU/m2 by intravenous infusion 5 days per week for 4 weeks, followed by 10 MU/m2 subcutaneously 3 days per week for 48 weeks) demonstrated that IFN-α-2b signiÞcantly prolongs both disease-free survival and overall survival compared with observation. At Þve years, the relapse-free survival and overall survival were 37% and 46%, respectively, in the high-dose IFN-α-2b arm, and 26% and 37%, respectively, in the observation arm; the statistical relevance of these values is controversial. At the 2001 American Society of Clinical Oncology (ASCO) meeting, researchers presented updated results based on a 12-year follow-up (Kirkwood JM, 2001[c]). These data conÞrmed the disease-free survival beneÞt but not the overall survival beneÞt. A three-arm trial (ECOG 1690) was designed to conÞrm the results of ECOG 1684 as well as to evaluate adjuvant high-dose and low-dose IFN-α-2b, administered for two years, versus observation. This trial, which recruited 651 patients

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with high-risk stage II and stage III disease, conÞrmed the disease-free survival beneÞt of high-dose IFN-α-2b over observation though it did not demonstrate an overall survival beneÞt for high- or low-dose IFN-α-2b (Kirkwood JM, 2000). At 52 months, survival was 52%, 53%, and 55% for high-dose, low-dose, and observation, respectively. The investigators speculate that differences in postrelapse treatment may account for the lack of overall survival beneÞt for the high-dose IFN-α-2b arm. Thirty-one percent of patients in the observation arm were treated with high-dose IFN-α-2b on relapse, which probably contributed to the improved Þve-year survival compared with the observation arm in ECOG 1684 (55% versus 37%). Intermediate-Dose Interferon-Alpha-2 Overview. Researchers have investigated the use of intermediate-dose IFN in an attempt to reduce the incidence of severe toxicity (particularly psychological disturbances that in the extreme may result in suicide) caused by high-dose IFN, while retaining the clinical activity and improvement in quality of life associated with this treatment. Intermediate-dose regimens involve IFN administered at doses of 5–10 MU/m2 . IFN-α-2a (Roche/Chugai’s Roferon A, Takeda’s Canferon A) and IFN-α-2b (Schering-Plough’s Intron A) tend to be used interchangeably in markets where both are available. We present the trial results for IFN-α-2b because this agent is marketed for adjuvant use in MM. Mechanism of Action. The IFNs are a family of naturally occurring cytokines that exhibit antiviral, antitumor, and immunomodulatory activities. Their antitumor effect derives from their ability to inhibit cell-cycle progression, reduce tumor-cell protein synthesis, and inhibit the formation of colonies of late progenitor cells. Their immunomodulatory role involves inducing the expression of major histocompatability antigens and modulating the expression and function of T cells, monocytes, and natural killer cells (large granular lymphocytes that bind to and kill cells by releasing cytotoxins). Clinical Performance. The toxicity of high-dose IFN-α schedules and the lack of efÞcacy of low-dose IFN-α schedules have prompted investigators to examine the efÞcacy of intermediate-dose IFN-α. A large Phase III trial in 1,418 stage IIB and stage III MM patients, sponsored by the European Organization for Research into the Treatment of Cancer (EORTC), has shown a distant-metastasis diseasefree survival beneÞt, compared with observation. The intermediate-dose schedule used in the study was 10 MU of IFN-α-2b subcutaneously Þve days per week for four weeks followed by two years of treatment with 5 MU subcutaneously three times per week. No advantage was gained with a shorter, higher-dose regimen (10 MU of IFN-α-2b subcutaneously Þve days per week for four weeks followed by one year of treatment with 10 MU subcutaneously three times per week). These data demonstrate that duration of IFN treatment may be more important to a successful outcome than the dose of IFN. At a median followup of two years, researchers found no signiÞcant difference in survival times

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between the arms (Eggermont AMM, 2001). The toxicity associated with these doses was manageable and comprised grade 3 and 4 ßu-like symptoms, fatigue and lethargy, and psychiatric effects occurring in approximately 10% of patients and hematological and renal effects in less than 2%. Low-Dose Interferon-Alpha-2 Regimen Overview. Owing to the severe toxicity associated with high-dose IFN-α-2 therapy and the lack of compelling evidence to recommend treatment with this regimen, an extended duration of low-dose treatment has been investigated. This approach aims to determine whether it is possible to preserve or improve on the response rate and time to disease progression obtained with the high-dose regimen. Low-dose regimens are administered at doses of 1–3 MU/m2 . IFN-α-2a (Roche/Chugai’s Roferon A; Takeda’s Canferon A) and IFN-α-2b (ScheringPlough’s Intron A) tend to be used interchangeably in markets where both treatments are available. Mechanism of Action. The IFNs are a family of naturally occurring cytokines that exhibit antiviral, antitumor, and immunomodulatory activities. Their antitumor effect derives from their ability to inhibit cell-cycle progression, induce a reduction in tumor-cell protein synthesis, and inhibit the formation of colonies of late progenitor cells. Their immunomodulatory role involves inducing the expression of major histocompatability antigens and modulating the expression and function of T cells, monocytes, and natural killer cells (large granular lymphocytes that bind to and kill cells by releasing cytotoxins). Clinical Performance. The efÞcacy of low-dose IFN-α (3 MU subcutaneously three times per week) has been tested in both low-risk stage II disease as well as high-risk stage II and stage III disease. In high-risk stage II and stage III disease, several clinical trials have found no beneÞt with low-dose IFN-α (see, for example, Hancock BW, 2001 [IFN-α-2a]; Kirkwood JM, 2000 [IFN-α-2b]). Published data from a United Kingdom Coordinating Committee on Cancer Research (UKCCCR) randomized Phase III trial of 674 patients, called the Adjuvant Interferon in High-Risk Melanoma (AIM-High) study, conÞrmed that low-dose IFN is not superior to observation alone as adjuvant treatment for completely resected high-risk MM patients (Hancock BW, 2004). The Þve-year analysis shows overall survival of 46% for the IFN-treated arm (3 MU subcutaneously three times per week for two years or until relapse) and 42% for the control arm. This difference is not statistically signiÞcant. Toxicity of low-dose IFN-α-2 in this study was modest, although one in six patients discontinued treatment because of side effects, primarily fatigue and mood disturbance. Several other trials, however, have suggested that low-dose IFN-α-2, taken for 18 months, is beneÞcial in terms of relapse-free survival in stage II patients (without factors denoting high risk). A French Cooperative Group on Melanoma trial compared low-dose IFN-α-2a (3 MU subcutaneously three times per week) for 18 months versus observation in 499 clinically node-negative patients with

538

MALIGNANT MELANOMA

tumors more than 1.5 mm thick. IFN-α-2a patients showed a signiÞcant improvement in relapse-free survival: the three-year relapse rate was 32% in the IFN-α-2a group and 44% in the control arm. The trend toward improved overall survival was not signiÞcant (Grob JJ, 1998). In a second trial conducted by the Austrian Malignant Melanoma Cooperative Group, 311 patients were randomized to receive IFN-α-2a (3 MU daily for three weeks, followed by 3 MU subcutaneously three times weekly for one year) or observation. After a median follow-up of 41 months, patients showed a signiÞcant improvement in relapse-free survival; the difference in overall survival was not signiÞcant, as would be expected with this short duration of follow-up (Pehamberger H, 1998). Longer-term data are not available. Aldesleukin Regimen Overview. Chiron’s genetically engineered (recombinant) interleukin-2 (rIL-2, aldesleukin, Proleukin; Shionogi’s Imunace) received FDA approval for mMM in 1998 after being granted orphan drug status in 1996. In Europe and Japan, it is approved only for renal cell carcinoma and angiosarcoma. RIL-2 {rIL-2} has many shortcomings; it is effective in only a small proportion of patients, is highly toxic (Schwartz R, 2002), and requires frequent intravenous infusions involving inpatient hospital administration and close monitoring. The approved high-dose regimen of aldesleukin involves two Þve-day cycles of treatment separated by nine days of rest. The approved dose is 600,000 IU/kg administered every eight hours by a 15-minute infusion for a maximum of 14 doses. In contrast to its success in renal cancer, low-dose aldesleukin has not shown activity in MM. Mechanism of Action. Aldesleukin is recombinant IL-2. RIL-2 is a naturally occurring protein that has no direct antitumor effect but enhances the body’s own immune response; it is naturally produced by lymphocytes to stimulate the growth and maturation of T cells, which can eliminate certain tumor cells. Clinical Performance. A meta-analysis of eight clinical trials with 270 mMM patients treated with aldesleukin in clinical studies has been reported (Atkins MB, 1999). The analysis revealed an overall response rate of 16%, comprising 6% with complete responses and 10% with partial responses. The median duration of response for patients achieving a partial response was six months. Although complete responses were rare, they were associated with a long median survival—the minimum median duration of response for complete responders is at least 59 months (the median duration of complete response has not yet been reported) (Atkins MB, 2000). Any patient who maintained a response for 30 months did not have their disease progress, and some of these patients responded for more than ten years (Proleukin prescribing information, Chiron, 2000). These prolonged responses were achieved primarily by patients who had not been previously treated and who had no visceral disease. The toxicities associated with aldesleukin are common and severe; in these trials, the most frequent were hypotension (71% of patients), diarrhea (67%), ßu-like symptoms (52%), and vomiting (50%) (Atkins M, 1999). Additionally,

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thrombocytopenia occurred in 37% of patients, anemia in 29%, and leukopenia in 16%. The most serious effect of aldesleukin is capillary leak syndrome (escape of plasma proteins and ßuid into the extravascular space), resulting in hypotension and reduced organ perfusion; the latter can be fatal. Drug-related deaths occurred in 2% of the 270 stage IV MM patients treated in the clinical studies covered in this meta-analysis. The authors concluded that the meta-analysis demonstrated that aldesleukin provides real clinical beneÞt for approximately 4% of patients and should be considered only as treatment for select patients with good performance status. As experience has grown with aldesleukin, toxicity management—particularly of capillary leak syndrome—and careful patient selection have reduced the incidence of severe toxicity. Control of capillary leak syndrome is achieved by careful monitoring of ßuid perfusion rates and urine output. Either intravenous ßuids or diuretics may be used to control any ßuid imbalance. The utility of high-dose aldesleukin has been limited by its toxicity. Patients with indwelling central lines who receive aldesleukin are particularly at risk for infection with gram-positive microorganisms. Antibiotic prophylaxis has been used to address this problem. Other side effects of high-dose aldesleukin include hypotension, cardiac arrhythmias, pulmonary edema, fever, increased capillary permeability, and, in rare instances, death. Researchers have explored many approaches for enhancing the therapeutic index of IL-2 by dissociating its toxic effects from its antitumor effects. Antibiotic prophylaxis has reduced the incidence of catheter-related sepsis and, given that sepsis was largely responsible for mortality from high-dose IL-2, has dramatically improved the safety of this therapy. Premedication with dexamethasone has been shown to prevent induction of TNF by IL-2 and to signiÞcantly reduce the incidence and severity of fever and hypotension. These maneuvers have enabled a threefold increase in the maximum tolerated dose. Possible interference with antitumor efÞcacy has, however, limited this approach, highlighting the need for more-selective ways of modifying IL-2 toxicity (Eastman ME, 2001). Alternative IL-2 dosing schedules have been tested in many clinical trials (Eton O, 2000[b]). Intermediate- and high-dose continuous infusion schedules remain common treatments and result in objective responses (15–20%) comparable to those of high-dose bolus IL-2 (Allen IE, 1998). In addition, durable remissions are observed in a small subset of patients. More recently, European investigators piloted continuous-infusion IL-2 in a decrescendo-dosing schedule (typically intravenous IL-2 at a dose of 18 MIU/m2 over 6 hours, followed by 18 MIU/m2 over 12 hours, then 18 MIU/m2 over 24 hours, and Þnally 4.5 MIU/m2 per day for three consecutive days). A small, randomized Phase III trial comparing continuous infusion IL-2, versus continuous infusion decrescendo IL-2, both with IFN-α-2, demonstrated improved response rates and reduced toxicity with decrescendo dosing (Keilholz U, 1993). This dosing schedule is now the standard for incorporating IL-2 in biochemotherapy regimens in Europe. The addition of decrescendo IL-2, in combination with cisplatin, dacarbazine, and IFN-α-2, has been compared with the same regimen without IL-2 (Keilholz U, 2003). Between 1995 and 2000, 363 mMM patients were recruited into this

540

MALIGNANT MELANOMA

study. The Þnal analysis showed that, despite a trend toward improved survival at two years (from 12.8% to 17.6%) and at three years (8.5% and 12.7%) when IL-2 was included, the differences were not statistically signiÞcant and did not justify its use. Further clinical trials are required to clarify the potential role of biochemotherapy regimens in clinical practice. Dacarbazine Overview. Dacarbazine (Bayer’s DTIC-Dome, generics) (Figure 2) has been established as standard treatment of mMM for more than 30 years and is the most widely prescribed drug for this indication. It can be given in short courses of less than Þve days or in longer courses of ten days. Despite its widespread use, complete or durable responses with dacarbazine are rare, median survival is not improved, and the Þve-year survival rate remains less than 2%. Mechanism of Action. Dacarbazine is an alkylating agent and a pro-drug of monomethyl triazeno imidazole carboxamide (MTIC). The cytotoxicity of MTIC is thought to be caused primarily by methylation of DNA at the O6 position of guanine, which prevents DNA replication. Because dacarbazine is a purine analogue, it may also inhibit cellular proliferation by its misincorporation into DNA. Clinical Performance. Dacarbazine achieves a response in up to 20% of patients and, in some trials, has improved survival versus observation alone. The median duration of response is Þve to six months (Kirkwood JM, 1993). The most signiÞcant toxicities associated with dacarbazine include myelotoxicity (which is dose-limiting), nausea, vomiting, and fever. A meta-analysis of 20 randomized clinical trials and 3,273 patients, comparing single-agent dacarbazine with combination treatments, has been published (Huncharek M, 2001). This analysis reveals that single-agent dacarbazine may be inferior to combination regimens, in particular to dacarbazine plus IFN-α-2, in terms of response rate, but researchers identiÞed no difference in overall survival. Cisplatin/Vinca Alkaloid/Dacarbazine Regimen Overview. The cisplatin/vinca alkaloid/dacarbazine (CVD) regimen comprises cisplatin (Bristol-Myers Squibb’s Platinol AQ/Platinex, Bristol/Nihon Kayaku’s Briplatin/Randa, generics), a vinca alkaloid, and dacarbazine (Bayer’s DTIC-Dome, generics). The vinca alkaloids vinblastine (Eli Lilly/EG Labo/Crinos/Clonmel’s Velbe, Eli Lilly/Shionogi’s Exal, generics) and vindesine O NH2 N

CH3

N N H

N

N CH3

FIGURE 2. Structure of dacarbazine.

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(Clonmel’s Eldisine) are considered equivalent and are used interchangeably. This regimen was developed in an attempt to improve the response rate and survival of mMM patients treated with dacarbazine alone. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

•

•

Cisplatin (Figure 3) is a platinum agent. Platinum agents act by generating highly reactive, charged platinum complexes that bind covalently to DNA, leading to intra-strand, inter-strand, and protein cross-linking of DNA, resulting in inhibition of transcription and/or DNA replication mechanisms. Vinblastine and vindesine are vinca alkaloids (Figure 4). Vinca alkaloids interact with tubulin and disrupt microtubular function in the mitotic spindle. This action leads to metaphase arrest, resulting in mitotic arrest and cell death. Dacarbazine (Figure 2) is an alkylating agent and a pro-drug of MTIC. The cytotoxicity of MTIC is thought to be caused primarily by methylation of DNA at the O6 position of guanine, which prevents DNA replication. Because dacarbazine is a purine analogue, it may also inhibit cellular proliferation by its mis-incorporation into DNA.

Clinical Performance. In a small trial recruiting 52 patients with advanced MM, CVD (the vinca alkaloid used in this study was vinblastine) produced a response in 40% of patients (including 4% who achieved complete responses) and a median duration of response of nine months (Legha SS, 1989). A multicenter trial comparing CVD with dacarbazine alone produced response rates of 19% and 14%, respectively, with no difference in response duration or survival (Lotze MT, 2001).

FIGURE 3. Structure of cisplatin.

CH3

N

HN

OH

H3CO O

H

N H

H3CO

CH3

OH N R1

R3 H

R2

FIGURE 4. Structure of vinblastine (R1 = CH3 , R2 = CO(OCH3 ), R3 = OCO(CH3 ) and vindesine(R1 = CH3 , R2 = CO(NH2 ), R3 = OH).

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MALIGNANT MELANOMA

In a European multicenter Phase III study (Jungnelius U, 1998), 326 patients with mMM were randomized to receive treatment with dacarbazine and vindesine with or without cisplatin. The results showed no statistically signiÞcant difference in overall survival between the two treatment arms but did show additional toxicity when cisplatin was added to the treatment regimen. A Phase III randomized trial reported by Eton and colleagues investigated the addition of cytokines to the CVD (using vinblastine) regimen (Eton O, 2002). The 92 patients receiving chemotherapy alone achieved a response rate of 25% and median survival of 9.2 months. These data are comparable to those reported at a recent meeting of the American Society of Clinical Oncology (ASCO), also comparing CVD (using vindesine) with and without biotherapy (Del Vecchio M, 2003). The results showed a response rate of 22% and a median overall survival of 12 months. The addition of cisplatin and a vinca alkaloid to dacarbazine does not appear to signiÞcantly and consistently improve overall survival compared with dacarbazine alone. Dartmouth Regimen Overview. The Dartmouth regimen consists of treatment with dacarbazine (Bayer’s DTIC-Dome, generics), cisplatin (Bristol-Myers Squibb’s Platinol AQ/Platinex, Bristol/Nihon Kayaku’s Briplatin/Randa, generics), carmustine (Bristol-Myers Squibb’s BICNU/Carmubris), and tamoxifen (AstraZeneca’s Nolvadex, generics). Approximately 25% of MMs have demonstrable estrogen receptors (Cohen C, 1990). Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the regimen’s overall activity: •

•

•

•

Dacarbazine (Figure 2) is an alkylating agent and a pro-drug of MTIC. The cytotoxicity of MTIC is thought to be caused primarily by methylation of DNA at the O6 position of guanine, which prevents DNA replication. Because dacarbazine is a purine analogue, it may also inhibit cellular proliferation by its misincorporation into DNA. Cisplatin (Figure 3) is a platinum agent. Platinum agents act by generating highly reactive, charged platinum complexes that bind covalently to DNA, leading to intra-strand, inter-strand, and protein cross-linking of DNA resulting in inhibition of transcription and/or DNA replication mechanisms. Carmustine (Figure 5) is a nitrosourea alkylating agent. Metabolites of carmustine cause alkylation and cross-linking of DNA, which prevent DNA replication and cell division. Other biologic effects include inhibition of DNA repair. Nitrosoureas generally lack cross-resistance with other alkylating agents. Tamoxifen (Figure 6) is a selective estrogen receptor modiÞer (SERM). SERMs act by competitively blocking estrogen receptors, thereby interfering with estrogen’s signaling of epithelial cell growth and reproduction.

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FIGURE 5. Structure of carmustine.

FIGURE 6. Structure of tamoxifen.

Clinical Performance. A randomized double-blind, placebo-controlled trial compared carmustine, dacarbazine, and cisplatin with tamoxifen (Dartmouth regimen) or without tamoxifen (Rusthoven JJ, 1996). The endpoints of the trial were response rate and survival. The study accrued 211 mMM patients between February 1992 and January 1995, 199 of whom were assessable for response and toxicity. The overall response rate was 21% in the placebo group and 30% in the tamoxifen group. The difference in response rate was not statistically signiÞcant. A Phase III multicenter randomized clinical trial was conducted to compare the Dartmouth regimen with single-agent dacarbazine following reports of several single-institution Phase II studies claiming response rates of up to 55% in stage IV MM patients (Chapman P, 1999). Some sporadic cases that maintained complete response to treatment were also noted. A total of 240 patients with measurable stage IV MM were randomized to receive either the Dartmouth regimen or dacarbazine; the treatment was repeated every three weeks. Median survival time was seven months from randomization; 25% of the patients survived for one year or longer. No difference in survival time was observed between the two arms when analyzed on an intent-to-treat basis or when considering only the 231 eligible treated patients. Investigators observed a nonsigniÞcant trend toward an improved response rate for the Dartmouth regimen of 18.5% compared with 10.2% for the dacarbazine single-agent arm (p = 0.09). Bone marrow suppression, nausea and vomiting, and fatigue were all signiÞcantly more common in the Dartmouth-treated arm. Based on these Þndings, the authors concluded that single-agent dacarbazine should remain the standard treatment choice for stage IV MM patients.

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MALIGNANT MELANOMA

Cisplatin/Vinca Alkaloid/Dacarbazine/Aldesleukin/ Interferon-Alpha-2 Regimen Overview. Numerous combinations of chemotherapeutic agents with one or more biological response modiÞers (biochemotherapy) have been studied since the 1990s, each with the aim of improving mediocre response rates and the abysmal survival of patients with mMM, at acceptable toxicity costs. Combining chemotherapy with a biological response modiÞer that has proven activity in MM has the advantage that the two approaches will have non-overlapping toxicity proÞles and can be administered simultaneously, and, it is hoped, will offer additive efÞcacy. A small Phase III study comparing dacarbazine (Bayer’s DTIC-Dome, generics) with or without IFN-α-2 (Roche/Chugai’s Roferon A; Schering-Plough’s Intron A; Takeda’s Canferon A) found an improved rate and duration of response on the addition of IFN-α-2. Other studies have reported response rates of up to 64% with occasional durable responses, providing impetus for the testing of further biochemotherapy combinations (Legha SS, 1998). However, several other studies, using different dosing schedules, have failed to conÞrm these Þndings, and central nervous system (CNS) relapse (i.e., brain metastases) continues to be a particular problem in these patients. Further clinical trials are required to optimize the doses and scheduling of biochemotherapy regimens before they are accepted into widespread use. Because so many different biochemotherapy combinations are possible, we focus on one regimen in this section, which incorporates cisplatin (Bristol-Myers Squibb’s Platinol AQ/Platinex, Bristol/Nihon Kayaku’s Briplatin/Randa, generics), a vinca alkaloid, dacarbazine, IFN-α-2, and aldesleukin (Chiron’s Proleukin; Shionogi’s Imunace). Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the overall regimen activity: •

•

•

•

Cisplatin (Figure 3) is a platinum agent. Platinum agents act by generating highly reactive, charged platinum complexes that bind covalently to DNA, leading to intra-strand, inter-strand, and protein cross-linking of DNA, resulting in inhibition of transcription and/or DNA replication mechanisms. Vinblastine and vindesine are vinca alkaloids (Figure 4). Vinca alkaloids interact with tubulin and disrupt microtubular function in the mitotic spindle. This action leads to metaphase arrest, resulting in mitotic arrest and cell death. Dacarbazine (Figure 2) is an alkylating agent and a pro-drug of MTIC. The cytotoxicity of MTIC is thought to be caused primarily by methylation of DNA at the O6 position of guanine, which prevents DNA replication. Because dacarbazine is a purine analogue, it may also inhibit cellular proliferation by its mis-incorporation into DNA. Aldesleukin is recombinant IL-2. IL-2 is a naturally occurring protein that has no direct antitumor effect but enhances the body’s own immune response; it is naturally produced by lymphocytes to stimulate the growth and maturation of T cells, which can eliminate certain tumor cells.

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•

545

The IFNs are a family of naturally occurring cytokines that exhibit antiviral, antitumor, and immunomodulatory activities. Their antitumor effect derives from their ability to inhibit cell-cycle progression, induce a reduction in tumor-cell protein synthesis, and inhibit the formation of colonies of late progenitor cells. Their immunomodulatory role involves inducing the expression of major histocompatability antigens and modulating the expression and function of T cells, monocytes, and natural killer cells (large granular lymphocytes that bind to and kill cells by releasing cytotoxins).

Clinical Performance. In 2000, Eton and colleagues presented the Þrst U.S. randomized study comparing sequential biochemotherapy (CVD + IL-2 + IFN-α-2) with combination chemotherapy (CVD) alone (Eton O, 2000). This 183patient study demonstrated a signiÞcant improvement in response rate from 25% to 48% and extended time to progression from 2.4 to 4.9 months, favoring the biochemotherapy arm. In addition, results showed a signiÞcant 30% improvement in median survival, from 9.2 months to 11.9 months. The accrual goal was increased to 482 patients in order to provide a 90% power that could detect a 33% improvement in median survival from 9 to 12 months, as seen in the Eton trial described previously. However, interim data presented at the ASCO meeting in 2003 did not look promising. The biochemotherapy arm exhibited a higher response rate than the CVD arm, 16.6% versus 11.9%, and response duration was 9.4 months versus 6.1 months, though this did not translate into the improved overall survival (9.1 months versus 8.4 months) seen in the Eton study (Atkins MB, 2003). None of the ECOG study data reached statistical signiÞcance, although subgroup analysis does suggest that patients who had not previously been treated with IFN-α-2 may beneÞt most. Further studies will be critical for determining the future role of biochemotherapy in the treatment of mMM and in adjuvant treatment of high-risk disease. Nonpharmacological Approaches Surgery is the mainstay of treatment for MM. In most patients, pathological conÞrmation of MM is made following an excisional biopsy (removal of the entire suspicious lesion). A wide local excision (removal of the tissue around the site of the MM) is the standard initial treatment of stage I-III MM. Guidelines recently published by the American Academy of Dermatology (Sorber AJ, 2001) recommend that excisional margins should vary according to the thickness of the lesion; from 5 mm for in situ lesions to 2 cm for lesions greater than 2 mm thick. The academy made this recommendation on the basis of several randomized trials that have shown no beneÞt in survival or recurrence with larger excision margins (see, e.g., Cohn-Cedermark G, 2000). For patients with thin tumors (less than 1 mm deep), relapse after surgery is rare, and patients require no further therapy. In patients with deeper tumors, risk of relapse after excision is higher, and sentinel lymph node biopsy or elective removal of the lymph nodes (elective lymph node dissection) and/or systemic adjuvant therapy may be advisable. Surgeons perform incisional biopsy (removal of part of a lesion, including the center) only when suspicion of MM is very low, the lesion is large, or excisional

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MALIGNANT MELANOMA

biopsy is either impractical for some reason (e.g., in the case of subungual [nail bed] lesions) or cosmetically undesirable (e.g., MMs of the face). In advanced MM, patients with isolated lung, liver, gastrointestinal, or distant lymph node metastases are all candidates for surgical resection. Elective lymph node dissection (ELND)—the prophylactic removal of all lymph nodes in the draining basin of a tumor—is a controversial procedure historically used for patients with lesions of intermediate thickness. ELND has two purposes. First, ELND provides prognostic information: the number of involved nodes is an important predictor of survival. Second, it reduces the risk of relapse after excision. This approach follows the theory that MM spreads through the draining lymph nodes before metastasizing to more distant sites. The practice of ELND was discredited in the late 1990s by a series of World Health Organization (WHO) and other trials that showed no survival beneÞt for patients undergoing this invasive procedure. Accordingly, ELND has been largely replaced by sentinel lymph node biopsy, followed by removal of nodes only in the case of a positive sentinel node. Further, despite recent Þndings that cast more positive light on ELND, the practice of sentinel lymph node biopsy has gained considerable momentum, and clinicians interviewed for this report do not expect to see a reversal in the trend away from ELND. Sentinel lymph node biopsy (SLNB) is a minimally invasive method of detecting micrometastatic nodal disease. Designated as the “standard of care” by the WHO (Cascinelli N, 2000), it is less invasive and less expensive than ELND and allows for the detection of subclinical disease 18–24 months before clinical manifestation of nodal disease. Two studies presented at ASCO in 2001 supported the reliability of SLNB as a prognostic tool. Both found a low likelihood of failure in sentinel-node-negative patients (Essner R, 2001; Balch CM, 2001)[b] Although the value of SLNB as a prognostic tool is well established, the role it should play in inßuencing therapy remains to be determined. It is still unknown whether SLNB followed by lymphadenectomy of clinically positive nodes is preferable to delaying surgery until nodal disease becomes palpable. WHO trial 14 found a survival beneÞt for the former scenario. The role of radiotherapy in the treatment of MM is primarily palliative—the alleviation of symptoms—because its use provides no survival advantage. Therefore, radiation therapy is generally reserved for those advanced cases of MM where surgery is not possible. EMERGING THERAPIES Owing to the lack of effective treatment for malignant melanoma (MM) and the large number of MM patients that currently receive no drug therapy, immense commercial and academic effort is focusing on the development of novel agents for the treatment of this disease. This effort is dominated by the search for an effective therapeutic vaccine, aimed at augmenting the host immune response against the tumor. Interviewed experts express great interest about the prospect of vaccine therapies; however, they doubt that any vaccine therapies currently in clinical development will be effective in, and eventually marketed for, the treatment of MM. Most believe that vaccine therapies will ultimately play a role in the

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treatment of MM. Further research is required to optimize the most appropriate MM targets for vaccines and to improve large-scale vaccine production. Anti–vascular endothelial growth factor (VEGF) approaches, such as the VEGF neutralizing antibody bevacizumab (Roche/Genentech’s Avastin), are also under investigation for the potential treatment of MM. Trials of these agents have now reached Phase II. Table 11 summarizes the majority of drug therapies in development for MM, as covered in this reference. Vaccines Overview. Although the concept of a therapeutic tumor vaccine is a viable one, these therapies remain largely unproven for the treatment of cancer. Phase III data subgroup analysis has shown that Dendreon’s Provenge, a vaccine for the treatment of metastatic prostate cancer, has clinical activity in patients with good performance status whose tumors have less aggressive characteristics. Together with an improvement in survival (approximately eight months), this factor may stimulate renewed interest in recruiting patients for MM vaccine trials. For Provenge, shown to activate T cells, the latest data available at the time of composing this reference indicates that a rolling BLA was to be Þled in 2006, for which the company would apply for priority review. Vaccines for MM can be broadly categorized into whole-cell vaccines (e.g., Avax Technologies’ M-Vax); peptide and ganglioside vaccines produced from puriÞed antigens (e.g., Ludwig Institute for Cancer Research’s Melan-A/MART1, Antigenics’ {Oncophage (vitespen, formerly HSPCC-96), Progenics Pharmaceutical’s GM2-KLH/QS-21); and antibody-based vaccines (e.g BioGen/IDEC’s Mel-Immune {Melimmune}-1 and -2). Most vaccines are administered with adjuvants such as bacille Calmette-Gu´erin (BCG, a nonspeciÞc immunostimulant) and QS-21 (a plant extract)—agents that stimulate the immune system to recognize the antigens in the vaccines. Some vaccines discussed in the following sections are coadministered with interleukin2 (IL-2) as an alternative means of stimulating the immune system. Because the role of the immune system is well established in MM and is characterized by sporadic spontaneous regression, MM is an attractive disease target for the development of vaccine treatments. Vaccines offer three major advantages over current treatments: they are generally well tolerated, the anticipated autoimmunity has not materialized as a clinical problem, and they have convenient dosing regimens. If vaccines are shown to be efÞcacious in any stage of MM, they will be rapidly adopted and widely used. Despite the promise of whole-cell vaccines in Phase II trials, available Phase III data show that these vaccines have not lived up to expectations, though many are ongoing and are yet to report results. For vaccines to be successful, experts believe that either concurrent administration of multiple vaccines or administration of a single vaccine that targets multiple antigens will be necessary. This strategy will maximize the likelihood of a patient’s tumors expressing an antigen targeted by the vaccines and minimize the chances of tumor cells lacking targeted antigens and escaping successful treatment. In a further step, Individuals

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MALIGNANT MELANOMA

TABLE 11. Emerging Therapies in Development for Malignant Melanoma Compound Vaccines Melacine United States Europe Japan

Development Phase

Marketing Company

D D —

Corixa Corixa —

Melan-A/MART-1 United States Europe Japan

— II —

— Ludwig Institute for Cancer Research —

Vitespen (Oncophage; formerly HSPCC-96) United States Europe Japan

III III —

Antigenic Antigenic

GM2-KLH/QS-21 United States Europe Japan

III III —

Progenics Progenics —

TriGem United States Europe Japan

S — —

Titan Pharmaceuticals — —

Antisense therapies Oblimersen (Genasense) United States Europe Japan

D — —

Genta; Aventis Genta; Aventis —

Alkylating agents Temozolomide (Temodar) United States Europe Japan

III III —

Schering-Plough Schering-Plough —

Immunomodulators Pegylated-interferon-alpha-2 United States Europe Japan

III — —

Roche; Schering-Plough — —

GM-CSF (Sargramostim, Leukine) United States Europe Japan

III — —

Berlex Laboratories — —

Thalidomide United States Europe Japan

II II —

Celgene Celgene —

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TABLE 11. (continued) Compound

Development Phase

Marketing Company

CC-5013 (Revimid) United States Europe Japan

III — —

Celgene — —

Histamine dihydrochloride (Ceplene) United States Europe Japan

III PR —

Maxim Maxim —

III III —

Bayer/Onyx Pharmaceuticals Bayer/Onyx Pharmaceuticals —

RAF kinase inhibitors (Sorafenib) United States Europe Japan

GM-CSF = Granulocyte-macrophage colony-stimulating factor. D = Discontinued; PR = Preregistered; S = Suspended.

may receive a cocktail of vaccines tailored to the characteristics of their tumor. A major impediment to progress, however, has been the inability to identify and measure the immune response that accounts for the clinical beneÞt. The lack of an immune surrogate for efÞcacy makes it much more difÞcult to improve vaccines as well as optimize their dosing regimens because trials designed to compare these factors use immunity endpoints that may not be related to the way the vaccine actually works. Mechanism of Action. Cancer vaccines are designed to stimulate the immune system to launch a response against the speciÞc target contained by the vaccine. In general, research has shown that the most effective antitumor immune responses are achieved by stimulating cytotoxic T cells, which can recognize and kill tumor cells directly. Melacine. (NOTE: the development of this vaccine has been discontinued in US, but approved in Canada!) Corixa’s Melacine is an allergenic whole-cell vaccine that consists of lysed cells from two MM cell lines. Melacine is administered in combination with Corixa’s proprietary vaccine adjuvant, Detox. Detox adjuvant includes monophosphoryl lipid (MPL) A adjuvant and mycobacterial cell-wall skeleton, both of which activate the human immune system. Melacine vaccine is administered in two injections and delivered over four six-month cycles, each consisting of ten treatments followed by a three-week rest. Interim analysis from a Phase III study, designed to detect a 50% improvement in survival, was reported at the American Society of Clinical Oncology (ASCO) meeting in May 2003 (Mitchell M, 2003). Between April 1997 and January 2003, 604 resected, stage III MM patients were recruited. The interim analysis, of 598 patients, showed no signiÞcant difference in disease-free survival (31 months

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and 25 months) between the two arms: Melacine plus low-dose IFN-α-2 versus high-dose IFN-α-2 alone. However, toxicity was lower in the combination group. Melacine is marketed only in Canada for the treatment of stage IV disease by Schering-Plough Canada (a subsidiary of Schering-Plough), which returned worldwide marketing rights for Melacine to Corixa in August 2002. Applications to the FDA and the European Agency for the Evaluation of Medicinal Products (EMEA) for stage IV disease have suffered setbacks, and in November 2003, Corixa opted to discontinue further development of this vaccine based on the FDA’s request for additional Phase III trials. The Phase III trial that formed the basis of the registration application in Canada showed that Melacine’s efÞcacy in stage IV disease is comparable to that of the combination regimen dacarbazine, cisplatin, carmustine, and tamoxifen (the Dartmouth regimen) and that Melacine’s safety and quality-of-life proÞle is superior (Mitchell MS, 1997). A different Phase III trial has shown that Melacine is effective in a subgroup of stage II patients. An initial analysis of this trial, which compared Melacine with observation in 689 patients, showed a signiÞcant improvement in diseasefree survival in patients who received Melacine: 30% of those patients relapsed or died, whereas 37% of the control patients relapsed or died. Patient response was found to be related to the human leukocyte antigen (HLA) phenotype; the 67% of patients treated with Melacine who had two or three of Þve prospectively deÞned HLA alleles had a longer period of disease-free survival than did control patients with the same number of matching alleles and Melacine patients who had fewer matching alleles (Sosman JA, 2001). M-Vax. M-Vax (Avax Technologies), a personalized vaccine, launched in Australia in 2000 for the treatment of stage III MM. However, in September 2002, Avax withdrew M-Vax because of Þnancial constraints faced by the company resulting from a delay in a reimbursement decision in Australia. In the United States, the FDA expressed concerns about the sterility of the fresh vaccine that was initially in development, and inspections of the manufacturing facility were undertaken. This concern led Avax Technologies to develop a frozen version of the vaccine. The company now states that no active clinical trials are under way with M-Vax, although M-Vax is available on a compassionate use basis in Australia. The data that supported the Australian approval were generated in 214 stage III MM patients and showed a Þve-year survival rate of 50% (among patients with one nodal area of involvement) and 35% (two areas of nodal involvement). These results were favorable compared with historical controls who underwent surgery alone, where Þve-year survival was only 20–25% (one nodal area of involvement) and 10% (two areas of nodal involvement). The drug has been launched in Switzerland in 2005; company is pursuing activity in EU and US. Melan-A/MART-1. The Ludwig Institute for Cancer Research (in collaboration with SEPPIC and GlaxoSmithKline Biologicals, which are both supplying vaccine adjuvants) has reported Phase II data for MelanA/MART-1 vaccine for

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the treatment of MM. MelanA/MART-1 is a melanoma-associated antigen, and the vaccine under development involves administering the peptide to patients, thereby enhancing their T-cell reactivity to the melanoma. In a Phase II trial, 37 patients with stage III/IV MM received one of seven different vaccine preparations, based on Melan-A (Cerottinni JP, 2003). Eight of the 15 patients who were tumor-free at entry remained so during the observation period. Of the 22 tumor-bearing patients, 18 showed unaltered progressive disease; two experienced disease stabilization; and two had regressing metastases of the skin, lymph nodes, and lung. After vaccination, 6 of 19 patients had activated tumor antigen–speciÞc T cells detectable ex vivo. We are not aware of any further clinical studies with MelanA/MART-1, HSPPC-96 (Vitespen). HSPPC-96 (Vitespen) (Antigenics’ Oncophage), a personalized peptide vaccine based on heat-shock protein (HSP)-96, is being tested in stage IV MM patients in an ongoing Phase III trial in the United States and Europe. The interaction of cancer-cell-speciÞc HSP-peptide complexes with the immune system is the basis for HSPPC-96 (Vitespen). When peptides are damaged—for example, by extremes of temperature—HSPs bind to them and ßag them for destruction. Cancer cells contain unique, damaged peptides that do not exist in other cells; HSP binds to these peptides and presents them on the cell surface. These cell-surface peptides act as antigens and enable the immune system to recognize the diseased cell and destroy it. According to Antigenics, destruction of the cell causes unbound HSP-peptide complexes to be released, thus amplifying the immune response. This ampliÞed immune response enables widespread destruction of all cells with the same surface antigen. To create the vaccine, surgeons remove a patient’s tumor and send a sample to Antigenics’ manufacturing plant for extraction of HSPs. Currently, seven grams of melanoma tissue are required for the process, although ongoing developments in the process will reduce this requirement in the future. The isolated HSPs are sterilized and used to produce a patient- and tumor-speciÞc vaccine that is administered to the patient to stimulate an immune response against any remaining tumor cells. HSPPC-96 (Vitespen) vaccine treatment begins after the patient has recovered from surgery (usually within four to eight weeks). The patient receives one injection of the personalized vaccine once per week for four weeks, then one further injection every other week. Treatment is designed to be given on an outpatient basis to minimize disruption of the patient’s everyday life. In a Phase II trial of 45 patients, 28 patients had residual measurable disease post-surgery. Of these 28 patients, 2 achieved a complete response, and 3 had long-term stable disease. Laboratory analysis of patients’ tumors and T cells have demonstrated that 5 of the 6 clinical responders (including the 2 complete responders and 3 patients with stable disease) but only 3 of 13 nonresponders displayed high expression of both class I HLA and melanoma antigens (Rivoltini G, 2002). These results conÞrm that vaccination of mMM patients with HSPPC-96 (Vitespen) is feasible. In addition, the trial shows that HSPPC-96 (Vitespen) is

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devoid of signiÞcant toxicity (no grade 3 or 4 toxicities were observed) and can induce clinical responses. In October 2005, Antigenics released data from a preliminary analysis of their randomized, international multicenter, open-label Phase III trial investigating HSPPC-96 versus a physician choice regimen in patients with stage IV melanoma, who were stratiÞed based on AJCC metastatic stage (M1a, M1b, and M1c). The endpoints of this trial, which was not intended for registration, include survival and safety. Preliminary data showed improved survival in patients with M1a melanoma who received HSPPC-96 versus the physicians’ choice. Antigenics is planning a Phase III trial for registration of HSPCC-96 in the M1a patient subgroup. The company will also be submitting results from potency testing of HSPPC-96 to the FDA inb March 2006. GM2-KLH/QS-21 Vaccine. GM2-KLH/QS-21 (Progenics Pharmaceuticals’ GMK) is a GM-2 ganglioside directed vaccine, combined with an adjuvant, in late-stage clinical development. GM2-KLH/QS-21 consists of the melanoma antigen ganglioside GM-2 (the most immunogenic ganglioside expressed on melanoma cells and one that is present on approximately 95% of melanoma tumors) conjugated to keyhole limpet hemocyanin (KLH) and combined with QS-21, a highly potent vaccine adjuvant. Although this vaccine has proved inferior to IFN-α-2b in stage IIB and stage III patients, a Phase III trial that compares GM2-KLH/QS-21 with observation in 1,300 resected stage II patients is under way in Europe and Australia, organized by the European Organization for Research into the Treatment of Cancer (EORTC) Melanoma Cooperative Group. Patient accrual is complete, and followup is continuing. A Phase III clinical study comparing GM2-KLH/QS-21 with high-dose IFN-α is also recruiting in Europe. Patients are being followed up for relapse and survival. Bristol-Myers Squibb held the rights to GM2-KLH/QS-21 but terminated the agreement in May 2001. A large U.S. Phase III trial sought to compare two years of therapy with GM2-KLH/QS-21 and one year of high-dose IFN-α-2b in stage IIB and stage III patients following tumor resection. This trial was terminated early, but an interim analysis showed that patients treated with IFN-α-2b experienced signiÞcantly superior survival (median follow-up of 16 months) (Kirkwood JM, 2001[b]). Patients who had an antibody response to GM-2 had better survival than those who did not, though the difference was not signiÞcant. Progenics continues to collect data on patients entered in this trial. Another small, randomized Phase II trial found that IFN-α does not diminish the immunologic response to GM2-KLH/QS21, indicating that the two agents could be used in combination (Kirkwood JM, 2001[a]). Immunologic response to GM2-KLH/QS-21—administered alone or in combination with high-dose IFN-α-2b—was compared in 107 fully resected stage IIB, III, and IV patients. Two of the three groups of patients were assigned to different IFN-α-2b regimens: in the Þrst group, IFN-α-2b and the vaccine were initiated at the same time; in the second group, IFN-α-2b was initiated one month after the Þrst vaccination. Results showed no signiÞcant difference in antibody responses among the three patient groups, and the presence of an

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immunologic response did not affect response to IFN-α-2b. A trend did favor improved disease-free survival in the two groups that received IFN-α-2b compared with the group that received vaccine alone, but the difference did not reach statistical signiÞcance in this small trial. GM2-KLH/QS-21 is administered subcutaneously once per week for 4 weeks, then once every 12 weeks for 92 weeks. The vaccine is well tolerated; no patients in the trial just described discontinued therapy. Local reactions at the injection site, liver enzyme abnormalities, and neurological toxicities were the most common grade 3 toxicities, and they occurred in less than 3% of patients. Despite the negative results of the terminated Phase III trial in stage IIB and III patients, many clinicians remain interested in GM2-KLH/QS-21. European clinicians are entering stage II patients into the ongoing randomized Phase III trial described earlier, and the potential of combining GM2-KLH/QS-21 with IFN-α-2 and additional peptide antigens is being investigated (Kirkwood JM, 2001[b]). The results of the European Phase III trial may be unavailable until midway through our forecast period because of the large number of patients and the long-term follow-up required to obtain robust, meaningful survival data. The potential of combining GM2-KLH/QS-21 with IFN-α-2 and additional peptide antigens is also being investigated (Kirkwood JM, 2001[b]). TriGem. (NOTE: most recent information indicates: the development has been suspended! ) Antisense Therapies Overview. To date, despite holding promise, no antisense therapy for the treatment of cancer has been approved. Mechanism of Action. Antisense oligonucleotides are short sequences of single-stranded DNA that can bind to a speciÞc region of corresponding messenger RNA (mRNA) sequence, thereby blocking both the expression and translation of the mRNA itself and the generation of the corresponding protein encoded by the mRNA. In this way, antisense molecules can block the expression of undesirable genes and their proteins. Oblimersen. Oblimersen (Genta and Aventis’s Genasense), an antisense oligonucleotide designed to block production of the Bcl-2 protein from bcl-2 (a proto-oncogene), is under development as an intravenous infusion for the potential treatment of various cancers including MM. Bcl-2 is known to inhibit cellular apoptosis (or programmed cell death), which can be a cause of resistance to cytotoxic drugs. The efÞcacy of cytotoxic agents may be augmented by inhibiting bcl-2. Genta submitted oblimersen to the FDA in September 2003, for fast-track review, for the treatment of MM. In December 2003, Genta completed the NDA Þling and applied for priority review status. On February 6, 2004, the FDA announced that it had accepted the NDA and priority review and will make a decision before June 8, 2004. In late April 2004, Genta and Aventis announced

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an expanded access program for eligible patients (those not previously treated with either dacarbazine or temozolomide) in the United States. However, In 2004, Genta withdrew its NDA in the US. In February 2006, Genta announced that it had conÞrmed the major efÞcacy variables from the randomized Phase III trial of Ganasense plus chemotherapy in patients with advanced metastatic melanoma in Europe. The data were presented in Paris by Dr. Alexander Eggermont of the EORTC. Updated results, including two years of follow up, from Genta’s pivotal Phase III trial have been further presented at recent international meetings. At ASCO in May 2005, the previously observed improvements in overall response rate (ORR) and progression free survival (PFS) in patients receiving dacarbazine plus oblimersen versus dacarbazine alone, were conÞrmed. Also in February of 2006, Genta announced that the EMEA had started its scientiÞc assessment of Genta’s MAA, which the company completed submission for in January 2006 (Press release, Genta, February 2006). The EMEA is expected to respond by June 2006. In May 2005, Genta had not yet made a decision on whether it would re-Þle in the United States. Alkylating Agents Overview. The alkylating agent dacarbazine is the only approved chemotherapeutic agent for the treatment of advanced MM. Although it received approval, this agent offers modest activity, yielding response rates of 5–20%, with only rare incidents of complete responses and no signiÞcant improvement in overall survival. Despite numerous studies of dacarbazine in combination with other chemotherapeutic agents or biological agents and non-dacarbazine-containing treatment regimens, nothing has proved superior to single-agent dacarbazine in randomized trials. Currently, temozolomide is under clinical evaluation for MM, in particular for patients with brain metastasis; temozolomide has already gained approval for brain tumors. Mechanism of Action. Alkylating agents methylate or alkylate DNA at speciÞc locations. Monomethyl triazeno imidazole carboxamide (MTIC), the active metabolite of both dacarbazine and temozolomide, methylates guanine at the O6 position, which is considered to be the primary toxic lesion that inhibits DNA replication and results in cell death. Temozolomide. Temozolomide (Schering-Plough’s Temodar) (Figure 7) is an alkylating agent that undergoes rapid chemical conversion at physiologic pH to the active compound MTIC. The cytotoxicity of MTIC is thought to result primarily from methylation of DNA at the O6 position of guanine, which prevents DNA replication. Unlike dacarbazine, temozolomide does not require metabolic activation by the cytochrome P450 and is an oral drug with reported 100% oral bioavailability. Temozolomide already has extensive off-label use for the treatment of mMM, particularly in Europe. A pivotal Phase III clinical investigation randomized 305 patients with advanced disease to receive either orally administered temozolomide (daily for

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FIGURE 7. Structure of temozolomide (R = CH3 ).

5 days every 28 days—the schedule used for temozolomide’s approved indication of refractory anaplastic astrocytoma) or dacarbazine (dosed daily for 5 days every 21 days) (Middleton MR, 2000). The median progression-free survival was superior in the temozolomide arm: 1.9 months versus 1.5 months for the dacarbazine arm, despite equivalent response rates (13.5% and 12.1% for temozolomide and dacarbazine, respectively). However, the increase in overall median survival, 7.7 months versus 6.4 months, was not statistically signiÞcant. The authors concluded that temozolomide exhibits efÞcacy equivalent to that of dacarbazine and provides a useful oral alternative to dacarbazine. Temozolomide exhibits extensive central nervous system (CNS) penetration and has recently (in March 2005) received FDA approval for the treatment of high-grade gliomas. Studies of temozolomide in advanced MM suggest a response rate in systemic disease comparable to that seen with dacarbazine (15–20%). Current clinical trials are evaluating the role of temozolomide in the prevention and treatment of MM CNS metastases—an increasingly common and important clinical challenge that occurs in up to half of mMM patients. A retrospective study compared the incidence of CNS relapse in mMM patients who had responded to treatment with dacarbazine or temozolomide. The study found signiÞcantly fewer CNS relapses in the temozolomide group (2/19 patients) than in the dacarbazine group (8/21) (Summers Y, 1999). In an attempt to improve the rate and durability of response to temozolomide, investigators have initiated a randomized Phase III clinical trial with extended treatment schedules involving six weeks of daily therapy followed by a two-week rest period. The rationale behind this extended treatment regimen, which is already showing efÞcacy in brain tumors, is that it reduces the recovery time needed to renew the DNA repair enzyme, O6 -alkylguanine-DNA alkyltransferase (AGT). This enzyme is responsible for the repair of the O6 methylation lesions of DNA produced by temozolomide. A further pharmacological maneuver is the development of agents, such as benzyl guanine, that inhibit AGT and have the potential to augment temozolomide efÞcacy. Several clinical trials are investigating the combination of temozolomide and thalidomide (Celgene’s Thalomid; discussed later on). Thalidomide is a immunomodulator that inßuences several cellular activities and exerts antiangiogenic effects. At ASCO 2003, researchers presented the results of two clinical trials evaluating temozolomide and thalidomide in advanced MM. One Phase II clinical study

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involved 38 chemotherapy-naive patients with stage III or stage IV MM (Hwu W, 2003). Temozolomide was administered at 75 mg/m2 daily for six weeks followed by a two-week break. Patients younger than age 70 received 200 mg of thalidomide daily. This dose was increased by 100 mg every two weeks to a maximum dose of 400 mg. Patients aged 70 or older started daily thalidomide at 100 mg with dose escalation of 50 mg every two weeks to a maximum dose of 250 mg. Response was evaluated every eight weeks. Of the 38 patients evaluated, 10 achieved partial response, 4 achieved a minor response, 8 experienced stable disease, 3 showed a mixed response (response at some sites of disease and progression at others), and 13 patients’ disease progressed. The second Phase II clinical trial presented at ASCO 2003 recruited 181 patients with metastatic MM and compared treatment with temozolomide and thalidomide, temozolomide and IFN-α-2, or temozolomide alone. Higher response rates and disease stabilization were observed in the combination arms—23.1% of patients treated with temozolomide plus thalidomide, 20.4% of patients treated with temozolomide plus IFN-α-2, and 13.7% of patients treated with temozolomide alone. A retrospective study compared the incidence of CNS relapse in metastatic patients who had responded to treatment with dacarbazine or temozolomide. The study found signiÞcantly fewer CNS relapses in the temozolomide group (2/19 patients) than in the dacarbazine group (8/21) (Summers Y, 1999). Another Phase II study was presented at the Chemotherapy Foundation Symposium, New York, Nov. 2003. See Journal of Clinical Oncology, 24(3);2006: 401–406.} As described earlier, the tolerability of temozolomide is similar to that of dacarbazine. The adverse effects seen most frequently with temozolomide are nausea (52%), vomiting (34%), pain (34%), and constipation (30%); in most cases, these effects are not severe. Clinically relevant hematologic toxicities occur in approximately 10% of patients. Immunomodulators Overview. Owing to the established role of the immune system in the development and treatment of MM, conÞrmed by the (albeit modest) efÞcacy of both IFN-α and IL-2, clinicians and developers have continued their interest in this approach to treating MM. Clinicians reportedly are investigating granulocytemacrophage colony-stimulating factor (GM-CSF) as adjuvant treatment; if trials demonstrate improved activity or toxicity over IFN-α-2, the uptake of GM-CSF will be widespread. Mechanism of Action. Immunomodulators exert their effects on the immune system through many targets and mechanisms, a majority of which are poorly understood. Immunomodulators may have direct effects on immune cells (e.g., T cells) or may synergize or antagonize the effects of immune-active cytokines. Pegylated Interferon-Alpha-2. Interferon-alpha-2b (IFN-α-2b; ScheringPlough’s Intron A) has been approved for adjuvant use in MM, and IFN-α-2a (Roche’s Roferon A) enjoys extensive off-label use for the same indication.

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Owing to the inherent difÞculties of maintaining active plasma levels of IFN-α-2 and the marked toxicity proÞle of the agent, both Schering-Plough (PEG-Intron) and Roche (Pegasys) have launched pegylated (polyethylene glycol–conjugated) versions of their drugs for hepatitis C and are investigating them for the treatment of MM. The role of polyethylene glycol (PEG) is to extend the plasma half-life of interferon by reducing its renal clearance and to eliminate the peaks and troughs of IFN concentration that are associated with three-times-weekly dosing of nonpegylated IFN-α. The IFNs are a family of naturally occurring cytokines that exhibit antiviral, antitumor, and immunomodulatory activities. The antitumor effect of IFNs derives from their ability to inhibit cell-cycle progression and to induce a reduction in tumor-cell protein synthesis. Their immunomodulatory role involves inducing the expression of major histocompatibility antigens and modulating the expression and function of T cells, monocytes, and natural killer cells (large granular lymphocytes that bind to and kill cells by releasing toxins). Clinical data from MM patients are not yet available. Phase I/II data, generated from 35 patients, (primarily with renal cell carcinoma), show that a once-weekly dose administration was active and well tolerated when given for up to one year (Berg W, 2000). The most common nonhematologic adverse events included mild to moderate nausea, anorexia, and fatigue. Six patients had grade 3 or 4 hematologic toxicity. At the time of composing this reference, recruitment is being conducted for the Phase III trial. Higher plasma concentrations of IFN-α can clearly be achieved by administering the pegylated form, with no apparent detriment in toxicity proÞle. However, the widespread skepticism of clinicians regarding the efÞcacy of IFN-α in MM will hamper the development of the pegylated form for this indication. If convincing efÞcacy can be achieved with the pegylated version, with acceptable toxicity, clinicians will certainly prescribe it for a disease badly lacking therapeutic options. GM-CSF. Recombinant GM-CSF (sargramostim; Berlex’s Leukine), a macrophage and monocyte activator, is in clinical trials for the adjuvant treatment of MM. An open-label, Phase II trial of GM-CSF as surgical adjuvant therapy was conducted in patients with high-risk (stage II [T4], III, or IV) melanoma that had been surgically excised, rendering the patients clinically tumor-free. GM-CSF was administered daily subcutaneously at 125 mg/m2 for 14 days, followed by 14 days off therapy, and was continued either until the tumor recurred, requiring systemic therapy, or until the patient had been tumor-free for three years. An interim analysis of 50 patients was conducted in February 2002 (Spitler L, 2000). Treatment was well tolerated. The major side effects were injection-site reactions, which occurred in more than half of patients. To evaluate survival, patients were matched by stage with 1,000 patients in the American Joint Committee on Cancer (AJCC) database. Preliminary results suggested a survival beneÞt for the patients treated with GM-CSF (p = 0.0005). The value of historical controls for comparison is questionable, and prospective randomized studies are ongoing to address this issue.

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O N O

O

NH

O

FIGURE 8. Structure of thalidomide.

Thalidomide. Thalidomide (Celgene’s Thalomid) (Figure 8) was licensed by the FDA in 1998 for the acute treatment of an inßammatory condition associated with leprosy and is in Phase II clinical trials for MM. It has been approved for use in relapsed and refractory multiple myeloma in Australia and is under review for this indication in several other markets. According to the data available at the time of composing this reference, it is still in Phase II trial. Thalidomide has several mechanisms of action; those that are important to its anticancer activity remain uncertain and controversial. Thalidomide possesses several immunomodulatory activities: it is potentially synergistic with GM-CSF, enhances IL-2 activity, induces and inhibits T-cell stimulation and proliferation, and suppresses TNF-α and downregulated IL-6. Thalidomide also inhibits angiogenesis induced by beta Þbroblast growth factor (β-FGF) and VEGF and has antimetastatic properties. A Phase I/II study in 11 high-risk MM patients, reported at the ASCO meeting in May 2003, provides evidence that thalidomide in combination with GM-CSF is well tolerated (Lutzky J, 2003). At the same conference, researchers presented the results of two clinical trials evaluating temozolomide and thalidomide in advanced MM. One Phase II clinical study involved 38 chemotherapy-naive patients with stage III or stage IV MM. Temozolomide was administered at 75 mg/m2 daily for six weeks followed by a two-week break. Patients younger than age 70 received 200 mg of thalidomide daily. This dose was increased by 100 mg every two weeks to a maximum dose of 400 mg. Patients aged 70 or older started daily thalidomide at 100 mg with dose escalation of 50 mg every two weeks to a maximum dose of 250 mg. Response was evaluated every eight weeks. Of the 38 patients evaluated, 10 achieved a partial response, 4 achieved a minor response, 8 experienced stable disease, 3 showed a mixed response, and 13 patients’ disease progressed. The second Phase II clinical trial presented at ASCO 2003 recruited 181 patients with mMM and compared treatment with temozolomide and thalidomide, temozolomide and IFN-α, or temozolomide alone. Higher response rates and disease stabilization were observed in the combination arms—23.1% of patients treated with temozolomide plus thalidomide, 20.4% of patients treated with temozolomide plus IFN-α-2, and 13.7% of patients treated with temozolomide alone (Delva R, 2003). For further study results, see attached (JCO 24(3);2003:401–406. Thalidomide is being extensively studied in MM, primarily in combination with other agents. The company’s newer agent, CC-5013, is 1,000 times more

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potent in vitroand is a more selective TNF-α inhibitor. CC-5013 is currently undergoing clinical investigation. However, results from thalidomide trials serve as useful proof of principle for this class of agents. CC-5013. CC-5013 (Celgene’s Revimid) is in Phase III studies for MM; enrollment into these trials was completed in January 2004; although by some indications the monotherapy melanoma trials in Phase III were terminated, due to lack of efÞcacy. CC-5013 acts by inhibiting TNF-α. It is an orally active analogue of thalidomide, but it features a more favorable preclinical activity and toxicity proÞle than the parent drug. A two-center Phase I study was performed to assess the tolerability and preliminary efÞcacy of CC-5013 (Liu Y, 2003). In this study, 21 patients with advanced MM, 5 with renal-cell carcinoma, and 10 patients with other solid tumors received oral CC-5013. T-cell surface markers and levels of cytokines were studied in venous blood. CC-5013 was well tolerated, and no serious adverse events were deÞnitely attributed to treatment. Myelosuppression was observed in 13 patients, including grade 4 neutropenia in 2 patients. All patients tested showed evidence of T-cell activation. Compared with baseline values, treatment was associated with signiÞcantly increased serum levels of GM-CSF, IL-2 receptor, and TNF-α. Partial responses were observed in three patients with MM. This study indicates the tolerability and potential clinical efÞcacy of CC-5013 in advanced MM. The demonstration of T-cell activity has enabled the early demonstration of proof of principle for this agent and may expedite its development. CC-5013 may be used as Þrst-line treatment and possibly in the adjuvant setting, where its oral administration will be viewed as a real beneÞt over current treatments. Histamine Dihydrochloride. In Phase III trials in the United States and Europe, histamine dihydrochloride (Maxim’s Ceplene) is the only histamine H2 receptor agonist identiÞed in clinical development for MM. Maxim Pharmaceuticals announced in November 2003 that it had Þled a European centralized procedure marketing authorization application for histamine dihydrochloride, in combination with IL-2, for the treatment of patients with mMM. Histamine and its analogues are known to stimulate cytokine release and enhance immune response. Histamine dihydrochloride is an immune-modulating histamine H2 receptor agonist that can prevent the production and release of oxygen free radicals. It has a cytoprotective effect, preventing or reversing damage induced by oxidative stress. Histamine dihydrochloride can enhance and support cytokine activity and is being tested as an adjunct to IL-2 therapy and/or IFN therapy. A Phase III clinical trial was conducted to study 305 mMM patients in 56 U.S. centers who received low-dose IL-2 with or without histamine dihydrochloride between July 1997 and March 1999 (Agarwala SS, 2002). Data from this study have been used to support the November 2003 application for European marketing authorization. At three years of follow-up, patients treated with the

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two agents showed a superior but not signiÞcant survival duration. Median survival was 9.1 months for the combination arm and 8.2 months for the IL-2 alone arm. However, disease-free progression was statistically superior for the combination arm. A subgroup analysis of patients with liver metastases also demonstrated significantly improved survival compared with single-agent IL treatment: 9.4 months versus 5.1 months, respectively (Agarwala SS, 2002). The study shows a low incidence of toxicities and a signiÞcantly improved quality of life for the patients treated with histamine and IL-2. Two-year survival rates for patients with liver metastases were 18.2% for the group treated with the histamine dihydrochloride combination versus 2.7% for the group treated with IL-2 alone. A further Phase III study, in the United States and Europe, had recruited 225 MM patients whose disease had metastasized to the liver. This randomized trial is designed to compare patient survival with the combination treatment of histamine dihydrochloride and IL-2 versus IL-2 alone and is expected to conclude in the second half of 2004. Given the lack of active agents for advanced MM and histamine dihydrochloride’s favorable toxicity proÞle, this agent is likely to obtain approval in both the United States and Europe for use in combination with IL-2, once data from the ongoing trial are available. Histamine dihydrochloride will be launched for MM patients with liver metastases, a poor prognosis group accounting for approximately 20% of stage IV patients. RAF Kinase Inhibitors Overview. RAF and other members of the mitogen-activated protein kinase (MAPK) pathway (such as RAF, MEK, and ERK) are attractive targets for the development of new anticancer agents. Aberrant RAF and RAS activity has been demonstrated in a variety of cancers and can result in deregulation of cellular proliferation, differentiation, or apoptosis. Mutations in the kinase domain of Braf have been reported in approximately 70% of mMMs (Davies H, 2002). Pharmaceutical companies have shown substantial interest in targeting kinaseregulated signaling pathways, including the MAPK pathway, as a novel approach to cancer treatment. The approval of the kinase inhibitors imatinib (Novartis’s Gleevec) for breast cancer and geÞtinib (AstraZeneca’s Iressa) for non-small-cell lung cancer has further fueled this interest. Mechanism of Action. RAF kinase inhibitors inhibit the RAF kinase constituent of the MAPK signaling cascade. This cascade causes inappropriate cell signaling in a large number of tumors, which results in cell proliferation. Sorafenib (BAY 43–9006). Bayer and Onyx Pharmaceuticals are developing Sorafenib, an oral cytostatic RAF kinase inhibitor, which also has VEGF-inhibitory activity, for the potential treatment of cancer. Sorafenib is the Þrst compound to target both the RAF/MEK/ERK signaling pathway to inhibit cell proliferation and the

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Sharma RA, et al. Tolerability of the novel oral thalidomide analog CC-5013 demonstrating extensive immune activation and clinical response. Proceedings of the American Society of Clinical Oncology. 2003;(22). Abstract 2862. Sherr CJ. CDK inhibitors: positive and negative regulators of G1-phase progression. Genes Development. 1999;13:1501–1512. Slingluff CL, et al. Surgical management of regional lymph nodes in patients with melanoma. Experience with 4,682 patients. Annals of Surgery. 1994;219:120–130. Sorber AJ, et al. Guidelines of care for primary cutaneous melanoma. Journal of the American Academy of Dermatologists. 2001;45:579–586. Sosman JA, et al. SigniÞcant impact of HLA class I alleles on outcome in T3N0 melanoma patients treated with Melacine (MEL): an allogeneic melanoma cell lysate vaccine: prospective analysis of Southwest Oncology Group (SWOG)-9035. Proceedings of the American Society of Clinical Oncology. 2001;(20). Abstract 1402. Spitler LE, et al. Adjuvant therapy of stage III and IV malignant melanoma using granulocyte-macrophage colony-stimulating factor. Journal of Clinical Oncology. 2000;18:1614–1621. Spitler LE, et al. Adjuvant therapy of stage III and IV melanoma using granulocytemacrophage colony-stimulating factor (GM-CSF). Proceedings of the American Society of Clinical Oncology. 2001;(20). Abstract 1429. Stang K, et al. Descriptive epidemiology of cutaneous malignant melanoma. Analyses of German Cancer Registry Data. In: Hasman A, et al., eds. Medical Infobahn for Europe. Amsterdam, the Netherlands: IOS Press; 2000. Stang A, et al. Skin melanoma in Saarland: incidence, survival and mortality 1970–1996. European Journal of Cancer Prevention. 2001;10:407–415. Stang A, et al. Site- and gender-speciÞc time trend analyses of the incidence of skin melanomas in the former German Democratic Republic (GDR) including 19,351 cases. European Journal of Cancer. 2003;39:1610–1618. Summers Y, et al. Effect of temozolomide (TMZ) on central nervous system (CNS) relapse in patients with advanced melanoma. Proceedings of the American Society of Clinical Oncology. 1999;(18). Abstract 2048. Swerdlow AJ, et al., eds. Cancer incidence and mortality in England and Wales: trends and risk factors.Oxford, U.K: Oxford University Press; 2001. Thurner B, et al. Vaccination with mage-3A1 peptide-pulsed mature monocyte-derived dendritic cells expands speciÞc cytotoxic T cells and induces regression of some metastases in advanced stage IV melanoma. Journal of Experimental Medicine. 1999;190; 1669–1678. Tsao H, et al. PTEN expression in normal skin, acquired melanocytic nevi, and cutaneous melanoma. Journal of the American Academy of Dermatology. 2003;49(5):865–872. [a] Tsao H, et al. The transformation rate of moles (melanocytic nevi) into cutaneous melanoma. Archives of Dermatology. 2003;139:282–288. [b] United Nations Population Division. Sex and Age Quinquennial 1950–2050 (The 1998 Revision).New York, NY: The United Nations, 1998. Veronesi U, et al. Thin stage I primary cutaneous malignant melanoma, comparison of an excision with margins of 1 or 3 cm. New England Journal of Medicine. 1988;318: 1159–1162.

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Wagner JD, et al. Current therapy of cutaneous melanoma. Plastic and Reconstructive Surgery. 2000;105:1774–1799. Wang F, et al. Phase I trial of a MART-1 peptide vaccine with incomplete Freund’s adjuvant for resected high-risk melanoma. Clinical Cancer Research. 1999:5;2756–2765. Wheatley K, et al. Interferon-a as adjuvant therapy for melanoma: a meta-analysis of the randomised trials. Proceedings of the American Society of Clinical Oncology. 2001;(20). Abstract 1394. Yamazaki N, et al. Dacarbazine, nimustine hydrochloride, cisplatin and tamoxifen combination chemotherapy for advanced malignant melanoma. Journal of Dermatology. 1999;26:489–493. Yang Y, Becker D. Antisense targeting of basic Þbroblast growth factor and Þbroblast growth factor receptor-1 in human melanomas blocks intratumoral angiogenesis and tumor growth. Nature Medicine. 1997;3:887–893.

Multiple Myeloma

ETIOLOGY AND PATHOPHYSIOLOGY Introduction Multiple myeloma (MM) is a progressive hematologic cancer characterized by the proliferation of malignant plasma cells, called myeloma cells (MCs), which inÞltrate the bone marrow (BM) at sites throughout the skeleton. This inÞltration causes BM failure, resulting in anemia and reduced immune function. In MM, MCs are normally conÞned to the BM, homing to the BM microenvironment (BMM) as a result of several interactions with cytokines, growth factors, and other cells within this environment. MCs cause abnormally high amounts of osteolysis (bone resorption), leading to disease-deÞning pathological fractures, bone pain, and high serum calcium (hypercalcemia). MM is always fatal and is associated with serious complications, including immune system failure. It is the most common primary cancer of the bone in adults. Pathophysiology Normal Plasma Cells. Normal, nonmalignant plasma cells play a key role in the humoral immune response, as summarized in Figure 1. These cells produce large quantities of antibodies that are essential to the identiÞcation and destruction of non-self cells and proteins (e.g., bacteria). The invading organism’s antigens interact with a corresponding receptor on a B lymphocyte. This association is followed by a complex sequence of interactions with other cells and cytokines to Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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FIGURE 1. Plasma cell and myeloma cell process.

produce a set of plasma cells that secrete an unbound copy of the B-cell receptor, or antibody. Antibodies, or monoclonal immunoglobulins, bind to the antigen on the offending cell, ßagging it for destruction by other parts of the immune system. Immunoglobulins can be produced by both normal and malignant plasma cells. Immunoglobulins are roughly Y-shaped; the stem of the Y is called the constant region, as shown in Figure 2. The two arms of the Y, called the variable

574

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FIGURE 2. Antibody immunoglobulin.

region, vary greatly between antibodies. These unique variations in shape of the antigen-binding site allow the antibody to bind to a matching antigen. Each immunoglobulin is made up of four polypeptide chains: two heavy and two light. The Þve classes of heavy chain (mu, delta, gamma, alpha, and epsilon) determine the Þve major classes of functional immunoglobulin: IgM, IgD, IgE, IgA, and IgG, the last of which is overwhelmingly the most abundant immunoglobulin. The two types of light chain are lambda and kappa. An immunoglobulin has either kappa or lambda chains, never a combination of the two. The ratio of the occurrence of the two chain types is normally two to one (kappa to lambda) in humans (Janeway CA, 2004). Distortion of this ratio or, indeed, circulation of free chains implies an abnormal proliferation of B cells. Monoclonal Gammopathies. Monoclonal gammopathies are a group of disorders characterized by monoclonal proliferation of plasma cells that produce a speciÞc immunoglobulin (International Myeloma Working Group, 2003). The monoclonal immunoglobulin is referred to as the M-protein. MM is just one type of monoclonal gammopathy at the opposite end of the spectrum from normal plasma cells. As normal cells acquire genetic mutations, they progress to monoclonal gammopathy of undetermined signiÞcance (MGUS), through asymptomatic myeloma, to symptomatic myeloma. The characteristics of these plasma cell disorders are outlined in Table 1.

TABLE 1. Conditions Related to Multiple Myeloma

Disease Monoclonal gammopathy of undetermined significance (MGUS) Asymptomatic multiple myeloma Symptomatic multiple myeloma Solitary bone plasmacytoma Extramedullary plasmacytoma

M-Protein Serum <3 g/dL

Serum >3 g/dL Serum/urine In 24–54% of cases No

a Kyle RA, 2002. b DeVita VT Jr., 2001. c Woodruff R, 1979.

Ig = Immunoglobulin; N.A. = Not applicable. Full source citations appear in ‘‘References.’’

Circulating Plasma Cells

Patients Progressing to MM in Ten Years (%)

Anemia

Renal Failure

Hypercalcemia

Lytic Lesions

<10%

No

No

No

No

<2 g/dL <3.5 g/dL

12a

10%

No

No

No

No

100b

>10%

Yes

Yes

Yes

Yes

No

No

No

No

One

No

No

No

No

No

<5 g/dL <7 g/dL >2 g/dL >3.5 g/dL N.A. N.A. N.A. N.A.

IgA/IgG

— ≤10c ≤10c

575

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MULTIPLE MYELOMA

Monoclonal Gammopathy of Undetermined Significance. MGUS is characterized by low plasma cell numbers and no overt tumor mass. No bone damage occurs, and blood counts tend to be normal. At this stage, patients have no symptoms. MGUS is reported to be present in 1% of the adult population. Of the patients diagnosed with MGUS, 1% per year progress to MM (Kyle RA, 2002); therefore, patients with MGUS require long-term follow-up. Asymptomatic Myeloma. This disorder is an intermediate condition between MGUS and symptomatic MM. M-protein levels and plasma cell numbers are elevated with respect to MGUS, but the patient does not experience anemia, renal insufÞciency, or bone damage. This disorder is sometimes referred to as smoldering MM . Symptomatic Multiple Myeloma. MCs proliferate in the BM and often invade the adjacent bone. The skeleton is destroyed from the inside, resulting in bone pain and fractures. Other symptoms include anemia, renal insufÞciency, and hypercalcemia. The monoclonal immunoglobulin is IgG in approximately 50% of MM patients and IgA in another 20% of patients. Their presence conÞrms the diagnosis and severity of MM. The higher the level of M-protein, the more advanced the disease. The class of immunoglobulin produced can inßuence the pathology of the disease: patients with IgA MM have a worse prognosis than IgG patients. In 15-20% of all patients, more kappa or lambda light chains are produced than can be combined with heavy chains. The excess light chains are small enough to be excreted and pass into the urine. When these protein chains are detected in the urine, they are called Bence-Jones proteins. BenceJones proteins may deposit in the kidney, putting these patients at greater risk of renal failure. Approximately 3% of symptomatic MM patients have nonsecretory myeloma, which means that the M-protein cannot be detected in the serum or urine (International Myeloma Working Group, 2003). Solitary Bone and Extramedullary Plasmacytoma. This tumor is a solitary mass of plasma cells. The plasma cells can be attached to bone, as in solitary bone plasmacytoma, or exist as an extramedullary plasmacytoma of the soft tissue. Detection of M-proteins in serum and urine is not always possible because they are generally present in low levels or, in some cases, not at all. Both bone and extramedullary plasmacytomas are normally removed by surgery and/or eradicated with radiotherapy; such treatment results in the disappearance of any M-proteins present. However, disease can recur or progress to disseminated disease or MM. The number of patients who progress to MM is difÞcult to quantify because plasmacytomas are indistinguishable from occult MM and because treatment is effective and survival exceeds ten years, thus making follow-up difÞcult. Given the difÞculties in predicting progression, this report assumes a progression rate of no more than 10% in ten years (see Table 1). Cytogenetic Changes. Multiple genetic abnormalities are required for the development of MCs. These genetic abnormalities prevent the differentiation

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and normal death of the MCs, which continue to proliferate and accumulate in the BM. Translocation of immunoglobulin genes is the mechanism of disease progression in MM. The proportion of MGUS patients who have two or more immunoglobulin translocations is approximately 5%, rising to 27% in patients with advanced MM (Kuehl M, 2005). A gene translocation is a coding section for a particular gene that has been transposed to an incorrect position on the same or different chromosome. This transposition leads to activation of oncogenes. Table 2 shows the common cytogenetic changes that occur during the progression from MGUS to MM, together with their prognostic signiÞcance; these events can be further divided into early and late events. Furthermore, approximately half of patients have hyperdiploidy, which means that they have three copies of some chromosomes rather than the normal two copies. These patients may have up to 75 chromosomes instead of the normal karyotype of 46 (Kuehl M, 2005). Early Events. Translocations between the immunoglobulin heavy-chain locus chromosome 14q32 and one of Þve partner loci (11q13, 4p16.3, 16q23, 6p21, and 20q11) play a role in the development of MM in about 50% of MM patients (Kyle RA, 2004). Table 2 outlines the consequences of these translocations at the molecular level. For example, a translocation at 4p16 causes expression of Þbroblast growth factor receptor 3 (FGFR3), a tyrosine kinase receptor not normally expressed by plasma cells. Expression of this receptor encourages tumor growth (Dalton W, 2001). Translocation at 16q23 (which codes for the c-maf gene) leads to disruptions in the basic cellular processes, including proliferation, differentiation, and response to growth factors (Dalton W, 2001). The deletion of certain sequences or parts of chromosomes has an important role in gene expression. In some cases of MM, one arm of chromosome 13q is deleted; the result is the inactivation of critical tumor suppressor genes. Patients with this chromosomal deletion have a shorter life expectancy after chemotherapy (Drach J, 2000). Late Events. Termed oncogenic, late events are detected in patients with advanced disease. For example, the N - and K-ras mutations are absent in MGUS, but they are present in patients who have developed MM and occur with greater frequency in late-stage disease (Drach J, 2002). N - and K-ras mutations can result in growth-factor-independent proliferation of MCs. Other late-stage oncogenic events, such as translocations in the c-myc genes and abnormalities of p53, also affect the progression of MM but to an unclear extent. Future clariÞcation of the cytogenetics of MM will lead to the development of new targeted therapies. Detection of these genetic abnormalities and understanding of their impact on patient survival will enable clinicians to treat aggressive disease with aggressive therapy. Epigenetic Changes. In all cells, DNA is packaged in chromatin, a highly organized and compact nucleoprotein structure. Chromatin structure is dynamic and has important roles in the regulation of transcription. The fundamental

578

MULTIPLE MYELOMA

TABLE 2. Molecular Biology of Multiple Myeloma Chromosomal Instability

Frequency in Patients

Significance of Mutation

50% (MGUS) 60–75% (MM)

Translocations involving this region of this chromosome result in deregulation of genes that function as oncogenes and growth factors.

Seidl S, 2003

14q32 translocation partners 11q13 cyclin D1

15% (MM)

Seidl S, 2003

4p16

FGFR-3

15% (MM)

16q23

c-maf

5% (MM)

6p21

cyclin D3

3% (MM)

20q11

mafB

2% (MM)

8q24

c-myc

3% (MM)

Most frequent translocation partnered with 14q32; cyclin D1 regulates the cell growth and differentiation cycle. FGFR-3 promotes myeloma cell proliferation and prevents apoptosis. c-maf is a transcription factor that stimulates cell cycle progression but also promotes pathological interactions between MCs and BMM. cyclin D3 regulates the cell growth and differentiation cycle. mafB is a transcription factor that inhibits differentiation without affecting cell proliferation. Patients who overexpress the c-myc gene have more aggressive disease. N.A. Deletion of chromosome 13q suggests a poor prognosis and a poor response to standard treatment.

Early events 14q32

Rare partners 13q

Gene IgH

17% (MM) 15–20%

Reference

Seidl S, 2003

Seidl S, 2003

Seidl S, 2003

Kyle RA, 2004 [b]

Kuehl M, 2005

Kuehl M, 2005 Drach J, 2000; Dalton W, 2001

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TABLE 2. (continued) Chromosomal Instability

Gene

Frequency in Patients

Late events 8q24

c-myc

40% in advanced disease

18q21

bcl-2

Variable

17p13

p53

20–40%

1p11–p13 and 12p12 N- and K-ras 49% at relapse

Significance of Mutation Occurs with greater frequency in advanced disease; patients who overexpress c-myc gene have more aggressive disease. These genes may play a role in myeloma cell proliferation because bcl-2 prevents apoptosis. p53 affects the progression of MM. N- and K-ras play a significant role in the development and perpetuation of MM. The ras oncogene product, p21, can be measured; if it is found in high concentrations, then the patient has a poor prognosis.

Reference Drach J, 2000; Dalton W, 2001

Drach J, 2002

Dalton W, 2001 Dalton W, 2001

N.A. = Not applicable. Full source citations appear in ‘‘References.’’

unit of chromatin is the nucleosome, which consists of DNA wound around a core of histone proteins. The amino-terminal amino acid residues of histone proteins—known as histone “tails”—can be post-translationally modiÞed by acetylation or methylation. The methylation patterns of DNA are also regulated. Hypermethylation inactivates genes, whereas hyperacetylation is associated with increased gene expression. These epigenetic effects on gene expression are potentially modiÞable with drugs, unlike the cytogenetic alterations previously discussed (Seidl S, 2003). Epigenetic events in MM include hypermethylation of genes such as p16/INK4A and p15/INK4B that inhibit G1 progression of the cell cycle. These mutations have been reported in approximately 75% of MM patients (Seidl S, 2003). Myeloma Cells. Analysis of the cell-surface signaling molecules of normal plasma cells shows that they usually express high levels of CD38 and CD138. They do express CD19 but not CD56 (Bataille R, 1997). Unlike normal plasma cells, MCs show characteristic features of long-lived BM cells: they generally

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MULTIPLE MYELOMA

express CD38, CD56, and CD138 but not usually CD19 (Bataille R, 1997; Seidl S, 2003). Deregulation of four major signaling pathways inside normal cells—the JAK/STAT, phosphatidylinositol 3-kinase/Akt (PI3-K/Akt), ras, and NF-κB pathways—is associated with MM. Signaling molecules in these pathways represent therapeutic targets, and novel agents are in development to exploit these signaling cascades (see “Emerging Therapies”). For example, mammalian target of rapamycin (mTOR), which is activated by oncogenic Ras protein, is being explored by some investigators as a therapeutic target. Bone Marrow Microenvironment. MCs must interact with the BMM to survive and proliferate. Figure 3 illustrates the steps involved in this process. Interference with components of this process is a target of emerging therapies (see “Emerging Therapies”). Cytokines and Adhesion Molecules. Adhesion of MCs to other cells or structures of the BM stroma (stroma are the functional units/cells of the BM) leads to further activation of speciÞc adhesion molecules. This positive feedback system is maintained by the secretion of interleukin (IL)-6 from both myeloma and stromal cells. The secretion of IL-6 by myeloma and stromal cells is a result of their interaction via vascular cell adhesion molecule-1 (VCAM-1). VCAM-1 is further ampliÞed by the secretion of tumor necrosis factor-alpha (TNF-α). These interactions result in the release of numerous other cytokines and growth factors, thus supporting the MCs’ proliferation and preventing their apoptosis (Drach J, 2000). IL-6 secretion may also be triggered by activation of the CD40 cell surface receptor (Urashima M, 1995). IL-6 is an important cytokine that promotes the survival of MCs and upregulates the expression of two anti-apoptotic genes: bcl-XL and mcl-1 (Puthier D, 1999). The BMM also produces the signaling molecule insulin-like growth factor1 (IGF-1). Molecules within the IGF-1 receptor (IGFR-1)/phosphatidylinositol 3-kinase/Akt (PI3-K/Akt) signaling pathway regulate cell survival and can protect MCs from death (Mitsiades CS, 2004[a]). Angiogenesis. Angiogenesis (new blood vessel formation) is vital for the survival of the tumor mass. High levels of neovascularization can be associated with other adverse prognostic factors, such as enhanced proliferation and the deletion of chromosome 13q (Drach J, 2002). MCs secrete vascular endothelial growth factor (VEGF), levels of which are further increased by interaction between the myeloma and stromal cells. Basic Þbroblast growth factor (bFGF) is also detectable in elevated amounts in MM patients (Vacca A, 1999). The secretion of these growth factors supports the development of new blood vessels, thus providing the MCs with the nutrients and oxygen they need to survive. In addition, VEGF promotes MCs’ proliferation and migration to the BMM. Researchers hope that a better understanding of these mechanisms will lead to breakthrough treatments for MM.

ETIOLOGY AND PATHOPHYSIOLOGY

FIGURE 3. The bone marrow microenvironment.

581

582

MULTIPLE MYELOMA

Bone Resorption. Bone resorption and deposition occur continually during normal homeostatic maintenance of the skeleton and of circulating free calcium levels. This process is mediated by osteoblasts, which make bone deposits, and by osteoclasts, which remove bone. In the healthy adult, bone mass remains constant, suggesting that the activity of osteoblasts and osteoclasts is equal. In MM, this balance is disturbed by an increased number of osteoclasts. Increased osteoclast activity results in hypercalcemia and fractures. The interaction between MCs and stromal cells is associated with the secretion of numerous cytokines, including IL-1β and TNF. These cytokines function as osteoclast-activating factors (Drach J, 2000; Greipp R, 1992). Two molecules of the TNF receptor ligand superfamily, osteoprotegerin (OPG) and its ligand (OPGL), are critical elements in the regulation of osteoclast activity (Anderson DM, 1997; Wong BR, 1999). Overproduction of OPGL—also known as TNFrelated activation-induced cytokine (TRANCE) and as receptor activator of nuclear factor kappa B ligand (RANKL)—causes activation and differentiation of osteoclasts directly by binding to its receptor (RANKL) on osteoclastic cells (Burgess TL, 1999). These factors encourage osteoclasts to be more active than osteoblasts; the result is more bone resorption than deposition. Consequently, bone density in close proximity to the tumor mass is reduced, leading to weakness and ultimately to fractures. OPG, a naturally occurring soluble factor, inhibits these processes, thereby preserving the integrity of bone mass (Hofbauer LC, 2004; Simonet WS, 1997). Researchers found that BM cultures from MM patients overexpressed OPGL mRNA, compared with healthy donors; immunohistochemical staining on BM biopsy specimens also showed an increase of OPGL and reduced OPG expression in MM patients, compared with healthy subjects (Giuliani N, 2001). This Þnding indicates that MCs induce the expression of OPGL and that MM is associated with an imbalance of the OPG/OPGL system in vivo. Therefore, inhibition of RANKL may emerge as a novel therapeutic option for patients with human bone disease (Hofbauer LC, 2004). How currently available bisphosphonates affect these bone resorption and deposition effectors is not clear. Researchers hope that better understanding of the BMM will lead to targeted treatments for MM. Clinical Manifestations. Figure 4 presents an overview of the clinical manifestations of MM. The sources of these manifestations can be classiÞed into the following three groups: •

•

Excess plasma cells. The presence of excess plasma cells disrupts the normal function of BM, causing anemia and bleeding. Additionally, as a result of MM, plasma cells manufacture one particular Ig to the detriment of all others, thereby reducing the ability of the immune system to recognize and destroy invading cells. The result is an increased risk of infection and sepsis. Lytic bone lesions. Changes in resorption weaken the bones and increase the risk of fracture. When lesions are in the spine, compression of the spinal cord can cause loss of autonomic control of bodily functions as well as severe

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FIGURE 4. Clinical manifestations of multiple myeloma.

•

pain. Greater calcium absorption increases the amount of free calcium salts circulating in the blood (hypercalcemia). These salts can be deposited in the kidneys and cause renal failure. Excessive production of M-proteins. The increased level of M-protein in the serum leads to hyperviscosity of the blood, which can cause organ damage, especially to fragile tissues like the eyes. M-proteins can also be deposited in the kidneys (amyloidosis), causing renal failure.

Staging and Prognostic Factors. Since 1975, the Durie-Salmon staging system has been the most widely used staging system for MM. This system classiÞes MM patients into three categories based on MC mass (Table 3). In addition to the Durie-Salmon system, serum beta-2 microglobulin concentration, widely recognized as the most useful prognostic factor—along with factors such as patient age, renal function, and calcium levels—has been used to help tailor treatments to the individual patient (for example, patients with more aggressive disease receive more aggressive treatment). In an attempt to simplify the Durie-Salmon staging system and improve its prognostic value, the International Myeloma Working Group (IMWG) has proposed an International Staging System (ISS; previously called the International Prognostic Index [IPI]) (Greipp PR, 2005). The updated staging system classiÞes patients based on their prognoses and comprises three stages based on easily measured serum levels of albumin and beta-2 microglobulin (Table 4). Serum beta-2 microglobulin (β2M) is present in small amounts in everyone, but in people with MM, its levels increase dramatically. Serum beta-2 microglobulin is

584

MULTIPLE MYELOMA

TABLE 3. Staging of Multiple Myeloma Stage

Criteria

I = Low cell mass

II = Intermediate cell mass III = High cell mass

• All of the following: • Hemoglobin >100 g/L • Serum calcium normal/ <2.6 mmol/L • Bone X ray normal/solitary plasmacytoma lesion only • Low myeloma protein production rate • Fits neither stage I nor stage III • • • •

More than one of the following: Hemoglobin <88 g/L Serum calcium >3.00 mmol/L Advanced lytic bone lesions

Myeloma Cell Mass (cells × 1012 /m2 ) <0.6

0.6–1.2 >1.2

• High myeloma protein production rate Note: Each stage is further subclassified according to renal function: normal = serum creatinine <170 µmol/L; abnormal = serum creatinine ≥170 µmol/L. Source: Adapted from Durie BG, Salmon SE. A clinical staging system for multiple myeloma. Correlation of measured myeloma cell mass with presenting clinical features, response to treatment, and survival. Cancer. 1975;36:842–854.

TABLE 4. International Staging System for Multiple Myeloma Stage I II III

Criteria Serum β2M <3.5 mg/L Serum albumin ≥3.5 g/dL Serum β2M <3.5 mg/L and albumin <3.5 g/dL or Serum β2M ≥3.5 to <5.5 mg/L Serum β2M >5.5 mg/L

β 2M = Beta-2 microglobulin.

Source: Based on data from Greipp PR, et al. International staging system for multiple myeloma. Journal of Clinical Oncology. 2005;23(15):3412–3420.

a reliable marker in evaluating tumor mass, response to treatment, and overall survival (Bataille R, 1984). Numerous studies have demonstrated that patients older than 60 years who have high serum β2M have a poorer prognosis than do patients younger than 60 who have a lower serum β2M (Durie BG, 1990). The use of serum albumin values provides greater consistency in prognostication (Greipp PR, 2005). The ISS appears to discriminate better between staging groups, and it has the advantage of being simple to use. The IMWG hopes that the ISS will enable accurate assessment of MM patients’ prognoses. Researchers noted that

CURRENT THERAPIES

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subclassiÞcation of stages I and II patients according to creatinine levels and subclassiÞcation of high-risk stage III patients according to low platelet or high LDH levels can further improve the discrimination and clinical usefulness of the ISS (Greipp PR, 2005). Etiology The exact cell origin of MM is unknown, but research suggests that a mutation of memory B cells is responsible (Drach J, 2000). Researchers believe that an initial oncogenic event causes MGUS and a later event leads to the development of MM (Salmon S, 1974). These oncogenic events can result from somatic mutations that occur randomly in individual cells throughout the body as a result of carcinogen exposure or errors in DNA replication. The occurrence of MGUS and MM in families indicates the presence of inherited genetic abnormalities (Salmon S, 1974). Only a few risk factors have been strongly implicated in the pathogenesis of MM. Both cohort analysis (Carli PM, 1998; Stagnaro E, 2001) and case-control (Baris D, 2000; Bray I, 2001; Brown LM, 1999; Gautier M, 1994; Hjalgrim H, 1998; Marcelin AG, 1997; Solomon L, 2002) studies have identiÞed the majority of associations between environmental exposures and MM in patients who work in the following industries: agriculture and the petrochemical, paint, rubber, sheet metal, textile, and wood industries (Bray I, 2001; Brown LM, 1999; Gautier M, 1994; Solomon L, 2002; Vineis P, 1996). The incidence of MM is higher in Western industrialized nations. Unlike many other cancers, MM has not been associated with cigarette smoking, alcohol consumption, or exposure to benzene (Pasqualetti P, 1990; Bezabeh S, 1996). MM is more common in African-Americans than in whites and in men than in women (by a ratio of three to two). This distribution may reßect the fact that men are more likely to work in the industries associated with risk factors for MM and that risks are greatest for people in the lowest socioeconomic stratum (Baris D, 2000). More recently, a link has been made between obesity and MM (Calle EE, 2003). Finally, the risk of developing MM increases with age across all populations (Schottenfeld D, 1996). CURRENT THERAPIES Treatment of multiple myeloma (MM) is balanced between periods of active treatment and periods of observation when the disease is in remission. A range of therapies are used to treat MM; the choice of therapy depends on the stage of disease. For most MM patients, the main treatment component is chemotherapy. The chemotherapy is usually accompanied by the corticosteroids prednisone (generics) or dexamethasone (Merck’s Decadron, generics) because these agents demonstrate considerable antitumor activity in MM. Treatment has been revolutionized by the introduction of two independently active compounds: thalidomide (Celgene’s Thalomid, generics) and bortezomib (Millennium Pharmaceuticals/ Johnson & Johnson’s Velcade).

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MULTIPLE MYELOMA

MM is generally considered to be an incurable disease. In the last few years, a handful of research groups have presented data on patients who have been in relapse-free remission for more than ten years. These patients received stemcell transplantations and constitute less than 15% of all patients undergoing such procedures. After exposing the patient to high-dose myeloablative therapy, physicians rescue the patient from the resulting life-threatening toxicities with one or two autologous progenitor stem-cell transplantations (auto-SCTs). A more aggressive approach is allogeneic progenitor stem-cell transplantation (allo-SCT). This method is appropriate only for Þt patients younger than age 55, who have high-risk cytogenetic abnormalities and human leukocyte antigen (HLA)-matched donors. Auto-SCT and allo-SCT are discussed in detail in the “Nonpharmacological Approaches” section. The success of all MM treatments is measured by the level of circulating M-proteins. Complete disappearance of M-proteins in both serum and urine is considered a complete response (CR); stabilization of M-protein levels in serum is deemed a partial response (PR). An objective response (OR) is the sum of CR and PR rates. Table 5 deÞnes the various responses to treatment. The majority of patients achieve only a PR that results in a temporary, stable plateau phase. This plateau phase can be sustained for as long as two years. Regimens that are used to achieve this plateau phase are called initial remission induction therapy. Patients who do not achieve a response—that is, MM progresses while they are still on induction chemotherapy—are described as having primary refractory disease or resistant MM. Secondary refractory patients are those who do respond to induction chemotherapy but do not respond to treatment after relapse. In the past, maintenance therapy was prescribed in an attempt to prolong the period of remission, but this use of conventional chemotherapy agents is controversial: data have shown that there is no survival beneÞt and that continued use of these agents through the remission period can result in drug resistance that makes construction of second- and third-line regimens more difÞcult. Few conventional chemotherapy agents are now used in this setting; instead, immunomodulatory drugs such as interferons (IFNs) and thalidomide are more commonly used despite the few data that support their use. The efÞcacy of the initial remission induction therapy has a bearing on all subsequent treatments after relapse. Clinicians construct subsequent regimens using agents to which the patient has not yet been exposed. These regimens are often combinations of multiple alkylating agents. Purely palliative interventions for MM include bisphosphonates, radiotherapy, and, less commonly, plasmapheresis (the mechanical removal of monoclonal proteins). Some of these approaches are discussed in the following sections. Table 6 lists the agents and procedures currently used in the treatment of MM. Melphalan and Prednisone (MP) Overview. The combination of the alkylating agent melphalan (GlaxoSmithKline/Celgene’s Alkeran, generics) and the corticosteroid prednisone (generics) is

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TABLE 5. Multiple Myeloma: Response-to-Treatment Definitions Response to Treatment

Description

Objective response (OR) Complete response (CR)

Near complete response (nCR)

Very good partial response (VGPR) Partial response (PR) Minimal response (MR) Stable disease (SD)

Progressive disease (PD)

Unable to detect MP in serum and urine using sensitive negative immunofixation electrophoresis. Normal percentage of plasma cells in the bone marrow or absence of MCs by staining techniques. Positive immunofixation electrophoresis result but unable to detect MP in serum and urine by less sensitive methods. 90% or more reduction in MP. 50% or more reduction in MP. Less than 50% reduction in MP. Some reduction in MP. Stable disease parameters, including number and extent of bone lesions. 25% or more increase in MP. Increase in plasma cells in the bone marrow. New or larger bone lesions.

MP = Myeloma protein. MCs = Myeloma cells.

Cl

H2N COOH N

Cl FIGURE 5. Structure of melphalan.

the most commonly prescribed therapy for MM and has been used for 30 years as a Þrst-line therapy. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the overall regimen activity: •

Melphalan (Figure 5) is an alkylating agent and a phenylalanine derivative of mechlorethamine. Alkylation of DNA causes both strand breaks and the formation of cross-links between strands of DNA. Both lesions result in aberrant DNA replication and transcription of RNA. Melphalan, like other alkylating agents, is cell-cycle-phase-nonspeciÞc. However, rapidly proliferating cells, such as MM cells, are more susceptible to the action of alkylating agents because less time is available for DNA enzymes to repair the cytotoxic lesion.

588

MULTIPLE MYELOMA

TABLE 6. Current Regimens and Drug Classes Used for Multiple Myeloma Regimen or Class

Regimen Components Agent

Melphalan, Melphalan prednisone/ (GlaxoSmith prednisolone Kline/Celgene’s (MP) Alkeran, generics)

Availability

Dose

US, F, Melphalan: 8 G, I, S, mg/m2 /d, PO, days 1–4 UK, J

Prednisone (generics)

US, F, G, I, S

Prednisone: 60 mg/m2 /d, PO, days 1–4

Prednisolone (generics)

UK, J

Prednisolone: 60 mg/m2 /d, PO, days 1–4 Cycle repeats every 28 days.

Common Toxicities • Neutropenia • Thrombocytopenia • Infection • Nausea, vomiting • Alopecia

US, F, Vincristine: G, I, S, 0.4 mg/d, IV UK, J continuous infusion, days 1–4

• Neutropenia • Thrombocytopenia • Sepsis

Doxorubicin (Pfizer’s Adriamycin, generics)

US, F, Doxorubicin: G, I, S, 9 mg/m2 /d, IV UK, J continuous infusion, days 1–4

• Cardiotoxicity

Dexamethasone (Merck’s Decadron, generics)

US, F, Dexamethasone: G, I, S, 40 mg, PO, days UK, J 1–4, 9–12, 17–20. Cycle repeats every 28–35 days.

Vincristine, Vincristine doxorubicin, (generics) dexamethasone (VAD)

High-dose dex- Dexamethasone amethasone, (Merck’s single agent Decadron, (HDD) generics)

US, F, Dexamethasone: G, I, S, 40 mg, PO, days UK, J 1–4, 9–12, 17–20 Cycle repeats every 28 days.

• Increased appetite • Altered mood • Impaired glucose tolerance • Proximal myopathy • Cushingoid features • Tremor • Gastric erosions or ulceration with bleeding • Pseudorheumatism

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TABLE 6. (continued) Regimen or Class

Regimen Components Agent

Availability

Dose

High-dose GlaxoSmithKline/ US, F, G, Melphalan: melphalan, Celgene’s I, S, 140–200 mg/m2 , IV single agent Alkeran, generics UK, J

Common Toxicities • • • •

Myelosuppression Nausea, vomiting Mucositis Diarrhea

• Alopecia • Infection Thalidomide, dexamethasone

Thalidomide (Celgene’s Thalomid, generics)

US, F, G, Thalidomide: I, S, 200 mg/day, PO UK, J

• Venous thromboembolism • Constipation • Rash

Dexamethasone (Merck’s Decadron, generics)

US, F, G, Dexamethasone: I, S, 40 mg on days UK, J 1–4, 9–12, and 17–20, PO, of each 28-day cycle Bortezomib, Bortezomib US, F, G, Bortezomib: single agent (Millennium I, S, 1.3 mg/m2 , IV push, on days 1, Pharmaceuticals/ UK 4, 8, and 11 of a Johnson & 21-day cycle for Johnson’s up to eight cycles Velcade) VBCMP

Vincristine (generics) US, F, G, Vincristine: I, S, 1 mg/m2 , IV, day 1 UK, J

Carmustine/ US, F, G, Carmustine: BCNU S, UK 12 mg/m2 , IV, (Bristol-Myers day 1 Squibb’s BiCNU/ Carmubris) CyclophosUS, F, G, Cyclophosphamide: phamide I, S, 250 mg/m2 , IV, day 1 (Bristol-Myers UK, J Squibb’s Cytoxan, Baxter’s Endoxan/ Endoxana, generics)

• Peripheral neuropathy • Thrombocytopenia • Gastrointestinal disorders • Nausea, vomiting • Peripheral neuropathy • Alopecia • Neutropenia • Thrombocytopenia • Infection

590

MULTIPLE MYELOMA

TABLE 6. (continued) Regimen or Class

Regimen Components Agent

Availability

Dose

Melphalan US, F, G, Melphalan: 6 (GlaxoSmithKline/ I, S, mg/m2 , PO, days 1–4 Celgene’s Alkeran, UK, J generics) Prednisone (generics) US, F, G, Prednisone: I, S, 40 mg/m2 , PO, days 1–7; then UK 20 mg/m2 , PO, days 8–14 (cy 1); Repeat every 35 days. VMCP/VBAP Vincristine US, F, G, Vincristine: (cycles (generics) I, S, 1 mg/m2 , IV, day 1 (maximum dose alternate) UK, J 1.5 mg) Melphalan US, F, G, Melphalan: (GlaxoSmithKline/ I, S, 6 mg/m2 /day, Celgene’s Alkeran, PO, days 1–4 UK, J generics) CyclophosUS, F, G, Cyclophosphamide: phamide I, S, 125 mg/m2 /day, PO, days 1–4 (Bristol-Myers UK, J Squibb’s Cytoxan, Baxter’s Endoxan/ Endoxana, generics) Prednisone (generics) US, F, G, Prednisone: I, S, 60 mg/m2 /day, PO, days 1–4 UK Repeat VMCP every three weeks for three cycles, followed by VBAP for three cycles. VBAP (cycles Vincristine US, F, G, Vincristine: alternate (generics) I, S, 1 mg/m2 , IV, day 1 (maximum dose with VMCP UK, J cycles) 1.5 mg)

Common Toxicities

• Myelosuppression • Fatigue • Sleeping difficulties • Aching of hands and feet

• Myelosuppression • Fatigue • Sleeping difficulties • Aching of hands and feet

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TABLE 6. (continued) Regimen or Class

VAMP and CVAMP

Regimen Components Agent

Availability

Carmustine/ US, F, G, BCNU S, UK (Bristol-Myers Squibb’s BiCNU/ Carmubris) Doxorubicin (Pfizer’s US, F, G, Adriamycin, I, S, generics) UK, J Prednisone US, F, G, (generics) I, S, UK Vincristine (generics) US, F, G, I, S, UK, J

Dose

Common Toxicities

Carmustine: 30 mg/m2 , IV, day 1

Doxorubicin: 30 mg/m2 , IV, day 1 Prednisone: 60 mg/m2 , PO, days 1–4 Vincristine: 0.4 mg/m2 , IV, by continuous infusion over 24 hours for four days

Doxorubicin (Pfizer’s US, F, G, Doxorubicin: Adriamycin, I, S, 9 mg/m2 , IV, by continuous generics) UK, J infusion over 24 hours for four days MethylpredUS, F, G, Methylprednisolnisolone (Pfizer’s I, UK, one: 1.5 g, IV, on Medrol/ J day one and Medrone/Medrate, tapered down to generics) zero (+/− cyclophosphamide) CyclophosUS, F, G, Cyclophosphamide: phamide I, S, 500 mg, IV, on (Bristol-Myers UK, J days 1, 8, and 15 Squibb’s Cytoxan, Administered Baxter’s Endoxan/ every four weeks. Endoxana, Weekly IV generics) cyclophosphamide can be administered between courses of VAMP to increase intensity.

• Cardiotoxicity

• Nausea, vomiting • Fatigue • Alopecia • Diarrhea • Irritation • Peripheral neuropathy • Cardiotoxicity

592

MULTIPLE MYELOMA

TABLE 6. (continued) Regimen or Class

Regimen Components Agent

Availability

Dose

Interferon-alpha IFN-α-2a US, F, G, IFN-α-2a (Roche/ChugI, S, (Roferon-A): ai’s Roferon-A, UK, J 3 × 106 –6 × 106 IU Takeda’s Canferon A)

Bisphosphonates

IFN-α-2b US, F, G, IFN-α-2b (ScheringI, S, (Intron A): 3 × Plough’s Intron UK, J 106 –6 × 106 IU A/Introna/ Viraferon) Zoledronic acid US, F, G, Zoledronic acid: (Novartis’ I, S, 4 mg, IV, every Zometa) UK, J four weeks

Common Toxicities • Transient fever and flu-like symptoms • Central nervous system toxicity • Fatigue • Myalgias • Nausea • Alopecia • Weight loss

• Gastrointestinal toxicity • Renal impairment • Fever • Anemia • Hypocalcemia • • • •

Hypophosphatemia Headache Flu-like symptoms Bone pain

• • • •

Fatigue Rigors Arthralgia Myalgia

• Injection site reaction • Fever • Blood creatinine and blood urea increased Pamidronate (Novartis’ Aredia, generics)

US, F, G, Pamidronate: I, S, 90 mg, IV, every four weeks UK, J

• Thrombocytopenia • Arthralgia • Myalgia • • • •

Insomnia Anemia Transient bone pain Fever

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TABLE 6. (continued) Regimen or Class

Regimen Components Agent

Availability

Dose

Common Toxicities • Myalgia • Flu-like symptoms • Injection site reaction • Headache • Gastrointestinal toxicity • Hypocalcemia • Hypophosphatemia • Hypomagnesemia • Increase in serum creatinine

Clodronate (Schering’s Bonefos/ Clastoban, generics)

F, G, I, S, Clodronate: UK 1,600–3,600 mg/day, PO, for 2–3 years

• Gastrointestinal toxicity • Hypocalcemia • Transient increase of serum creatinine • Increase of parathyroid hormone level

IV = Intravenous; PO = By mouth. US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan.

•

Prednisone (Figure 6) is a glucocorticosteroid, and prednisolone is the major active metabolite of prednisone. Glucocorticosteroids reduce the inßammatory response to tumor tissue by preventing white blood cells from functioning to produce swelling and pain around the tumor site. Corticosteroids induce lysis of peripheral lymphocytes and slow lymphocyte production. Addition of corticosteroids to chemotherapy regimens may augment their efÞcacy.

Clinical Performance. The MP regimen was a breakthrough in MM treatment when it was introduced more than 30 years ago; the protocol has remained largely unaltered since then. In their landmark study, Alexanian and colleagues randomized 183 patients not previously treated with an alkylating agent to receive melphalan plus prednisone or melphalan alone (Alexanian R, 1969). They compared two MP and two melphalan regimens. Seventy-nine patients received treatment with 0.025 mg/kg/day melphalan for four days every six weeks and either alternate-day prednisone (28 patients received 1.0 mg/kg every Monday, Wednesday, and Friday morning) or concurrent prednisone (51 patients received 2.0 mg/ for four days every six

594

MULTIPLE MYELOMA

CH2OH C O

H3C

O OH

H3C

O FIGURE 6. Structure of prednisone.

weeks). One hundred and four patients received treatment with 0.025 mg/kg/day melphalan alone: 35 patients were treated every day; the other 69 received melphalan for four days every six weeks. In evaluable patients, investigators observed a PR rate of 61% in the melphalan plus alternate-day prednisone group and 65% in the melphalan plus concurrent prednisone group. The PR rates for the daily melphalan and intermittent melphalan patients were 17% and 32%, respectively. Patients treated with MP had a median survival of 24 months, compared with only 18 months for the melphalan patients. Patient response to MP is gradual; maximum beneÞt is usually achieved only after several months. It is rare for MP to induce complete remission; instead, patients reach a plateau phase. This phase is characterized by stabilization of serum myeloma protein levels, which permits discontinuation of treatment. MP has two signiÞcant disadvantages. Melphalan and other alkylating agents should not be given to patients who are expected to undergo a transplant because these agents reduce the number of stem cells that can be harvested. In addition, long-term use of MP increases the risk of patients developing treatment-related acute myeloid leukemia (Cuzick J, 1987). Vincristine, Doxorubicin, and Dexamethasone (VAD) Overview. The VAD chemotherapy regimen is made up of vincristine (generics), doxorubicin (PÞzer’s Adriamycin, generics), and dexamethasone (Merck’s Decadron, generics). The relative success of VAD, coupled with its rapid onset of action, has prompted use of this regimen as Þrst-line therapy to reduce the tumor burden of myeloma cells (MCs) before stem-cell transplantation. VAD is ideal for this purpose because, unlike other combination regimens, it does not damage stem cells. VAD is usually given for four courses to reduce the tumor burden and the number of MCs in the peripheral circulation. VAD may also be used as a Þrst-line therapy in patients with renal failure and for relapsed or refractory MM. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the overall regimen activity: •

Vincristine (Figure 7) is a vinca alkaloid. Vinca alkaloids bind with microtubular proteins of the mitotic spindle, thus leading to mitotic arrest or cell

CURRENT THERAPIES

N

OH CH3

NH H3CO H3CO

595

N

O

H N R

H HO

CH3 OAc O

H3CO

FIGURE 7. Structure of vincristine (R = CHO).

FIGURE 8. Structure of doxorubicin (R = OCH3 , R1 = OH, R2 = H, R3 = H, R4 = OH).

•

•

death. The drug achieves its selective toxicity against cancer cells by acting on proliferating cells. Doxorubicin (Figure 8) is an anthracycline. Anthracyclines interact with several different cellular targets, most importantly topoisomerase II. By inhibiting this DNA religation enzyme, anthracyclines exert their cytotoxic effect. Another mechanism leading to cell death, known as DNA intercalation, involves insertion of the anthracycline molecule between base pairs. This phenomenon causes single- and double-stranded breaks in DNA that inhibit cell proliferation. The free radicals generated by the reductive metabolism of anthracyclines may also damage cellular structures. Dexamethasone (Figure 9) is a glucocorticosteroid. Corticosteroids reduce the inßammatory response to tumor tissue by preventing activation of signaling molecules such as tumor necrosis factor-alpha (TNF-α). Corticosteroids induce lysis of malignant plasma cells and slow MC production. Addition of corticosteroids to chemotherapy regimens may augment their efÞcacy.

596

MULTIPLE MYELOMA

OR O HO

OH

F O

FIGURE 9. Structure of dexamethasone (R = H).

Clinical Performance. VAD is associated with a high overall response rate in the Þrst-line setting. In an early study of 60 MM patients, the 28-day VAD regimen consisted of 0.4 mg/day vincristine on days 1–4, 9 mg/m2 /day doxorubicin on days 1–4, and 40 mg/day dexamethasone on days 1–4, 9–12, and 17–21 (odd cycles), or on days 1–4 only (even cycles) (Alexanian R, 1990). There was a 28-day interval between cycles. Treatment was continued until the maximum response was observed. In this study, Þrst-line treatment with VAD resulted in an overall response rate of 55%; the CR was 12%. The response was rapid, with a median tumorhalving time of 0.4 months; most patients achieved maximum response after only two cycles. The mean remission duration was 22 months; median survival was 36 months. Because of the rapid response time, VAD is the ideal regimen for newly diagnosed patients who require rapid control of MM. Unfortunately, VAD does not produce any signiÞcant increase in overall survival when compared with other combination chemotherapy regimens (DeVita V, 2001). VAD is also an important regimen in relapsing disease. In a key trial of 36 patients with relapsing MM, VAD was administered to 17 patients as in the 60-patient study previously described; the other 19 patients received 40 mg/day dexamethasone on days 1–4, 9–12, and 17–21 (Alexanian R, 1986). VAD induced a PR or CR in 65% of patients; dexamethasone alone induced remission in only 21% of patients. The mean remission duration was 9 months; median survival was 22 months for both treatment groups. MM patients with renal failure are also good candidates for the VAD protocol. Because the chemotherapy agents of VAD are not renally excreted, the regimen is safe and effective, and there is no need to modify the dose (Aitchison RG, 1990). Yet another group of patients for whom VAD is well suited are those who are resistant to standard melphalan-containing therapy; VAD has elicited a PR in 25% of these patients (Barlogie B, 1984). The VAD regimen is associated with signiÞcantly more adverse events than dexamethasone monotherapy (described later). When 177 patients treated with VAD were compared with 112 patients treated with dexamethasone only, 27% of patients treated with VAD experienced serious adverse events, compared with 4% of patients treated with dexamethasone only (Alexanian R, 1992). The most common adverse events associated with VAD that required hospitalization were neutropenia, catheter-related complications such as venous thromboembolism

CURRENT THERAPIES

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(VTE), and infection. Virtually all VAD patients had mild peripheral neuropathy, alopecia, and mild cushingoid features. Most patients treated with dexamethasone monotherapy experienced cushingoid features and/or insomnia but not myelosuppression. The VAD regimen requires a 96-hour infusion of doxorubicin through a central venous catheter, a method that requires hospitalization and may lead to catheterrelated infections or VTE (Alexanian R, 1990). Doxorubicin is also associated with cardiotoxicity (Rinehart JJ, 1974). For patients whose disease becomes refractory to VAD, the addition of cyclophosphamide (CVAD) can elicit responses in up to 40% of patients (DeVita V, 2001). High-Dose Dexamethasone, Single Agent Overview. High doses of the corticosteroid dexamethasone are an important component of many chemotherapy regimens, but they can also be used as Þrst-line monotherapy. High-dose dexamethasone monotherapy is an effective alternative to complex combination therapy. Dexamethasone’s efÞcacy has been proved by comparing the efÞcacy of single-agent treatment to that of combination regimens. Because dexamethasone is not as toxic as combination chemotherapy, highdose dexamethasone is used in patients who are contraindicated for chemotherapy. High-dose dexamethasone is also an initial treatment for patients with renal failure because it is not dependent on the kidneys for excretion. Dexamethasone is administered alone as intermittent oral therapy, thereby providing a convenient, potent alternative to the more complex combination regimens. Mechanism of Action. Dexamethasone (Figure 9) is a glucocorticosteroid. Corticosteroids reduce the inßammatory response to tumor tissue by preventing activation of signaling molecules such as tumor necrosis factor-alpha (TNF-α). Corticosteroids induce lysis of malignant plasma cells and slow MC production. Addition of corticosteroids to chemotherapy regimens may augment their efÞcacy. Clinical Performance. High-dose dexamethasone has been shown to be almost as effective as VAD chemotherapy in patients with refractory MM (Alexanian R, 1986). Fifty-two MM patients who had previously been unresponsive to chemotherapy were treated with either the VAD regimen previously described or 40 mg/day dexamethasone on days 1–4, 9–12, and 17–20. Twenty-four patients had been resistant to chemotherapy for more than 12 months. VAD induced a PR or CR in 32% of patients, and dexamethasone alone induced remission in 27% of patients. The mean remission duration was 9 months; median survival for both treatment groups was 22 months. Side effects of high-dose dexamethasone include increased appetite, altered mood, impaired glucose tolerance, proximal myopathy, cushingoid features, tremor, gastric erosions or ulceration with bleeding, and pseudorheumatism. Most of these side effects are reversible when therapy is discontinued.

598

MULTIPLE MYELOMA

High-Dose Melphalan, Single Agent Overview. In an attempt to improve patient survival, clinicians have used moreintense treatments. The use of high-dose intravenous (IV) melphalan as part of a complex sequence of regimens is now the preferred treatment option in eligible patients. Patients receive induction therapy (usually VAD) for four cycles. A stem-cell harvest is performed. The patient has a single infusion of high-dose melphalan, which kills the majority of bone marrow cells and MCs. After highdose melphalan, the patient is rescued by reinfusing the autologous progenitor stem cells (described further on). Several studies have demonstrated that highdose melphalan is effective for all stages of MM. Mechanism of Action. Melphalan (Figure 5) is an alkylating agent and a phenylalanine derivative of mechlorethamine. Alkylation of DNA causes both strand breaks and the formation of cross-links between strands of DNA. Both of these lesions result in aberrant DNA replication and transcription of RNA. Melphalan, like other alkylating agents, is cell-cycle-phase-nonspeciÞc. However, rapidly proliferating cells, such as MM cells, are more susceptible to the action of alkylating agents because less time is available for DNA enzymes to repair the cytotoxic lesion. Clinical Performance. McElwain and Powles were the Þrst to demonstrate that a single high dose of melphalan could induce high response rates even in refractory MM (McElwain TJ, 1983). They treated eight MM patients, four of whom were previously untreated, with 100-140 mg/m2 melphalan. All patients responded to treatment. The same investigators continued their work in 58 MM patients younger than 63 years (Selby PJ, 1987). Patients were treated with 140 mg/m2 melphalan. Forty-one patients had not received previous treatment for MM; of these patients, 11 achieved CR and another 21 achieved PR. The median duration of remission was 19 months. Another 15 patients with refractory MM were treated; 10 responded to therapy, including 2 patients who experienced CR. However, all patients in this group relapsed within 12 months. Long-term follow-up of patients treated with 140 mg/m2 melphalan showed that the therapy was associated with high response rates, relatively long remissions, and good symptom control (Cunningham D, 1994). Of 63 previously untreated MM patients, 51 responded and 20 achieved CR. The median duration of response was 18 months. After a median follow-up of 74 months, 23 patients were still alive; median survival duration was 47 months, and 35% of patients were expected to be alive at nine years. High-dose melphalan is associated with serious adverse events, including severe myelosuppression, moderate nausea, vomiting, mucositis, diarrhea, and alopecia (Selby PJ, 1987). Myelosuppression may lead to death from an opportunistic infection. In the study by Selby and colleagues, 12% of patients treated with 140 mg/m2 melphalan died within two months of therapy due to infection (Selby PJ, 1987). The profound myelosuppression induced by high-dose melphalan led to the use of auto-SCT to rescue myeloproliferation.

CURRENT THERAPIES

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The combination of high-dose melphalan and auto-SCT has become standard therapy for all patients younger than age 65 with good performance status (PS) (Attal M, 1996). Current high-dose melphalan therapy consists of either 140 mg/m2 dose of melphalan (MEL140) combined with 8 Gy total body irradiation (TBI) or a 200 mg/m2 dose of melphalan (MEL200). The latter regimen has been proved more effective and less toxic than MEL140-TBI, but both regimens are still widely used (Moreau P, 2002). The advantage of MEL200 over MEL140-TBI is its lower transplant-related mortality (TRM) rate and better overall survival. The survival beneÞt was not due to a better or longer response to transplant but to better survival after relapse. Although single-agent melphalan is usually followed by auto-SCT, investigations have explored the beneÞts of the drug alone in patient groups not normally eligible for transplants. Studies have shown that intermediate-dose melphalan, 100 mg/m2 (MEL100), is superior to standard MP in elderly patients. Median event-free survival was 34 months with MEL-100 and 17.7 months with standard MP (Palumbo A, 1999). A more recent trial conducted by the same research group demonstrated that tandem cycles of MEL100 elicited CR in 25% of patients aged 50-70, compared with a 6% CR following standard MP (Palumbo A, 2004[b]). These trials support the use of intermediate-dose melphalan as an effective treatment option in elderly patients. In fact, use of this treatment in the elderly is becoming more widespread. Thalidomide and Dexamethasone Overview. Thalidomide (Celgene’s Thalomid, generics), an orally bioavailable immunomodulatory/antiangiogenic agent, was launched in Europe in the 1960s as a sedative/hypnotic. Thalidomide was very effective in treating the nausea associated with pregnancy, but it proved to be teratogenic (causing birth defects) and was removed from the market. However, thalidomide enjoyed a renaissance when it was discovered that the agent was a potent inhibitor of angiogenesis (D’Amato RJ, 1994). Because MM is associated with increased angiogenesis, investigators were inspired to try thalidomide therapy in cases of relapsed MM. They observed a 32% response rate, and two patients experienced CR. These dramatic results sent thalidomide to the top of the list of investigational drugs for MM treatment. Thalidomide is not currently approved for the treatment of MM in the United States, France, Germany, Italy, Spain, the United Kingdom, or Japan. Instead, the agent is used off-label. In 1998, the FDA approved thalidomide for the treatment of erythema nodosum leprosum (ENL), an inßammatory condition associated with leprosy, but its distribution is restricted. In the United States, Celgene is the sole provider, despite the fact that the patent protecting thalidomide has expired. Generics competition is generally limited by the costs associated with operating restrictive access schemes established to maximize safety. Under Celgene’s System for Thalidomide Education and Prescribing Safety (STEPS), each patient who is prescribed thalidomide must register with Celgene by signing a declaration that conÞrms his or her understanding of the risks associated with thalidomide therapy.

600

MULTIPLE MYELOMA

The FDA has granted thalidomide orphan drug status for the treatment of MM, and in October 2004, Celgene’s Thalomide received an approvable letter from the FDA for the treatment of MM. The FDA requires Celgene to submit data from the E1A00 trial, a large, randomized Eastern Cooperative Oncology Group (ECOG) study that compared thalidomide plus dexamethasone with dexamethasone alone in previously untreated MM patients. This trial has been completed and is discussed later here. In Europe and Japan, thalidomide is distributed for off-label use only by a small number of generics companies via restricted access schemes. However, other companies are trying to gain approval for thalidomide in the treatment of MM in these territories. In Europe, Pharmion purchased the right to market thalidomide from Celgene, and in November 2001, the European Medicines Agency (EMEA) granted the drug orphan drug status for the treatment of MM. In July 2002, while seeking regulatory approval for thalidomide, Pharmion began selling thalidomide in Europe on a compassionate-use or named-patient basis; the company established a strict patient registration program, similar to STEPS, called the Pharmion Risk Management Program. In May 2004, Pharmion withdrew its MM application from the EMEA so as to prepare a submission with data from two additional clinical trials. The Þrst study will compare survival and additional clinical end points for two doses of thalidomide in 400 patients with relapsed, refractory MM. The second study’s endpoints will include time to progression and survival in 436 newly diagnosed patients treated with thalidomide plus dexamethasone or with dexamethasone alone. In January 2005, Japan’s Ministry of Health, Labor, and Welfare (MHLW) granted thalidomide, which is under development by Fujimoto Pharmaceutical, orphan drug status for the treatment of MM. Initial interest in thalidomide focused on its use as a single agent. A Phase II study involving 84 patients with treatment-refractory MM demonstrated promising results with thalidomide and provided the rationale for further trials. Of these patients, 90% had received high-dose melphalan and auto-SCT, and 69% had received two or more cycles of intensive chemotherapy. Patients received 200 mg thalidomide daily as a single agent for a mean duration of 80 days. The dose was increased by 200 mg every two weeks until the maximum dose of 800 mg per day was reached. Two patients achieved a complete remission; eight achieved at least a 90% reduction in serum and urine M-proteins; six achieved at least a 75% reduction in M-proteins; seven achieved at least a 50% reduction; and six achieved at least a 25% reduction. The overall rate of response (deÞned as a greater than 25% reduction in M-proteins) was 32% (Singhal S, 1999). Side effects occurred at all dose levels and increased with escalating doses. Most adverse effects were mild to moderate. One-third or more of patients experienced constipation, weakness or fatigue, and somnolence. Nine patients could not tolerate thalidomide and discontinued treatment after a median of 36 days. One patient died suddenly despite responding to treatment; the cause of death, possibly thalidomide, is unknown. The median event-free survival for all 84

CURRENT THERAPIES

601

O N O

O

NH

O

FIGURE 10. Structure of thalidomide.

patients was three months. After 12 months, only 22% of the patients remained event-free (Singhal S, 1999). More recent studies have supported the use of thalidomide plus dexamethasone instead of VAD as part of an induction regimen. This section focuses on the combination of thalidomide and dexamethasone rather than thalidomide monotherapy. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the overall regimen activity: •

•

Thalidomide (Figure 10) is a glutamic acid derivative. Its mechanism of action in MM has not been fully elucidated. In preclinical trials, the agent exhibited both immunomodulatory and antiangiogenic effects. Investigators have shown that thalidomide inhibits the production of TNF-α and increases the effect of anti-TNF-α proteins. Thalidomide stimulates the proliferation of cytotoxic T cells able to destroy tumor cells. In addition, the compound increases secretions of cytokines such as IFN-gamma and IL-2; both exhibit an antitumor effect. Thalidomide also modulates the expression of adhesion molecules on the surfaces of myeloma and stromal cells, a process that is important in tumor survival. Thalidomide exerts potent antiangiogenic activity by blocking two growth factors that control angiogenesis—VEGF and bFGF. Dexamethasone (Figure 9) is a glucocorticosteroid. Corticosteroids reduce the inßammatory response to tumor tissue by preventing white blood cells from functioning to produce swelling and pain around the tumor site. Corticosteroids induce lysis of peripheral lymphocytes and slow lymphocyte production. The addition of corticosteroids to chemotherapy regimens may augment their efÞcacy.

Clinical Performance. Data from the Phase III ECOG E1A00 trial were presented at the American Society of Hematology annual meeting in December 2004. The study compared dexamethasone plus thalidomide with dexamethasone alone as Þrst-line induction therapy in 207 patients with newly diagnosed, advanced MM. Patients received 200 mg/day of thalidomide and/or 40 mg of dexamethasone administered on days 1–4, 9–12, and 17–20 of each 28-day cycle. The primary endpoint was best response at four months; after that point, patients could undergo auto-SCT or continue with drug therapy. In this trial, response was deÞned as a reduction in serum M-protein of at least 50%. After four months, there was a signiÞcant difference between the treatment groups: 63% of patients receiving thalidomide plus dexamethasone had responded, compared with 41% of patients who received only dexamethasone.

602

MULTIPLE MYELOMA

Although 67% of patients in the thalidomide plus dexamethasone group experienced serious adverse events, compared with only 42% in the dexamethasone group, there were fewer deaths in the thalidomide plus dexamethasone group (7%) than in the dexamethasone group (11%). The most common serious adverse event was VTE, which affected 18% of patients receiving thalidomide plus dexamethasone but only 3% of patients receiving dexamethasone monotherapy (Rajkumar SV, 2004[b]). Because thalidomide treatment for MM is essentially palliative rather than curative—treatment aims to shrink malignant lesions and provide symptomatic relief—the impact of such treatment on quality of life is particularly important. Given thalidomide’s potential side effects, it may not have a beneÞcial impact on a patient’s quality of life. Preliminary data presented at the American Society of Clinical Oncology (ASCO) meeting in 2003 suggest that 81 mg aspirin (Bayer’s Bayer aspirin, generics) may be sufÞcient to prevent VTE (Tso E, 2003). Bortezomib, Single Agent Overview. Bortezomib (Millennium Pharmaceuticals/Johnson & Johnson’s Velcade) is the Þrst proteasome inhibitor to be approved for the treatment of MM. Although only Phase II studies had been completed, both the European and U.S. regulatory authorities recognized bortezomib as an important new therapy for MM and reviewed the drug under special procedures. In January 2003, Millennium submitted a new drug application (NDA) to the FDA for accelerated review, and the FDA granted bortezomib orphan drug status. In May 2003, the FDA approved bortezomib for relapsed and refractory MM patients who have received at least two prior therapies and demonstrated disease progression during the last therapy; bortezomib was launched within a week. In September 2004, Millennium submitted a supplemental NDA to the FDA for bortezomib as a second-line treatment for MM patients who have received at least one previous therapy; this indication was approved in March 2005. In January 2003, Millennium also submitted a marketing authorization application to the EMEA. At that time, no procedure existed for accelerated review of applications, but under regulatory requirements for anticancer products in humans, the agency was able to accept an application without Phase III comparator trials. In April 2004, the European Commission granted European Union (EU) marketing authorization under exceptional circumstances; the Þrst European launch occurred in July 2004 in the United Kingdom. In March 2005, the EMEA approved bortezomib as a monotherapy for the treatment of progressive MM in patients who have received at least one previous therapy and have already undergone, or are unsuitable for, stem-cell transplantation. In December 2003, the MHLW granted bortezomib orphan drug status in Japan. In January 2005, Millennium and Johnson & Johnson announced the initiation of the Phase III Velcade as Initial Standard Therapy in Multiple Myeloma: Assessment with Melphalan and Prednisone (VISTA) study. VISTA will compare bortezomib combined with MP versus MP alone in patients with newly diagnosed MM who are not eligible for a stem-cell transplant. The Evaluation of Velcade Employed as

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Retreatment for EfÞcacy, Safety, and Tolerability (EVEREST) trial was announced in March 2005. This trial will assess bortezomib in MM patients who previously responded to bortezomib and relapsed following a treatment-free remission. Mechanism of Action. Bortezomib is a small-molecule proteasome inhibitor that works through multiple pathways, including those that inßuence apoptosis and angiogenesis. Proteasomes are enzyme complexes responsible for breaking down proteins, including those that regulate cell division, notably NF-κB. MM is dependent on NF-κB and NF-κB-dependent genes as growth factors and for adhesion of plasma cells to the bone marrow stroma. Bortezomib has high selectivity for the proteasome over other proteases (e.g., thrombin) and has demonstrated in vitro cytotoxicity against a wide range of tumor cell lines. In MM, the drug inhibits malignant plasma cells and stromal cells in the bone marrow. Studies in MM cell lines have demonstrated growth inhibition in doxorubicin-resistant, melphalan-resistant, and mitoxantrone-resistant cells. The addition of dexamethasone to bortezomib provides additional dose-dependent activity. Clinical Performance. Applications for approval were based on the results of the Phase II Study of Uncontrolled Multiple Myeloma Managed with Proteasome Inhibition Therapy (SUMMIT) (Richardson PG, 2003[b]). In this trial, 202 patients with advanced, relapsed/refractory MM received 1.3 mg/m2 bortezomib by IV push on days 1, 4, 8, and 11 of a 21-day cycle for up to eight cycles. The addition of 20 mg dexamethasone was permitted in patients with progressive disease after two cycles and in patients with stable disease after four cycles. Of the 193 evaluable patients, 173 had previously been treated with three or more standard MM therapies; the other 20 patients had received either two standard therapies or a stem-cell transplant. Sixty-seven percent of patients achieved a response (deÞned as stable disease or better); in this group, the CR rate was 10%. The overall response rate (reduction of M-proteins by 50%) was 35%. The cohort of patients who received additional dexamethasone achieved a slightly better overall response rate of 39%; of these patients, 17% achieved a CR. Median survival for all patients was 16 months; the median time to progression was seven months. In patients who responded, the duration of response obtained with bortezomib was 12 months. Investigators described therapy with dexamethasone and bortezomib as well tolerated. Low-grade gastrointestinal toxicities and low-grade thrombocytopenia occurred in 40% of patients. Thirty-one percent experienced grades I-III peripheral neuropathy, but 80% of all patients in this group entered the trial with peripheral neuropathy as a result of prior therapy; of more concern was the fact that 38% of these patients developed peripheral neuropathy as a result of bortezomib therapy with or without dexamethasone (Richardson PG, 2003[b]). The smaller Clinical Response and EfÞcacy Study of Bortezomib in the Treatment of Relapsing Multiple Myeloma (CREST) also supported the regulatory submission in both Europe and the United States (Jagannath S, 2002). The Phase II trial involved 54 patients with earlier-stage disease—relapsed or refractory MM after Þrst-line therapy. Patients were randomized to receive one of two

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doses: 1 mg/m2 or 1.3 mg/m2 of bortezomib by IV push following the regimen used in the SUMMIT trial. Again, the addition of 20 mg dexamethasone was permitted in patients with progressive disease after two cycles and in patients with stable disease after four cycles. In the higher-dose arm, the overall response rate was 50%; 19% of responders achieved stable disease. In the lower-dose arm, the rates were 33% and 26%, respectively. Median time to disease progression was 11 months in both arms; median overall survival had not been reached at 8.8 months follow-up. The addition of dexamethasone in patients who failed to respond after two cycles of treatment increased the overall response rates to 62% in the higher-dose arm and 44% in the lower-dose arm. The principal adverse events were gastrointestinal, including constipation, diarrhea, nausea, and vomiting. Peripheral neuropathy occurred in 18% of patients in the lower-dose arm versus 58% in the higher-dose arm. The special procedures under which bortezomib was reviewed in Europe and the United States require that data from Phase III trials be submitted to the regulatory authorities after approval. The Phase III Assessment of Proteasome Inhibition for Extending Remissions (APEX) trial involved 669 patients with relapsing or refractory MM who had received one to three previous therapies. APEX was terminated early because of favorable results for bortezomib therapy. Interim results were announced at the 2004 American Society of Hematology meeting (Richardson PG, 2004[a]). The primary endpoint of APEX was time to disease progression; secondary end points included survival, response rate, response duration, time to skeletal events, incidence of severe infection, and safety. Patients were randomized to receive either bortezomib or high-dose dexamethasone. Bortezomib was administered at 1.3 mg/m2 by IV push on days 1, 4, 8, and 11 of a 21-day cycle for eight cycles (induction), followed by 1.3 mg/m2 IV push on days 1, 8, 15, and 22 of a 35day cycle for three cycles (maintenance). Patients in the comparator arm received 40 mg dexamethasone orally on days 1–4, 9–12, and 17–20 of a 35-day cycle for four cycles (induction), followed by 40 mg dexamethasone orally on days 1–4 of a 28-day cycle for Þve cycles (maintenance). Thus, patients in the bortezomib and dexamethasone groups received 39 and 40 weeks of treatment, respectively. The mean time to response was 43 days in both treatment arms. Thirty-eight percent of patients receiving bortezomib responded, compared with 18% of dexamethasone patients. Furthermore, the CR and near CR rates for the bortezomib group were 6% and 7%, respectively, compared with only 1% and 1% in the comparator group. Compared with dexamethasone treatment, time to disease progression increased by 78% and risk of death fell by 41% in patients receiving bortezomib. After one year, 80% of the bortezomib group and only 66% of the dexamethasone group were still alive. The incidence of adverse events was similar in the two groups. The incidence of severe adverse events was 44% in the bortezomib arm and 43% in the dexamethasone arm. The most common adverse events related to bortezomib therapy were thrombocytopenia and neutropenia. In May 2005, preliminary data were presented at the ASCO meeting on the use of bortezomib and dexamethasone as an induction regimen before auto-SCT

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in previously untreated MM patients (Harousseau J, 2005). Patients received four consecutive 21-day cycles. Bortezomib was administered as a 1.3 mg/m2 IV push on days 1, 4, 8, and 11 and 40 mg dexamethasone was given orally on days 1–4 and 9–12 of the Þrst two cycles and on days 1–4 for the latter two cycles. Stem-cell harvest was performed just before the fourth cycle. In 48 evaluable patients, investigators observed a pretransplant OR rate of 67%, including 21% who achieved CR and 10% who had a very good partial response. The patients who underwent auto-SCT had an OR rate of 90%, including CR and very good PR rates of 33% and 21%, respectively. At this time, no data are available on the duration of response. Side effects were mild and most commonly gastrointestinal toxicities (49%), fatigue (34%), peripheral neuropathy (29%), skin reactions (26%), and thrombocytopenia (17%). Other Alkylator-Containing Combination Chemotherapy Regimens Overview. Many other chemotherapy regimens are based on alkylating agents collectively referred to as combination chemotherapy (CCT ). Each of these regimens includes four to six of the following compounds (see Table 6 for examples): • • • • • •

Vincristine (V) (generics). Cyclophosphamide (C) (Bristol-Myers Squibb’s Cytoxan, Baxter’s Endoxan/Endoxana, generics). Carmustine/BCNU (B) (Bristol-Myers Squibb’s BiCNU/Carmubris). Melphalan (M) (GlaxoSmithKline/Celgene’s Alkeran). Doxorubicin (A) (generics). Prednisone (P) (generics)/methylprednisolone (M) (PÞzer’s Medrol/Medrone/Medrate, generics).

Typical regimens are VBCMP and VMCP/VBAP; each has been compared with the MP regimen in clinical trials in the hope of establishing the optimal combination (Blad´e J, 1993; Oken MM, 1997). Numerous randomized trials have compared various combinations of alkylatorbased regimens with MP; some showed improved survival (Durie BG, 1986; Delvyn C, 1977; Oken MM, 1997; Blad´e J, 1993) and others showed equivalence (Gregory WM, 1992; Pavlovsky S, 1988; Peest D, 1988). Researchers conducted a meta-analysis of 6,633 patients in 27 randomized trials that compared combination chemotherapy with MP; they found no improvement in overall survival beneÞt effected by any combination chemotherapy compared with MP (Myeloma Trialists’ Collaborative Group, 1998). The toxicity problems associated with alkylator-based combination chemotherapy are similar to those associated with MP. Alkylating agents cause toxicity to stem cells in varying amounts. Also, the more agents used in the combination, the more toxic the therapy is. Toxicities include myelosuppression, vomiting, alopecia, and infection, as well as cardiotoxicity, which is associated with doxorubicin. These morbidities occur without improvement in overall survival. Because of their failure to consistently demonstrate an improvement over MP, a convenient oral regimen familiar to most clinicians, these combination regimens are generally used as second- and third-line treatments. The combination

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regimens in Table 6 are not an exhaustive list. The table does not include all regimens prescribed because there is little difference between the many alternatives. The following paragraphs discuss particular advantages that can be achieved with particular regimens. Mechanism of Action. The individual agents contribute the following mechanisms to achieve overall regimen activity: •

•

•

•

•

Vincristine is a vinca alkaloid. Vinca alkaloids bind with microtubular proteins of the mitotic spindle, thus leading to mitotic arrest or cell death. The drug achieves its selective toxicity against cancer cells by acting on proliferating cells. Cyclophosphamide (Figure 11) is an alkylating agent. These agents alkylate DNA bases, thereby producing cross-links that covalently link the two DNA strands and prevent cell replication. Carmustine (Figure 12) (BCNU) is a nitrosourea-alkylating agent. Metabolites of carmustine cause alkylation and cross-linking of DNA, which prevent DNA replication and cell division. Other biological effects include inhibition of DNA repair. Generally, nitrosoureas are not cross-resistant with other alkylating agents. Melphalan (Figure 5) is an alkylating agent and a phenylalanine derivative of mechlorethamine. Alkylation of DNA causes both strand breaks and the formation of cross-links between strands of DNA. Both of these lesions result in aberrant DNA replication and transcription of RNA. Melphalan, like other alkylating agents, is cell-cycle-phase-nonspeciÞc. However, rapidly proliferating cells are more susceptible to the action of alkylating agents because of the reduced time available for DNA enzymes to repair the cytotoxic lesion. Doxorubicin (Figure 8) is an anthracycline. Anthracyclines interact with several different cellular targets, most importantly topoisomerase II. By inhibiting this DNA religation enzyme, anthracyclines exert their cytotoxic effect. Another mechanism leading to cell death, known as DNA intercalation, involves insertion of the anthracycline molecule between base pairs. This phenomenon causes single- and double-stranded breaks in DNA that inhibit cell proliferation. The free radicals generated by the reductive metabolism of anthracyclines may also damage cellular structures.

FIGURE 11. Structure of cyclophosphamide.

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FIGURE 12. Structure of carmustine.

•

Prednisone (Figure 6) and methylprednisolone are glucocorticosteroids. Glucocorticosteroids reduce the inßammatory response to tumor tissue by preventing white blood cells from functioning to produce swelling and pain around the tumor site. Corticosteroids induce lysis of peripheral lymphocytes and slow lymphocyte production. Addition of corticosteroids to chemotherapy regimens may augment their efÞcacy.

Clinical Performance. VBCMP. This combination (vincristine, carmustine, cyclophosphamide, melphalan, prednisone) is used in patients who have had a poor response to less toxic combinations like VAD and MP but whose disease is considered too aggressive for single-agent steroid treatment. One trial randomized 479 patients with previously untreated MM to either four-week cycles of MP or Þve-week cycles of VBCMP for one year (Oken MM, 1997). Patients then received MP or VBCMP maintenance therapy at six- and eight-week intervals, respectively, until relapse. Responses were observed in 72% patients receiving VBCMP and in 51% of patients receiving MP. Median remission duration and Þve-year survival rates were signiÞcantly different: median remission durations were 24 and 18 months for VBCMP and MP, respectively; the median survival rates were 26% and 19%, respectively. VBCMP was associated with more nausea, peripheral neuropathy, alopecia, and neutropenia than MP. VMCP/VBAP. This combination (vincristine, melphalan, cyclophosphamide, prednisone/vincristine, carmustine, doxorubicin, prednisone) is one of the most intense regimens and is reserved for patients who have had a very poor response to Þrst-line therapy. The patient undergoes a course of chemotherapy every four weeks. The Þrst combination administered is VMCP; the second is VBAP. The treatment is alternated in this way until remission is achieved. This particular combination allows patients to be treated with the greatest number of drugs. Although more side effects are likely to occur, this treatment should, in theory, reduce the incidence of resistance. A Spanish trial of 449 symptomatic MM patients compared MP with alternating courses of VMCP and VBAP. The objective response rates were 62.7% with VMCP/VBAP and 51% with MP. No signiÞcant differences were found between survival rates in the two treatment groups. However, patients with IgA myeloma who were treated with VMCP/VBAP survived for a median of 38.4 months, signiÞcantly longer than the median MP survival of 20.2 months (Blad´e J, 1993).

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VAMP and CVAMP. Another slight variation on VAD is VAMP (vincristine, doxorubicin, methylprednisolone, cyclophosphamide), in which high-dose dexamethasone is replaced by methylprednisolone, which has less tendency to cause steroidrelated toxicities. Although no studies compare overall response rates between VAD and VAMP, rates appear similar in practice (Gore ME, 1989). This regimen is ideal for patients who have intolerable steroid-related side effects. The addition of cyclophosphamide (CVAMP) can provide greater intensity when needed. Interferon-Alpha Overview. Interferons (IFNs) demonstrate antiproliferative activity against human viruses and human tumor cells. Most studies of IFN in MM have investigated two types, α-2a (Roche/Chugai’s Roferon-A; Takeda’s Canferon A) and α-2b (Schering-Plough’s Introna/Intron A/Viraferon), which appear clinically equivalent and tend to be used interchangeably in markets where both are available. The many trials that have tested IFN as induction and maintenance therapy in MM have produced conßicting results. Less-toxic IFNs incorporated into polyethylene-glycol-coated liposomes are now available; these IFNs are associated with better efÞcacy and tolerability and may lead to better compliance at higher doses. Mechanism of Action. The IFNs are a family of naturally occurring cytokines that exhibit antiviral, antitumor, and immunomodulatory activities. The antitumor effect of IFNs derives from their ability to inhibit cell-cycle progression, induce a reduction in tumor-cell protein synthesis, and inhibit late progenitor colonies. Their immunomodulatory role involves inducing the expression of major histocompatibility antigens and modulating the expression and function of T cells, monocytes, and natural killer cells (large granular lymphocytes that bind to and kill cells by releasing cytotoxins). Clinical Performance. A meta-analysis of all published data found that IFN as induction therapy in combination with chemotherapy prolonged median survival by 4.8 months and overall survival by 3.1 months (Fritz E, 2000). This small improvement is not considered substantial enough to warrant use of IFN as remission-induction treatment. Studies of IFNs as maintenance therapy also produced conßicting results. One study found that IFN therapy prolonged relapse-free survival by 4.4 months and overall survival by 7 months (Fritz E, 2000). These improvements are considered minimal and were accompanied by debilitating ßu-like symptoms, which can severely affect a patient’s quality of life. Another study, by the Nordic Myeloma Group, demonstrated no beneÞt in survival and a signiÞcant reduction in quality of life in the Þrst year of treatment with IFN (Wisloff F, 2000). A meta-analysis showed that IFN maintenance therapy provided only a modest improvement in survival (Myeloma Trialists’ Collaborative Group, 2001); more recent results from an intergroup trial showed no beneÞt from IFN maintenance therapy (Crowley J, 2004). These conßicting data and the cost implication of long-term IFN treatment have limited the use of IFN as maintenance therapy for MM.

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Bisphosphonates Overview. Bisphosphonates can reduce the substantial bone damage caused by MM’s increasing osteoclast activity. Physicians initiate bisphosphonate treatment at diagnosis, regardless of stage, to almost all patients with MM. The only patients who may be excluded are those with renal failure, for whom bisphosphonates are contraindicated. The degree of renal impairment is crucial, given that a moderate dose adjustment may allow safe treatment. The decision to treat remains that of the clinician. Mechanism of Action. Bisphosphonates have a high afÞnity for bone and are preferentially delivered to sites of increased bone formation or resorption. These compounds bind to calcium crystals and bone mineral and are potent inhibitors of osteoclast resorption activity. Third-generation agents like zoledronic acid (Novartis’s Zometa) are 10,000–100,000 times more potent than the Þrstgeneration agents; pamidronate (Novartis’s Aredia, generics), a second-generation agent, is 100 times more potent (Berenson JR, 2001). Currently, clinical trial evidence is difÞcult to compare, given that positive end points differ between trials. Some trials use time-to-Þrst-fracture, termed the Þrst skeletal event. Other trials use radiographic evidence of progression of osteolytic lesions. Formulation. Another difference between bisphosphonates, other than potency, is the method of administration. Zoledronic acid has considerable advantage over pamidronate because it can be infused over 15 minutes rather than 90 minutes. Clodronate (Schering’s Bonefos/Clastoban, generics) is the only oral bisphosphonate that has demonstrated an improvement in MM-associated osteolysis. Clinical Performance. A clinical trial compared 4 or 8 mg zoledronic acid, IV, every four weeks with pamidronate IV every four weeks for 12 months (Rosen LS, 2001). The study included 513 patients with Durie-Salmon stage III MM and at least one osteolytic bone lesion. The incidence of skeletal events was similar in all three groups: 47%, 49%, and 49% for 4 mg zoledronic acid, 8 mg zoledronic acid, and 90 mg pamidronate, respectively. The median time to Þrst event was 12.3 months for 4 mg zoledronic acid and 12 months for pamidronate. The 8 mg dose of zoledronic acid was discontinued because of adverse effects on renal function. A placebo-controlled pamidronate study demonstrated an increase in time to onset of Þrst skeletal event of 21 months, compared with 12 months for placebo, and a reduction in the number of skeletal events per year (Berenson JR, 1996). Oral clodronate (Schering’s Bonefos), a Þrst-generation bisphosphonate, has a beneÞcial effect on the bone symptoms of MM. Trials showed a reduction in radiographic progression of osteolytic lesions in comparison with placebo (Latinen R, 1992). All patients experience a reduction in bone pain and a reduced need for analgesics, and, therefore, an improved quality of life. Unfortunately, none of these studies has shown that bisphosphonates have a signiÞcant impact on overall survival, although a particular subpopulation in a long-term treatment trial with pamidronate lived longer than the placebo group.

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Generally, however, treatment with pamidronate does not increase overall survival (Berenson JR, 1998). A meta-analysis of the role of bisphosphonates in MM reviewed data from 11 studies (Djulbegovic B, 2000): two trials of etidronate (Procter & Gamble’s Didronel); four trials of pamidronate; four trials of clodronate; and one trial of ibandronate (Roche’s Boniva). The reviewers concluded that bisphosphonate therapy in MM reduces vertebral fractures and pain but not mortality. Nonpharmacological Approaches Stem-Cell Transplants. The stem cells used in transplantation can be harvested either from a donor (allo-SCT) or from the patient (auto-SCT). Allogeneic transplants are more effective than autologous transplants in treating MM, but they are associated with high procedure-related mortality and morbidity, and not all patients have a suitable donor. Allogeneic Progenitor Stem-Cell Transplantation. The use of allo-SCT is limited by the availability of an appropriately matched donor and the toxicity of the procedure. The donor must have the same human leukocyte antigen (HLA) type as the patient. Because HLA type is genetically determined, siblings have a one in four chance of having a matching HLA type. Because of the trend toward small families, less than 30% of patients have an appropriate, related HLAmatched donor. Fortunately, matched unrelated donors (MUDs) can be found for an additional 30% of patients through transplant registries. Given the age of most MM patients, allo-SCT is seldom used. As stated previously, allo-SCT is a highly toxic procedure associated with signiÞcant morbidity and mortality. The morbidity is the result of an immunological effect known as graft-versus-host disease (GVHD). In this condition, the donated stem cells recognize the patient as foreign and establish a hostile immune response. The symptoms of GVHD, which can occur in the Þrst few months after transplantation (acute) or can be long-term (chronic), include skin rashes, gastrointestinal toxicities, and liver damage. They increase with the age of the patient and the degree of HLA mismatch. GVHD, however, can be beneÞcial: the infused cells can target and destroy any residual MCs in the patient, a graft-versus-myeloma (GVM) effect, because the graft cells recognize the MCs as foreign and attempt to destroy them (Tricot G, 1996). The presence of this effect boosts the efÞcacy of the allogeneic transplant in comparison with an autologous transplant. Patients are prepared for therapy with various induction regimens of either high-dose chemotherapy or radiotherapy. Allo-SCT is performed only in patients younger than 55 years. This procedure is further restricted to patients who have an HLA-matched donor. Because the median age at diagnosis of MM is 65 years, only a small number of patients are eligible for this treatment. The other limiting factor is the high transplant-related mortality (TRM), a consequence of infection and GVHD (Gahrton G, 1995). These risks have fostered the view that allo-SCT should be restricted to patients

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with prognostic markers indicative of poor outcome, such as abnormalities, deletion of chromosome 13, or hypodiploidy. Several medical centers have reported on the outcome of allo-SCT. All found this procedure to be associated with high TRM in comparison with autologous transplantation (Bensinger W, 1996; Gahrton G, 1995). The European Group for Blood and Marrow Transplantation (EBMT) recently assessed the outcomes of allo-SCT by comparing mortality rates for transplantations performed during two speciÞc time periods: 1983–1993 and 1994–1998. The mortality rate two years after transplantation declined from 46% to 37% in favor of transplantations performed after 1994 (Gahrton G, 2001). This reduction is the result of improved patient selection and better treatment of infections (Gahrton G, 2001). More recent advances in transplantation techniques and donor matching have reduced the rate of TRM to approximately 30% (Majolino I, 2003). Unlike autologous transplants, allo-SCTs are never contaminated with MCs. This lack of contamination results in a greater number of patients obtaining a molecular remission. Polymerase chain reaction (PCR) is used to detect MCs; if no cells are detected, then the patient is in molecular remission. Molecular remission is achieved in 50% of patients treated with allo-SCTs compared with 7% in those treated with autologous transplants (Corradinni P, 1999). The overall relapse-free survival of patients in complete remission after allo-SCT can be as high as 34% at six years (Gahrton G, 1995). The sizable fraction of patients who achieve complete remission after alloSCT makes it an attractive option for eligible patients. If transplant mortality could be reduced further, allogeneic transplantation would be a more promising approach than autologous transplantation for the treatment of younger patients (Gahrton G, 2001). Autologous Progenitor Stem-Cell Transplantation. Because auto-SCT uses a patient’s own progenitor cells, it is more widely applicable than allo-SCT, and because it does not induce GVHD, it is less toxic. The TRM of auto-SCT is approximately 7%, which is comparable with the 2–10% TRM observed during the Þrst six months of standard chemotherapy (Vesole DH, 1996). However, auto-SCT does not invoke a GVM effect. Although the cells are harvested while the patient is in remission or has a low myeloma burden, a small risk remains that MCs may be harvested and reintroduced into the patient following high-dose therapy. In an effort to remove residual MCs, the harvested cells may be treated with ex vivo chemotherapy or with one of several immunological or biological purging techniques that either remove unwanted MCs or select the nonmyeloma cells, which will then be returned to the patient. High-dose melphalan followed by auto-SCT offers the best results when it is given as Þrst-line therapy. Unlike other treatment options, this regimen can elicit a CR in 22% of patients and a PR in 81% (Attal M, 1996). The overall survival achieved by this treatment approach ranges from four to Þve years (Attal M, 1996; Fermand JP, 1998). Despite initial hopes, high-dose melphalan followed by auto-SCT is not curative in the majority of patients; most patients relapse within Þve years. One randomized trial compared this regimen with standard MP

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therapy for Þrst-line treatment. In this study, 200 patients younger than 65 years were randomized to receive either high-dose melphalan followed by auto-SCT or standard chemotherapy with MP. Because many patients were deemed ineligible, only 74 out of 100 patients in the high-dose melphalan/auto-SCT arm received treatment. At Þve years, the event-free survival rates were 28% for the highdose melphalan/auto-SCT group and 10% for the standard MP group; overall survival rates were 52% and 12%, respectively (Attal M, 1996). As this study demonstrates, high-dose melphalan followed by auto-SCT provides a signiÞcant survival beneÞt compared with standard therapy. A randomized study comparing high-dose melphalan/auto-SCT as Þrst-line therapy (early auto-SCT group) with high-dose melphalan/auto-SCT as rescue therapy in patients who relapsed after treatment with VMCP (late auto-SCT group) showed no signiÞcant difference in overall survival between treatment arms (Fermand J, 1998). One important factor to note is that stem-cell harvest was performed before randomization in this study, an approach that does not reßect current medical practice. This procedure allowed optimal collection of peripheral stem cells because any prior alkylator exposure would lead to stem-cell damage. In current medical practice, patients receive combination chemotherapy Þrst and then high-dose melphalan, which would reduce the success of this treatment as rescue therapy. One difference between the two arms was the duration of Þrst remission: 39 months in the early auto-SCT arm and only 13 months in the late auto-SCT arm (Fermand J, 1998). Until recently, most high-dose melphalan/auto-SCT studies have been conducted in patients younger than 60 years—a group that does not reßect the majority of patients with MM, given that median age at diagnosis is typically 65 (Siegel DS, 1999). More-recent studies assessing the impact of age on the outcome of treatment have been constructed with two patient groups of different ages. The younger patient group is selected from a historical database of high-dose melphalan/auto-SCT-treated patients; their characteristics are matched to those of the second group of patients, who are selected to receive high-dose melphalan despite their advanced age (Palumbo A, 1999; Powles R, 2000; Siegel DS, 1999). These trials are subject to a selection bias, which may have an impact on the outcomes. In one trial, the incidence of CR was 20% in the older group of patients versus 43% for the younger group. Event-free survival and overall survival were comparable (Siegel DS, 1999). Historically, patients with renal failure have been excluded from high-dose melphalan/auto-SCT treatment because of fears of poor outcome. A recent study found that patients with abnormal renal function at diagnosis and after stem-cell transplantation have a signiÞcantly reduced three-year survival rate compared with patients who have normal renal function. However, these Þndings do not support the exclusion of patients with poor renal function from high-dose melphalan/ auto-SCT treatment because the beneÞt to overall survival is considerably better than the beneÞt provided by standard-dose alternatives (San Miguel J, 2000). Another research group found that renal failure had no impact on the quality of stem-cell collections (Badros A, 2001). M-protein levels were reduced by at

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least 75% in 70–80% of renal failure patients after high-dose melphalan/autoSCT, and 50% of patients experienced an improvement or normalization of renal function (Lee C, 2002). Tandem Auto-SCT. Given that patients achieve better results with one cycle of high-dose melphalan/auto-SCT than with standard therapy, it follows that two cycles within three to four months may confer an even greater improvement in survival. This double-transplantation procedure is termed tandem auto-SCT or double auto-SCT. The dose-limiting toxicity of auto-SCT is mucositis induced by high-dose melphalan. If mucous membranes are allowed to recover for three to four months, repeated applications of high-dose melphalan can be made safely. Clinical studies have compared double auto-SCT patients with patients selected from a historical database who received standard therapy. Double auto-SCT induced higher PR rates than standard therapy (85% versus 52%) and achieved overall survival of 62 months, compared with 48 months for standard therapy (Barlogie B, 1997). A randomized study by the Intergroupe Francophone du My´elome (IFM) of 399 previously untreated MM patients younger than 60 years showed the superiority of the tandem approach over single auto-SCT (Attal M, 2003). CR or very good PR was experienced by 42% of single-transplantation patients and 50% of double-transplantation patients. The probability of surviving seven years after the diagnosis of MM was 21% for the single-transplantation patients and 42% for the double-transplantation group. The beneÞts of double transplantation were not evident until after four years of follow-up. If patients achieved CR after the Þrst transplantation, they did not beneÞt from the second. Nonmyeloablative Allo-SCT. Several studies are evaluating nonmyeloablative allo-SCT, also referred to as NM-allo-SCT ; the purpose of the chemotherapy is to cause enough immune suppression to allow donor engraftment. It is hoped that these procedures will reduce GVHD and TRM while maintaining the GVM effect. The results seem promising in the short term; the rate of TRM is about 15% (Gerull S, 2004). However, relapse is more frequent than with standard allo-SCT because the GVM effect is reduced (Pandit S, 2002). A variation on NM-allo-SCT is to combine it with an auto-SCT. In this double transplantation procedure, the patient receives a standard auto-SCT followed by NM-allo-SCT. A German study of 17 patients showed that when an NM-alloSCT was given a median of 119 days after an auto-SCT, the rate of CR was 73%. After a median follow-up of 13 months after the NM-allo-SCT, 13 patients were still alive and 12 had not experienced relapse or disease progression (Kr¨oger N, 2002). Researchers found that when the interval between the two transplantations was a median of 62 days, 57% of patients achieved CR and 26% achieved PR (Maloney DG, 2003). TRM was 15–20%. After approximately 18 months of follow-up, 78% of patients were still alive. The Intergroupe Francophone du My´elome designed the IFM9903 and IFM9904 protocols to compare double autoSCT with auto-SCT followed by NM-allo-SCT (Moreau P, 2003). Preliminary data suggest that the latter approach has no effect on event-free survival or overall survival. However, it should be noted that the patients enrolled in the IFM9903

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and IFM9904 trials had serum beta-2 microglobulin levels of >3 mg/dL and deletion of chromosome 13, both of which indicate poor prognosis. Radiation Therapy. The indication for radiation therapy in MM is mainly for symptom management. Radiation therapy is reserved for isolated lytic lesions that have the potential to progress to pathological fractures or cause spinal cord compression. Radiation therapy is also very effective for the treatment of localized pain due to isolated lytic lesions. In the past, TBI has been used as Þrst-line therapy with severe bone marrow suppression (DeVita VT, 2001). Today, TBI is used only rarely as part of the conditioning regimen in some allo-SCT protocols because studies indicate increased toxicity with no improvement in overall survival (Pandit S, 2002). EMERGING THERAPIES Several new agents show promise for the treatment of multiple myeloma (MM). Key areas of activity include the development of thalidomide-derived immunomodulatory drugs and farnesyl transferase inhibitors and the use of agents approved for other neoplastic diseases. Capitalizing on advances in the understanding of the pathophysiology of MM (see “Etiology and Pathophysiology”), researchers have identiÞed possible therapeutic approaches at every level of gene expression, from inhibition of the chromatin machinery active in myeloma cells (MCs) and degradation of messenger RNA (mRNA) to inhibition of receptors or their downstream signaling molecules to post-translational modiÞcation of transcription factors. Targeted therapies based on knowledge of molecular events are at an early stage of clinical development; therefore, the sections that follow discuss the most promising classes of emerging therapies. Other noteworthy compounds in preclinical development that are not discussed in detail include cyclooxygenase-2 inhibitors, cytokine inhibitors, heatshock protein-90 (Hsp-90) inhibitors, IGF-1 inhibitors, monoclonal antibodies, mTOR inhibitors, telomerase inhibitors, and tyrosine kinase inhibitors. In recent years, one emerging therapy has suffered a high-proÞle failure. In 2004, oblimersen (Genta/SanoÞ-Aventis’s Genasense, formerly G-3139) failed to meet its primary endpoint, time to progression, in Phase III trials in MM patients. Oblimersen is an antisense oligonucleotide designed to degrade bcl-2 mRNA and block the production of the Bcl-2 protein that prevents apoptosis. Table 7 summarizes the drug therapies in late-stage development for MM. Most agents in the later stages of development have already been granted orphan drug status for this indication. The accompanying sidebar, “Orphan Drug Status in the Major Markets,” explains the commercial signiÞcance of orphan drug status. Table 8 summarizes key attributes of the therapies in early-stage development for MM. Thalidomide-Derived Immunomodulatory Drugs Overview. Thalidomide-based therapy has revolutionized the treatment of MM in recent years, but the toxicity associated with thalidomide—notably, its well-known teratogenic effects and the risk of venous thromboembolism

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TABLE 7. Emerging Therapies in Development for Multiple Myeloma Compound

Development Phase

Thalidomide-derived immunomodulatory drugs Lenalidomide (Revlimid) United States Europe Japan

Marketing Company

III III —

Celgene Celgene —

— I/II (United Kingdom) —

— Celgene —

Apoptosis stimulators Arsenic trioxide (Trisenox) United States Europe Japan

II II —

Cephalon Cephalon —

Cytotoxic agents Liposomal doxorubicin (Doxil) United States Europe Japan

III — —

Johnson & Johnson — —

Farnesyl transferase inhibitors Tipifarnib (Zarnestra) United States Europe Japan

II — —

Johnson & Johnson — —

VEGF inhibitors Bevacizumab (Avastin) United States Europe Japan

II — —

Genentech/Roche — —

Vatalanib (PTK-787) United States Europe Japan

II — —

Novartis — —

II II —

Curagen/TopoTarget Curagen/TopoTarget —

I — —

Merck — —

II II —

Johnson & Johnson Johnson & Johnson —

CC-4047 (Actimid) United States Europe Japan

Histone deacetylase inhibitors PXD-101 United States Europe Japan Suberanilohydroxamic acid (SAHA) United States Europe Japan MAP kinase inhibitors SCIO-469 United States Europe Japan

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TABLE 7. (continued) Compound

Development Phase

Vaccines Dendritic cell myeloma fusions United States

Europe Japan Radiopharmaceuticals Holmium-166-DOTMP ( 166 Ho-DOTMP, Skeletal Targeted Radiotherapy [STR]) United States Europe Japan

I

Marketing Company

— —

Beth Israel Deaconess Medical Center/DanaFarber Cancer Institute — —

D D —

NeoRx NeoRx —

Samarium-153-EDTMP(153 Sm-EDTMP, Quadramet) United States I Europe — Japan —

Cytogen — —

D = Discontinued.

(VTE)—remains problematic. The shortcomings of thalidomide have prompted the development of more-potent thalidomide analogues with improved safety proÞles. Celgene screened a series of compounds and identiÞed four potential thalidomide-derived immunomodulatory drugs (IMiDs). Two of these, lenalidomide and CC-4047, are in clinical trials for MM; CC-11006 and CC-10050 are at an earlier development stage. Mechanism of Action. The IMiDs have a mechanism of action similar to that of thalidomide. In CC-4047 and in lenalidomide, the phthaloyl ring of thalidomide has been modiÞed to improve the inhibition of tumor necrosis factor-alpha (TNF-α). CC-4047 was generated by the addition of an amino group at the 4 position of the ring; the removal of a carbonyl on the ring created lenalidomide. These modiÞcations resulted in analogues that were up to 50,000 times more potent inhibitors of TNF-α than thalidomide in vitro and more stable (Corral LG, 1999). Like thalidomide, IMiDs stimulate the proliferation of activated T cells, increase levels of interferon-gamma and interleukin-2, and promote naturalkiller-cell-mediated killing of myeloma cells (Davies FE, 2001). IMiDs may also directly inhibit mitogen-activated protein kinase (MAPK) signaling, which may account for the down-regulation of interleukin-6 seen with IMiD treatment of cells in vitro (Hideshima T, 2000). Lenalidomide. Lenalidomide (Celgene’s Revlimid) is the lead compound in a series of small-molecule, orally bioavailable IMiDs in development by Celgene under license from Entremed. The compound was previously known as CDC-501. The FDA and the European Medicines Agency (EMEA) granted lenalidomide

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TABLE 8. Select Investigational Therapies in Early-Stage Development for Multiple Myeloma, 2005 Compound

Development Phase

Cyclooxygenase-2 inhibitors SDX-101 II (United States)

An oral compound and the R-enantiomer of etodolac. Induces apoptosis in cultured myeloma cells that are resistant to killing by bortezomib, dexamethasone, and doxorubicin. Also induces apoptosis in cells isolated from MM patients refractory to treatment (Yasui H, 2005).

Callisto Pharmaceuticals

An oral inhibitor of IL-6 production in myeloma cells in vitro and in a transgenic mouse model of human MM (Tassone P, 2005). In January 2004, the FDA granted it orphan drug status for MM.

Kosan Biosciences

An injectable inhibitor of Hsp-90, a protein chaperone that binds to signaling proteins, including IL-6 and IGF-1 receptor (IGF-1R). When 17AAG binds to Hsp90, it disrupts the Hsp-90-signaling protein complexes, leading to their degradation and cell death. In October 2004, the FDA granted it orphan drug status for MM.

I (United States)

Pfizer

Preclinical (United States)

Novartis

A fully humanized IgG2 antibody with a high affinity for IGF-1R, blocking binding of IGF-1 to its receptor and IGF-1-induced receptor autophosphorylation. CP-751871 has been shown to reduce IGF-1R expression on granulocytes of MM patients. An oral IGF-1R kinase inhibitor that prevents IGF-1-mediated survival and growth in cells isolated from MM patients that are resistant to killing by bortezomib and lenalidomide (Mitsiades CS, 2004[a]).

Hsp-90 inhibitors 17-allylaminoI (United geldanamycin States) (17AAG)

ADW-742

Comments

Salmedix

Cytokine inhibitors Atiprimod II (United States)

IGF-1 inhibitors CP-751871

Developing Company

Monoclonal antibodies Anti-CD40 I (United (SGN-40) States)

Seattle Genetics

Blocks CD40-ligand-induced proliferation and survival in myeloma cells. In August 2004, the FDA granted it orphan drug status for MM.

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TABLE 8. (continued) Compound Anti-CD40 (CHIR-12.12) Anti-CD56

mTOR inhibitors CCI-779

Development Phase

Developing Company

Comments

Preclinical (United States) Preclinical (United States)

Chiron

Preclinical (United States)

Wyeth

An oral inhibitor that significantly reduces subcutaneous growth of myeloma cell lines in a mouse immunodeficiency model (Frost P, 2004).

Geron

An oligonucleotide that disrupts telomere integrity and induces apoptotic death of myeloma cells (Shammas MA, 2004).

Chiron

An oral inhibitor of fibroblast growth factor receptor 3 that induces apoptosis in myeloma cells (Trudel S, 2005).

Telomerase inhibitors GRN-163 Preclinical (United States) Tyrosine kinase inhibitors CHIR-258 I (United States)

ImmunoGen

Blocks CD40-ligand-induced proliferation and survival in myeloma cells. A humanized anti-CD56 antibody conjugated to a cytotoxic agent, DM1. Able to kill myeloma cells expressing CD56 even in the presence of bone marrow stromal cells.

Full source citations appear in ‘‘References.’’

orphan drug status in October 2001 and February 2004, respectively. In February 2003, lenalidomide was designated for fast-track approval by the FDA for relapsed and refractory MM. No development has been reported in Japan. Currently, Celgene is evaluating lenalidomide in two Phase III studies, the Þrst in newly diagnosed MM, the second in relapsed or refractory MM. Lenalidomide was apparently nonteratogenic when tested in New Zealand rabbits, the only animal model in which the fetal toxicity of thalidomide can be detected. Based on these and other preclinical data, no onerous patient registration programs are in place to control the supply of lenalidomide. At the 2003 American Society of Hematology (ASH) meeting, investigators presented Phase II data from 91 evaluable patients with relapsed or refractory MM (Richardson PG, 2003[a]). The patients had a median age of 58; 62% had received prior SCT, 53% had received thalidomide and bortezomib, and 8% had received prior bortezomib. Fifty-three percent of patients had relapsed or refractory disease. Patients were randomized to receive either 15 mg lenalidomide twice daily or 30 mg lenalidomide once daily for three weeks, followed by a one-week rest period. After four weeks in patients with progressive disease and after eight weeks in patients with stable disease, dexamethasone was added at a dose of 40 mg daily for four days every two weeks. The study was continued until patients

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achieved their best response. The rate of complete response (CR) was 10%; 27% of patients achieved partial response (PR) or minimal response. Forty-nine patients were subsequently treated with dexamethasone and 20 patients achieved a response. Median progression-free survival was 5.5 months. SigniÞcant adverse events included thrombocytopenia in 18% of patients and neutropenia in 28%; these myelosuppressive hematologic events occurred less frequently in the group that received 30 mg lenalidomide once daily. Researchers did not observe the side effects associated with thalidomide, such as signiÞcant somnolence, constipation, and neuropathy, with either regimen. Rates of VTE were not reported. A Phase II trial of lenalidomide in 30 newly diagnosed MM patients with a median age of 64 years was presented at the 2004 ASH meeting (Rajkumar SV, 2004[a]). Patients were treated with 25 mg lenalidomide daily on days 1–21 of a 28-day cycle and high-dose dexamethasone (HDD; 40 mg daily) on days 1–4, 9–12, and 17–20. Patients also received 81 mg aspirin daily as prophylaxis against VTE. Therapy was continued until patients achieved their maximal response. The combined rate of CR and PR was 83%. Serious adverse events were experienced by 33% of patients, including one incident each of CD4 T-cell count <200/mm3 , anemia, neutropenia, increased liver enzymes, muscle weakness, agitation, hyperglycemia, cardiac arrhythmia, pneumonitis, erlichiosis, and colonic perforation. No VTE events were observed. Phase III trials of lenalidomide in combination with other agents in a variety of settings are ongoing in the United States, Europe, and elsewhere. The CC-5013-MM-009 (United States) and CC-5013-MM-010 (international) protocols are multicenter, randomized, parallel-group, double-blind, placebo-controlled Phase III studies of lenalidomide plus dexamethasone versus dexamethasone alone. Seven hundred and Þve patients with symptomatic refractory MM (disease progression after two or three cycles of therapy) or relapsed MM (progression after previous treatment) were enrolled in the trials. Previous treatment with thalidomide was not an exclusion criterion, and approximately 75% of patients enrolled had developed resistance to thalidomide therapy. Patients were treated with 25 mg lenalidomide daily plus HDD or placebo plus HDD in 28-day cycles. Lenalidomide was administered at 25 mg once daily on days 1–21 of each cycle, and 40 mg HDD was given on days 1–4, 9–12, and 17–20 of each cycle. After four cycles, the HDD schedule was reduced to 40 mg on days 1–4 of each cycle. The primary endpoint was time to disease progression, calculated as the time from randomization to the Þrst documentation of progressive disease, based on Blad´e response criteria (Blad´e J, 1998). Therapy was continued until the primary endpoint was reached. In March 2005, Celgene announced that the CC-5013-MM-009 and CC-5013MM-010 trials would be unblinded early because lenalidomide had exceeded the prespeciÞed measure of statistically signiÞcant improvement in time-to-disease progression. In May 2005, at the American Society of Clinical Oncology meeting, data were presented from both trials. Interim analysis of CC-5013-MM009 showed that of 171 evaluable patients receiving both lenalidomide and HDD, 61.2% achieved objective response compared with 22.8% of 171 patients

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receiving HDD alone. The percentages of patients achieving CR with lenalidomide and HDD and with HDD alone were 26.5% and 4.1%, respectively. Timeto-disease progression was 60.1 months for lenalidomide patients compared with only 20.7 months for patients receiving HDD alone. In CC-5013-MM-010, 351 patients were evaluated. The response rates were lower than in the U.S. trial: the objective response rates for lenalidomide and HDD and for HDD alone were 47.5% and 18.4%, respectively; CR rates were 9.1% and 1.2%. For patients receiving both lenalidomide and HDD, time to disease progression was 58.0 months compared with 21.7 months for patients receiving HDD alone. The Eastern Cooperative Oncology Group (ECOG) and the Southwest Oncology Group (SWOG) are carrying out two other important, double-blind, Phase III lenalidomide trials. The ECOG-E4A03 trial will randomize 412 patients with newly diagnosed MM to treatment with 25 mg lenalidomide daily plus HDD or a lower dose of dexamethasone (40 mg daily on days 1, 8, 15, and 22 for four cycles). If patients do not respond to therapy, they will be given salvage therapy consisting of 200 mg thalidomide daily on days 1–28 for four cycles and dexamethasone at the dose they had previously received. The SWOG S0232 study will randomize 500 patients with newly diagnosed MM who are not candidates for transplant to treatment with 25 mg lenalidomide daily plus HDD or placebo daily plus HDD for three 28-day cycles separated by 35-day intervals. Patients achieving a response or stable disease after three cycles will switch to 21-day cycles, separated by 28-day intervals, of lenalidomide/HDD or placebo/HDD maintenance therapy until progression or relapse. The key potential shortcoming of lenalidomide is long-term myelosuppressive toxicity. Researchers will carefully examine the degree of myelosuppression in the Phase III studies. CC-4047. CC-4047 (Celgene’s Actimid) is another member of Celgene’s series of small-molecule, orally available IMiDs in development for MM under license from Entremed for use in Europe and the United States. The agent is also known as CC-394 and is in Phase I/II trials in the United Kingdom both as monotherapy and in combination with other agents. Although CC-4047 has exhibited in vitro potency ten times greater than that of lenalidomide, Celgene has not determined, at this time, whether to develop the compound commercially. CC-4047 has been shown to enhance the efÞcacy of vaccination against tumor cells in murine models (Dredge K, 2002). Two Phase I/II trials of CC-4047 have been reported. In the Þrst trial, 24 patients with relapsed or refractory MM and a median of three prior courses of treatment were enrolled in a dose-escalation trial to determine the maximum tolerated dose (MTD) of CC-4047 (Schey SA, 2004). Patients entered one of four CC-4047 dosing groups: 1, 2, 5, or 10 mg daily. The dose-escalation trial followed a standard design based on cohorts of three patients. Three patients begin treatment at the lowest dose, and the incidence of dose-limiting toxicity (DLT; deÞned as any treatment-related grade III or IV toxicity) is assessed over 28 days. If no patients have experienced DLT during that period, another cohort of three patients begins treatment at the next highest dose. If DLT is seen in one

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patient receiving a particular dose, three more patients join at that same dose. If at least two patients experience DLT with a particular dose, the researchers conclude that the previous dose is the MTD, and an additional six patients are given the MTD to gain additional toxicity data. The median duration of treatment in the dose-escalation trial was 28 weeks. Four of the 24 patients achieved CR by 12–43 weeks of therapy; the rate of PR was 54% (13 patients). The responses occurred despite dose reduction, and two of the four CRs occurred after de-escalation steps. The DLT associated with CC-4047 treatment was neutropenia. Across all dose groups, grade III and IV neutropenia occurred in 58% of patients. However, after the initial fourweek period, only one new patient developed neutropenia after escalation to the 5 mg/day dose. Therefore, the MTD of CC-4047 was determined to be 2 mg/day. Two patients in the 2 mg/day group developed VTE after the initial four-week period at four and nine months, and one patient from the 5 mg/day group developed VTE after 11 months. The median overall survival was 21 months. Minor side effects included rash, neuropathy, constipation, diarrhea, nausea, edema, and hypotension. The same researchers carried out a second Phase I trial that used alternateday dosing with CC-4047 (Streetly M, 2004). Data were presented at the ASH 2004 meeting. Twenty patients with relapsed or refractory MM received 1, 2, 5, or 10 mg CC-4047 every other day. Using the same dose-escalation protocol previously described, the MTD for alternate-day dosing proved to be 5 mg every other day; the DLT was neutropenia, which occurred in 45% of patients. After a mean duration of ten months of therapy, two patients had CR, and another ten achieved PR. Median overall survival was 23 months. Apoptosis Stimulators Overview. Apoptosis, or programmed cell death, is a universal and exquisitely efÞcient cellular suicide pathway. As understanding of its vital role in normal development has deepened, researchers have identiÞed numerous genes that encode apoptotic regulators, some of which are familiar oncogenes or tumor suppressor genes. Targeting apoptosis is successful only if the therapeutic index is sufÞcient to selectively destroy cancer cells rather than normal cells. In drug-curable malignancies, such as common pediatric leukemias and certain solid tumors, apoptosis is a prominent mechanism associated with the induction of tumor remission. Many cytotoxic agents’ ultimate mechanisms of action take effect via the general apoptotic pathway. Mechanism of Action. Apoptosis is an evolutionarily conserved cell-death pathway that occurs in a variety of physiological situations. An apoptotic stimulus induces an initiation and commitment phase followed by a degradation phase. This last stage is regulated by cysteine proteases (caspases 1–14) and follows a characteristic pattern of morphological changes that comprises membrane rufßing, mitochondrial dysfunction, cytoplasmic and organelle shrinkage, nuclear contraction, and endonuclease activation, resulting in deoxyribonucleic

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acid (DNA) fragmentation. Two central pathways mediate apoptosis. The type I “extrinsic,” or death receptor, pathway generates an apoptotic signal following the aggregation of death ligands that involves caspase-8 and caspase-10. The type II, or “intrinsic,” pathway generates signals through mitochondria that involve caspase-9. In some cases, type I activation may also proceed down the mitochondrial pathway. A drug that activates apoptosis might achieve a suitable therapeutic index in several ways. First, it might activate a death cascade via a drug target that is uniquely expressed in a cancer cell. Alternatively, it might be delivered to the target tissue in a manner that is selective for the cancer cell. A third possibility—perhaps the most promising one—is to exploit a pathway that is activated by oncogenes in order to provoke apoptosis selectively in cancer cells. It is now clear that oncoproteins can interact with apoptotic regulatory pathways. Thus, overexpression of Myc sensitizes cells to a wide assortment of apoptotic triggers, probably reßecting the role of apoptosis in the intracellular immunity that prevents normal cells from persisting in the body once they acquire cancercausing genetic defects. However, many human tumors that overexpress Myc are highly resistant to apoptotic triggers, probably owing to a variety of downstream lesions that blunt the death pathway. Still, the recognition that oncogenes can sensitize cells to pro-apoptotic treatments suggests that if such lesions can be circumvented, drugs that induce cell death could prove highly selective for cancer cells. Arsenic Trioxide. Arsenic trioxide (Cephalon’s Trisenox) is in Phase II trials in the United States and Europe for use as a single agent and in combination with high-dose steroids. In both cases, multicenter trials are investigating the beneÞt of arsenic trioxide for relapsed MM (up to 110 patients in total). Arsenic trioxide has also completed a Phase II trial in combination with ascorbic acid and melphalan for relapsed/refractory MM, and it has been granted orphan drug status in the United States and Europe for the treatment of MM. Arsenic trioxide is approved in Europe, Japan, and the United States for relapsed or refractory acute promyelocytic leukemia (APL). The results of arsenic trioxide monotherapy and combination therapy have shown clinically signiÞcant responses in relapsed MM patients. Arsenic trioxide’s mechanism of action has not been fully elucidated, but the compound has been shown to inhibit growth and induce apoptosis in MC lines. MCs thrive on IL-6 and vascular endothelial growth factor (VEGF) secreted in the bone marrow microenvironment (BMM; see “Etiology and Pathophysiology”). Arsenic trioxide appears to reduce the production of IL-6 and VEGF in the BMM, an action that partially explains the agent’s growth-inhibitory effects (Hayashi T, 2002). Furthermore, arsenic trioxide activates important caspases in both the extrinsic and intrinsic apoptotic pathways. Caspase-9 is activated by arsenic trioxide in the presence of functional p53; if p53 is absent, caspase-8 and caspase-10 are triggered (Liu Q, 2003). p53 is a tumor suppressor gene involved in the control of normal cellular proliferation, differentiation, and apoptosis. Inactivation of p53 has been observed in many solid tumors, and there is a correlation between p53

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623

mutation frequency and tumor progression. By contrast, p53 mutations are very rare in newly diagnosed MM; they occur in less than 20% of patients. In MM, p53 mutations are associated with advanced or aggressive disease (Portier M, 1992). At the 2002 ASH meeting, researchers presented the results of a Phase II trial for relapsed and refractory MM (Hussein MA, 2002). Patients underwent treatment with arsenic trioxide 0.25 mg/kg/day, IV, for Þve days per week for two weeks, followed by two weeks without treatment. This 28-day cycle was repeated until a response was achieved. Nine of the 21 evaluable patients (43%) achieved a signiÞcant improvement (>25% reduction) in M protein levels. Most patients had received at least two prior chemotherapy regimens. Five patients experienced serious, treatment-related adverse events: leukocytopenia/anemia, anemia/thrombocytopenia, fatigue, febrile neutropenia, and pulmonary edema. Given the modest activity of arsenic trioxide in single-agent trials, researchers investigated the compound’s potential in combination therapy. In August 2001, researchers reported that ascorbic acid enhances the activity of arsenic trioxide against drug-resistant MM in vitro by reducing intracellular glutathione (GSH), which has been implicated as an inhibitor of arsenic-trioxide-induced cell death (Grad JM, 2001). This Þnding provided the basis for the Phase I/II trial of arsenic trioxide in combination with ascorbic acid that is discussed in the following paragraph. At ASH in December 2003, investigators presented the results of a Phase I/II trial investigating the combination of arsenic trioxide with ascorbic acid and melphalan (Borad M, 2003). Ten patients with refractory MM, including Þve with renal failure, underwent treatment with 0.25 mg/kg arsenic trioxide, administered over one to two hours twice weekly; 1,000 mg/day ascorbic acid after each arsenic trioxide dose; and 0.05–0.1 mg/kg/day melphalan for four days every four to six weeks. Therapy was continued until disease progression was observed. All patients responded with a reduction of 37–85% of serum M-protein (MP). Furthermore, all patients with renal failure experienced improvement in renal function with a reduction in the serum creatinine ranging from 35–68%. Four patients progressed, with times-to-progression of 14, 21, 26, and 40 weeks; two of these four patients died with progressive disease. At the time the data were presented, the other six patients remained on the regimen after 15, 15, 20, 44, 52, and 58 weeks, including three of the patients with renal failure. During the trial, three patients required a reduction of the melphalan dose due to myelosuppression. Grade III toxicities included anemia, leukocytopenia, and thrombocytopenia. Arsenic trioxide may prove to be a useful adjunct to low-dose melphalan therapy in heavily pretreated MM patients with renal failure. Cytotoxic Agents Overview. Cytotoxic anticancer drugs are collectively the largest and most established chemotherapy group. Compounds that are marketed for other cancer indications have undergone investigation in MM without yielding improved response rates; these compounds include docetaxel (SanoÞ-Aventis’s Taxotere) and paclitaxel (Bristol-Myers Squibb’s Taxol). However, in an effort to improve

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patients’ tolerance for conventional chemotherapy regimens used to treat MM, modiÞed formulations may be of beneÞt. The most notable example is liposomal doxorubicin (Johnson & Johnson’s [J&J’s] Doxil, Schering-Plough’s Caelyx, Zeneus Pharma’s Myocet). Conventional doxorubicin is a key component of the VAD regimen: vincristine (generics), doxorubicin (PÞzer’s Adriamycin, generics), and dexamethasone (Merck’s Decadron, generics) (see “Current Therapies”). The VAD regimen requires a 96-hour infusion of doxorubicin using a central venous catheter, a procedure that requires hospitalization and may lead to catheter-related infections or thrombosis (Alexanian R, 1990). Doxorubicin is also associated with cardiotoxicity (Rinehart JJ, 1974). Mechanism of Action. The fundamental aim of all cytotoxic agents is to exert selective toxicity on cancer cells. They do so via a variety of pathways. Doxorubicin is an anthracycline. Anthracyclines interact with several different cellular targets, most importantly topoisomerase II. By inhibiting this DNA religation enzyme, anthracyclines exert their cytotoxic effect. Another mechanism leading to cell death, known as DNA intercalation, involves insertion of the anthracycline molecule between base pairs. This phenomenon causes single- and double-stranded breaks in DNA that inhibit cell proliferation. The free radicals generated by the reductive metabolism of anthracyclines may also damage cellular structures. Liposomal Doxorubicin. Liposomal doxorubicin (J&J’s Doxil, ScheringPlough’s Caelyx, Zeneus Pharma’s Myocet) is approved in Europe and the United States for metastatic breast cancer, ovarian cancer, and AIDS-related Kaposi’s sarcoma. This agent consists of doxorubicin incorporated into polyethyleneglycol-coated liposomes. The liposomes signiÞcantly increase the length of time that doxorubicin is circulating in the body, compared with the conventional doxorubicin formulation. Liposomal doxorubicin is associated with less myelotoxicity, cardiotoxicity, and nephrotoxicity than conventional doxorubicin in the treatment of AIDS-related Kaposi’s sarcoma and metastatic breast cancer. Preferential accumulation of liposomes in tumor sites enables administration of lower doses of the active agent (Gabizon AA, 2001). The use of liposomal doxorubicin for MM is currently off-label, but a growing number of clinical trials support its use. J&J is sponsoring Phase III clinical trials to assess thalidomide plus dexamethasone therapy, with or without liposomal doxorubicin, in newly diagnosed MM patients. In December 2004, the FDA granted J&J orphan drug status for the use of liposomal doxorubicin in MM. In a prospective, randomized trial of 259 patients, conventional VAD was compared with VAD that included liposomal doxorubicin as Þrst-line treatment for MM (Dimopoulos MA, 2003). The modiÞed regimen is known as DVd. VAD was administered as a bolus every day for four days; DVd was given as a single-day infusion. The VAD bolus consisted of 0.4 mg vincristine and 9 mg/m2 unmodiÞed doxorubicin, both administered by IV rapid infusion on day one; 40 mg dexamethasone was administered daily for four consecutive days. DVd consisted of 2 mg vincristine and 40 mg/m2 liposomal doxorubicin, given over

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one hour on day 1, and 40 mg dexamethasone, given daily for four consecutive days. Both regimens were given every 28 days for four cycles. The primary endpoint was best response. The response rates for the VAD and DVd regimens were 61.4% and 61.3%, respectively. VAD was associated with greater incidence of neutropenia (20% versus 15%), thrombocytopenia (10% versus 5%), and mild alopecia (55% versus 37%). However, the incidence of erythrodysesthesia (also known as hand-foot syndrome) was much greater in patients treated with DVd (13% versus 2%); erythrodysesthesia is known to be associated with liposomal doxorubicin. Although liposomal doxorubicin has transformed the traditional VAD induction regimen for MM into DVd, which is less myelotoxic, regimens containing vincristine are becoming increasingly outmoded in the United States. Vincristine is neurotoxic (neurotoxicity rates for VAD and DVd are 13% and 15%, respectively [Dimopoulos MA, 2003]). Key shortcomings of liposomal doxorubicin include its high cost compared with that of unmodiÞed doxorubicin and the incidence of erythrodysesthesia, a toxicity that is not associated with unmodiÞed doxorubicin. Two key ongoing Phase III trials are examining liposomal doxorubicin in this indication. The Þrst trial, sponsored by J&J, is a randomized, open-label, multicenter trial comparing thalidomide plus dexamethasone, with or without liposomal doxorubicin, in patients with newly diagnosed MM. In the second study, also sponsored by J&J, bortezomib with or without liposomal doxorubicin will be assessed in relapsed MM. If the outcomes of these trials are favorable for liposomal doxorubicin, this agent will Þnd a role in improving responses in patients who cannot tolerate more aggressive therapies. Farnesyl Transferase Inhibitors Overview. The proto-oncogene ras, which encodes the small GTP-binding protein Ras, is required for both normal intracellular signaling via receptor tyrosine kinases (such as VEGF receptor tyrosine kinase) and appropriate control of cell division. Mutated Ras occurs in up to 39% of newly diagnosed MM patients (Liu P, 1996), and substantial research has been invested in identifying inhibitors of Ras function. Farnesyl transferase inhibitors (FTIs) have emerged as potential therapies for MM and other neoplastic diseases because they catalyze a critical post-translational modiÞcation step required for Ras activity. In preclinical models, unmodiÞed Ras molecules were unable to stimulate MAP kinase, thereby inhibiting cell division (McGeady P, 1995). Tipifarnib (J&J’s Zarnestra, previously R-115777) is the FTI most advanced in clinical development, but promising preclinical data on MCs treated with lonafarnib were presented at ASH in 2004. Lonafarnib potentiated killing by bortezomib, thereby increasing the proportion of apoptotic MCs from 20% to 88% (David E, 2004). Mechanism of Action. Ras normally transduces intracellular signals from receptor tyrosine kinases; Ras signaling activates MAPK, thereby stimulating the promitogenic transcription factors Fos and Jun and promoting cell proliferation.

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Mutations of Ras stimulate uncontrolled cell proliferation by constitutive activation of MAPK. Ras function requires the addition of a farnesyl (lipid) group to a cysteine residue at the carboxy terminus of the protein. This post-translational modiÞcation is catalyzed by farnesyl transferase; small-molecule inhibitors of this enzyme antagonize the modiÞcation of Ras, which in turn generates a naive form of Ras that is incapable of stimulating MAP-kinase-associated cell division. Tipifarnib. Janssen Pharmaceutica and its parent company, J&J, are developing the oral FTI tipifarnib (Zarnestra, previously R-115777) for several hematologic malignancies but not speciÞcally for MM. J&J submitted an NDA to the FDA in January 2005 seeking authorization for tipifarnib for the treatment of acute myeloid leukemia, based on Phase II data. However, in May 2005 the FDA stated that Phase III data would be required for approval. Phase III trials of tipifarnib are ongoing. Investigators at the Lee MofÞtt Cancer Center (Tampa, Florida) conducted a Phase II trial of tipifarnib in 43 patients with relapsed/refractory MM who had received a median of four chemotherapy regimens before the study (Alsina M, 2004). The patients were treated with 300 mg tipifarnib, given orally twice daily for three weeks, followed by a week of rest. The four-week cycle was repeated and treatment continued if a response or disease stabilization was observed. Sixty-four percent of patients experienced disease stabilization, deÞned in this study as a less than 50% reduction of MP levels. Tipifarnib therapy suppressed farnesyl transferase activity in the bone marrow and peripheral blood mononuclear cells and reduced levels of phosphorylated Akt and STAT3 in bone marrow cells. However, this inhibition of signaling did not correlate with the rate of disease stabilization. The most frequent adverse event was fatigue, which occurred in 66% of patients; of these patients, 12.5% experienced grade III/IV fatigue. Other side effects included diarrhea, nausea, neuropathy, anemia, and thrombocytopenia. Tipifarnib may be used in combination with other agents to treat MM. In a presentation of preclinical data at the Tenth International Multiple Myeloma Workshop in April 2005, investigators showed that the combination of bortezomib and tipifarnib is effective and synergistic in a human MC line (Lonial S, 2005). The combination achieved rapid caspase-dependent and -independent apoptosis. In particular, at a dose of 4 µM tipifarnib, the combination killed more than 90% of cells versus 40% for bortezomib alone. Clinical studies of the same combination are planned. VEGF Inhibitors Overview. As discussed in the “Etiology and Pathophysiology” section, VEGF is a growth factor vital for the survival of MCs. Several current and emerging therapies for MM affect VEGF indirectly through signal transduction, but only VEGF inhibitors selectively block this particular angiogenic pathway. Several big pharmaceutical companies are developing VEGF inhibitors for the treatment of tumors, but their development programs do not necessarily include MM patients; instead, academic researchers are independently testing

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some promising compounds in MM patients. GlaxoSmithKline’s VEGF-2 receptor tyrosine kinase inhibitor (GW-786034) is in Phase II trials at the DanaFarber Cancer Institute in Boston. The following sections discuss bevacizumab (Genentech/Roche’s Avastin) and vatalanib (Novartis/Schering’s PTK-787), both of which are in Phase II development. Mechanism of Action. Although VEGF is a target for inhibiting angiogenesis, inhibition of a single stimulatory signal is likely to have an effect only in a subset of patients and therefore cannot be universally applied to all cancers, especially end-stage cancers, when multiple proangiogenic factors are expressed. A more promising approach may be to inhibit VEGF receptors; there are only two VEGF receptors versus four VEGF ligands. In addition, because the VEGF receptor is expressed on normal endothelial cells, these cells have a much lower mutation rate than MCs and are therefore less likely to become resistant to a VEGF-receptor-targeted drug. Bevacizumab. Bevacizumab (Genentech/Roche’s Avastin) is a recombinant, humanized monoclonal antibody to VEGF that has been approved for colorectal cancer in Europe and the United States. Roche acquired worldwide rights to bevacizumab except in the United States, where Genentech retained exclusive rights. In June 2003, the NCI initiated a Phase II trial of bevacizumab in patients with relapsed/refractory MM. Patients were randomized to receive an IV infusion of bevacizumab over 30–90 minutes on days 1, 15, 29, and 43, in combination with thalidomide once daily, versus bevacizumab alone. No clinical data have been made public so far. Vatalanib. Vatalanib is an orally administered, small-molecule angiogenesis inhibitor that targets the VEGF-1, -2, and -3 receptors. The lead indication for vatalanib is colorectal tumors, for which the compound is undergoing Phase III development in Europe and the United States by Novartis and Schering. Investigators at the Dana-Farber Cancer Institute are using vatalanib in Phase II trials to treat MM. A research group at the institute has shown that the agent inhibits growth and migration of MCs in the BMM (Lin B, 2002). Histone Deacetylase Inhibitors Overview. A wide range of histone deacetylase (HDAC) inhibitors are in development for MM and other cancers. This approach is a good example of targeting epigenetic phenomena associated with the disease (see “Etiology and Pathophysiology”). Levels of histone acetylation are probably regulated by histone acetyltransferase (HAT) and HDAC complexes. Hyperacetylation of histones correlates with recruitment of HATs and increased transcription. Conversely, hypoacetylation correlates with the recruitment of HDACs and reduced transcription. Clinicians have also shown that some HAT complexes contain known transcriptional activators and some HDAC complexes contain known transcriptional repressors (Struhl K, 1998). Loss of HAT function has been implicated

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in the development of cancers, while inhibition of HDAC activity, and therefore increased gene expression, may lead to differentiation, cell cycle arrest, and apoptosis of tumor cells (Marks PA, 2000). These observations prompted investigators to treat cultured MCs with HDAC inhibitors, and they found that such compounds induced apoptosis in the malignant cells (Mitsiades N, 2003). This section reviews the most promising HDAC inhibitors in development for MM. Mechanism of Action. The mechanisms by which HDAC inhibitors exert their effect are not fully understood. Treatment of cells with these agents activates a small set of genes that usually includes the cell-cycle kinase inhibitor p21W AF 1 and other kinases associated with cell growth, such as p16INK4a and p27Kip (Richon VM, 2002). PXD-101. Curagen and TopoTarget are developing a series of HDAC inhibitors, of which PXD-101, a novel hydroxamate-type compound, is the Þrst to enter clinical development in Europe and the United States. No development has been reported in Japan. The compound is currently in Phase II development both as a single agent and in combination with dexamethasone for the treatment on multiple myeloma. A Phase I study of PXD-101 was designed to evaluate its safety, efÞcacy, pharmacokinetics, and pharmacodynamics and to accrue 24–36 patients (Plumb JA, 2004). In September 2004, investigators presented preliminary data from the study at the 16th joint meeting of the European Organization for Research and Treatment of Cancer, the NCI, and the American Association for Cancer Research (EORTC-NCI-AACR). Patients received 150–600 mg/m 2 PXD-101 as a 30-minute IV infusion daily for Þve days. Blood samples were taken at regular intervals to monitor acetylation of histones extracted from peripheral blood mononuclear cells. In all patients, acetylation of histone H4 increased following treatment with PXD-101. In the 150 mg/m2 dose group, acetylation had returned to base levels by two hours after treatment. After treatment with the 600 mg/m2 dose, acetylation levels remained elevated for two hours after infusion and, in some patients, were sustained for six hours. Induction of proteins involved in cellcycle arrest and apoptosis was noted, including p21WAF1 , p19INK4d , and Apaf-1. Another Phase I study involving 15 patients is also ongoing. In February 2005, Phase II trials of PXD-101 began. The open-label, multicenter study is expected to enroll approximately 50 MM patients who have failed two previous therapies. Patients will be treated for two 21-day cycles with 900 mg/m2 PXD-101 daily for Þve days, followed by two weeks of monitoring. If patients achieve an objective response to therapy, they will receive additional cycles of PXD-101 up to a maximum of six. If patients experience disease progression, they will be treated with up to six cycles of PXD-101 plus 40 mg dexamethasone on days 2–5 and 10–13 of each cycle. The study is expected to be completed by mid 2006. Suberanilohydroxamic Acid. Merck is developing suberanilohydroxamic acid (SAHA); the company gained the compound when it acquired Aton Pharma

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in 2004. Aton Pharma had been investigating the potential of SAHA in several cancers, including MM. In October 2003, Aton initiated a Phase I trial in patients with advanced MM, and the FDA granted orphan drug designation for this indication. At this time, Merck is pursuing the indication of cutaneous T-cell lymphoma, an aggressive form of non-Hodgkin’s lymphoma, and expects to submit an NDA to the FDA in 2006. At the ASH meeting in 2004, investigators presented preliminary data from a Phase I study carried out by Aton Pharma in collaboration with researchers at the Dana-Farber Cancer Institute (Richardson PG, 2004[b]). A dose-escalation trial was designed to determine the MTD of SAHA; oral doses of 200, 250, and 300 mg were given daily for Þve days, followed by two days of rest, in fourweek cycles. At the time of the ASH meeting, Þve patients with relapsed MM and three patients with relapsed, refractory MM had been treated with 200 mg or 250 mg SAHA for a median of three cycles (range two to nine); seven patients were available for evaluation. One patient in the 250-mg-dose group developed DLT with grade III fatigue, prompting dose reduction in the next cycle. Other, less severe side effects included fatigue, diarrhea, indigestion, and dehydration. No signiÞcant myelosuppression, neuropathy, or sedation were observed. In the seven evaluable patients, minimal responses were observed in two patients, two patients had stable disease, and two patients experienced disease progression. The trial is ongoing. SAHA and other HDAC inhibitors may be most useful when combined with other agents. Some investigators involved in the Phase I trial of SAHA showed that cultured MCs treated with SAHA are nearly twice as susceptible to killing by bortezomib (Mitsiades CS, 2004[b]). MAP Kinase Inhibitors Overview. Oral, small-molecule inhibitors of mitogen-activated protein (MAP) kinase are in development for the treatment of cancers as well as inßammatory disorders. Few agents have advanced to late-stage clinical trials owing to the high discontinuation rate of early-stage therapies due to their toxicity or lack of demonstrated efÞcacy. This section discusses SCIO-469, which is furthest advanced. Mechanism of Action. The enzyme p38 MAP kinase is part of an intracellular signal transduction cascade that is involved in regulating the expression of several cytokines, including IL-6, VEGF, IL-1β, TNF-α, and receptor activator of nuclear factor kappaB ligand (RANKL). Researchers believe that targeting this pathway may provide effective and complete blockade of cytokines that are up-regulated in MCs. The potential therapeutic beneÞt of targeting the MAP kinase pathway in MM is also suggested by preclinical studies with selective inhibitors of the p38 MAP kinase pathway. SCIO-469. Scios, a J&J subsidiary, is developing SCIO-469, an oral p38 MAP kinase inhibitor for the treatment of MM and inßammatory diseases. The agent

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is in Phase II development in the United States and Europe; no development has been reported in Japan. Scios initiated a Phase II study of SCIO-469 in patients with relapsed/ refractory MM in May 2004. This open-label, active-control, safety/efÞcacy study was to enroll 60 patients. At this time, no clinical data have been made public. SCIO-469 has been shown to inhibit MC proliferation in a mouse xenograft plasmacytoma model of MM (Nguyen AN, 2004). The compound also inhibited secretion of osteoclast-stimulating factors (RANKL, IL-11, and MIP1a) in the BMM in vitro. Like the HDAC inhibitors, the MAP kinase inhibitors may be most useful in combination therapy. SCIO-469 has been shown to enhance the cytotoxicity of bortezomib even in bortezomib-resistant MC lines and MCs harvested from patients (Hideshima T, 2004). Vaccines Overview. Immunotherapeutic approaches for MM involve both speciÞc and nonspeciÞc stimulation of the immune system. SpeciÞc stimulation occurs when tumor antigens are used in a vaccine formulation to direct the immune system against the MCs, avoiding healthy cells. NonspeciÞc stimulation occurs when an adjuvant (a substance such as keyhole limpet hemocyanin [KLH] that evokes a generalized immune response) is administered. The idea of using vaccines to stimulate therapeutic tumor-antigen-speciÞc immune responses holds promise as a complementary approach to current treatment options in most cancers. Cancer vaccines may incorporate both tumor antigens and an immunostimulatory adjuvant. Questions that must be addressed by clinical trials include choice of antigen and optimal timing of vaccination. Diverse vaccines are under investigation for MM. Much interest has centered on vaccines directed speciÞcally against the myeloma idiotype of an individual patient, but idiotypes are only weakly antigenic and have not been used successfully as immunotherapy. Subsequently, idiotypic vaccination has been combined with dendritic cells (DCs), thus exploiting the fact that DCs are the most potent antigen-presenting cells in the immune system. Several research institutes are investigating DC-based vaccines in MM patients; this section discusses the Phase I experience of the Beth Israel Deaconess Medical Center, Boston, and the Dana-Farber Cancer Institute. Cellular vaccines may offer an alternative therapeutic approach, using MCs to stimulate an immune response. A large body of evidence shows that allogeneic progenitor stem-cell transplantation (allo-SCT) elicits a graft-versus-myeloma effect, but autologous progenitor stem-cell transplant (auto-SCT) does not (see “Current Therapies”). By activating T cells in the period immediately following auto-SCT, it may be possible to enhance the killing of MCs. DNA vaccines and gene-modiÞed vaccines are also being investigated for MM but are at a much earlier stage of development. Mechanism of Action. Cancer vaccines are designed to stimulate the immune system to launch a response against the speciÞc epitope contained in the

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vaccine. In general, research has shown that the most effective antitumor immune responses are achieved by stimulating T lymphocytes, which can recognize and kill tumor cells directly. Immunostimulatory adjuvants also activate other cells of the immune system, including macrophages, natural killer (NK) cells, and DCs; these cells are capable of recognizing and destroying tumor cells. Dendritic Cell Myeloma Fusions. Attempts by biotechnology companies to produce efÞcacious vaccines have not been successful; for example, the development of Dendreon’s Mylovenge, stablemate to the prostate cancer vaccine Provenge, has stalled. Instead, academic researchers lead DC vaccine research, and at this time, it is not clear what commercial opportunity lies in such tailormade approaches. Investigators at the Beth Israel Deaconess Medical Center and the Dana-Farber Cancer Institute are developing a patient-speciÞc vaccine using autologous DCs fused with MC lines. Mature DCs are harvested from patients and then fused with MCs. The resulting fusion cells (FCs) present tumor antigens and stimulate cytotoxic, T-lymphocyte responses against autologous tumors. FCs have been shown to effectively protect mice from proliferation of MCs (and therefore from death) when MCs are injected after vaccination with FCs (Gong J, 2002). At the ASH meeting in 2004, investigators presented data from a Phase I trial of FC therapy (Avigan D, 2004). Four patients were vaccinated with FCs plus GM-CSF, a hormone that plays a key role in stimulating the immune response. Three patients received 1 × 106 FCs; the fourth patient was vaccinated with 2 × 106 FCs. Vaccination resulted in T-cell activation as measured by increased percentages of CD4+ and CD8+ T cells expressing interferon-γ . The researchers are now evaluating whether vaccination has affected prognostic markers in the patients. Vaccine-related adverse events included muscle aches and stiffness, transient fever, pruritis, rash, and fatigue. GVAX. NOTE: at the time of composing this reference, this GVAX has been discontinued. Radiopharmaceuticals Overview. Painful osteolytic bone lesions can be associated with MM and other cancers that have metastasized to the bone. The lesions may be severe and difÞcult to treat. Radiopharmaceuticals incorporate a bone-seeking radioisotope to deliver radiation directly to bone metastases, thereby reducing the side effects associated with external-beam radiotherapy. The use of radiopharmaceuticals for the treatment of bone pain related to cancer has a long history. Until recently, the adverse reactions associated with these agents tended to outweigh their clinical beneÞt. Two types of radiopharmaceutical have been used in the treatment of bone metastases: the pure, “bone-seeking” radioisotopes, such as strontium-89, and radioisotopes that are complexed with other bone-seeking agents, such as samarium-153-EDTMP (153 Sm-EDTMP; Cytogen’s Quadramet). Side effects, including leukocytopenia and thrombocytopenia, are problematic in the treatment

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of many cancers. However, such myelosuppression may be of beneÞt in the treatment of MM. As previously discussed, auto-SCT does not cure MM because the myeloablative conditioning regimen that precedes transplantation does not kill every MC (see “Current Therapies”). The combination of targeted radiopharmaceuticals with standard high-dose melphalan and auto-SCT may provide a better method for purging tumor cells. Mechanism of Action. Radiopharmaceuticals are designed to deliver high doses of radiation to tumor sites throughout the skeleton with minimal damage to organs outside the bone. Prior to administration of radiopharmaceutical therapy, the patient’s peripheral blood stem cells are collected; after radiopharmaceutical therapy, the patient receives an infusion of the collected stem cells to restore bone marrow function. Holmium-166-DOTMP. Holmium-166-DOTMP (166 Ho-DOTMP; NeoRx’s Skeletal Targeted Radiotherapy [STR]), a complex of holmium-166 and a small molecule that targets bone, was in Phase III development for the treatment of MM in Europe and the United States. The half-life of 166 Ho-DOTMP is 26.8 hours, which is about half that of 153 Sm-EDTMP (46.3 hours). Theoretically, this halflife means that reinfusion of the patient’s harvested stem cells can take place only six to eight days after 166 Ho-DOTMP therapy; reinfusion is performed approximately 12 days after 153 Sm-EDTMP therapy and auto-SCT. Phase I/II trials assessed one of three doses of 166 Ho-DOTMP in combination with one of three doses of melphalan in 83 patients (Giralt S, 2003). Patients had received a median of two therapies before transplantation, and 23 patients (28%) had disease refractory to induction chemotherapy. Fifteen patients received 20 Gy of 166 Ho-DOTMP and 140 mg/m2 of melphalan (MEL140; 5 patients), 140 mg/m2 of melphalan combined with 8 Gy total body irradiation (MEL140TBI; 6 patients), or 200 mg/m2 of melphalan (MEL200; 4 patients). In the 30 Gy 166 Ho-DOTMP group, the numbers of patients receiving MEL140, MEL140TBI, and MEL200 were 4, 9, and 7, respectively. The highest-dose group was 166 Ho-DOTMP 40 Gy, and most patients received MEL200 (30 patients); only seven patients were treated with MEL140 and 10 with MEL140-TBI. 166 HoDOTMP was given seven to ten days before auto-SCT and melphalan three days before transplantation. TBI was divided into four fractions, given three, two, and one day before auto-SCT, with the Þnal fraction given on the day of the transplantation. All patients received prophylaxis for hemorrhagic cystitis: one of three participating centers instituted continuous bladder irrigation (CBI) until the day following 166 Ho-DOTMP administration to minimize the contact of radioactive urine with the bladder; the other two centers used hydration at 200 mL/h with IV saline-dextrose, forced diuresis, and voiding every two hours. Across all dosing groups, 29 (35%) patients achieved CR. When data were analyzed according to melphalan dose, higher rates of CR were observed with MEL140-TBI (40%) and MEL200 (39%) than with MEL140 (19%). Patients who

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responded to induction chemotherapy had CR and PR rates of 40% and 22%, respectively, compared with patients with primary refractory disease, whose rates of CR and PR were 23% and 45%, respectively. These data agree with data seen in the IFM 94 prospective, randomized trial of single and double auto-SCT (Attal M, 2003). At the time of publication of the 166 Ho-DOTMP trial, the median survival time had not been reached because the median follow-up time was only 31.4 months (Giralt S, 2003). Overall survival at one and two years was 84% and 71%, respectively; event-free survival was 22 months. This result is also comparable to the IFM 94 Þndings: at median follow-up of 75 months after a single auto-SCT, event-free survival and overall survival were 25 and 48 months, respectively (Attal M, 2003). Grades II/III hemorrhagic cystitis were observed in 27 166 Ho-DOTMP patients, only one of whom had been treated with CBI (Giralt S, 2003). CBI with hydration appeared to minimize bladder toxicity. Seven patients developed a serious, treatment-related, delayed toxicity, referred to as thrombotic thrombocytopenia purpura/hemolytic-uremic syndrome (TTP/HUS). At the request of the FDA, Phase III trials were delayed and NeoRx initiated a Phase II dosimetry trial. Data from this study were submitted directly to the FDA in February 2003; in April 2003, the FDA removed its restriction on Phase III trials. The Phase III trial began in March 2004. The study was designed to enroll 240 patients with primary refractory MM who have been undergoing treatment for less than 18 months. All patients will receive MEL200 followed by auto-SCT; before the myeloablative conditioning, half of the patients will be randomized to treatment with 750 mCi/m2 166 Ho-DOTMP. The primary end point of the trial is CR at six months post-transplantation. Event-free survival and OS will also be assessed. In May 2005, NeoRx announced that it was discontinuing clinical development of 166 Ho-DOTMP. Samarium-153-EDTMP. 153 Sm-EDTMP is approved for the relief of metastatic bone pain in Europe and the United States. Cytogen is investigating whether 153 Sm-EDTMP has direct antitumor activity and enhances the activity of conventional therapies. At this time, the company’s focus is the treatment of hormone-refractory prostate cancer. Physicians at the Mayo Clinic (Rochester, Minnesota) have used Cytogen’s product in an uncontrolled Phase I study in MM patients who were scheduled to undergo auto-SCT (Dispenzieri A, 2005). Twelve patients with MM were enrolled in a standard dose-escalation trial based on cohorts of three patients receiving doses of 6, 12, 19.8, and 30 mCi/kg 153 Sm-EDTMP and MEL200. No DLT was observed, but the investigators decided to change the protocol from dosing based on body mass to 40 Gy targeted to the red bone marrow, where myeloproliferation occurs. Six patients received the 40 Gy targeted dose. The time between 153 Sm-EDTMP dosing and MEL200 ranged from 7 to 12 days; the median time was 11 days. Prophylaxis for hemorrhagic cystitis consisted of oral and IV ßuids before 153 Sm-EDTMP administration and 200 mL/h hydration after infusion until the day following radiopharmaceutical treatment.

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Five (28%) patients achieved CR and another Þve had very good PR. Six patients died at a median of 21 months. No hemorrhagic cystitis or TTP/HUS was observed at a median follow-up of 31 months. The difference seen in the incidence of adverse events with 153 Sm-EDTMP and 166 Ho-DOTMP may be due to a more aggressive hydration strategy in the Mayo Clinic study or to differences in the retention times or emission path lengths of the two agents. The 40 Gy targeted dose will be used in Phase II studies at the Mayo Clinic. REFERENCES Aitchison RG, et al. Vincristine, adriamycin and high-dose steroids in myeloma complicated by renal failure. British Journal of Cancer. 1990;61:765–766. Alexander F, et al. Cancer registration of leukaemias and lymphomas: results of a comparison with a specialist registry. Community Medicine. 1989;11(2):81–89. Alexanian R, et al. Treatment for multiple myeloma: combination chemotherapy with different melphalan dose regimens. Journal of the American Medical Association. 1969;208(9):1680–1685. Alexanian R, et al. High-dose glucocorticoid treatment of resistant myeloma. Annals of Internal Medicine. 1986;105:8–11. Alexanian R, et al. VAD-based regimens as primary treatment for multiple myeloma. American Journal of Hematology. 1990;33(2):86–89. Alexanian R, et al. Primary dexamethasone treatment of multiple myeloma. Blood. 1992;80(4):887–890. Alsina M, et al. Farnesyl transferase inhibitor tipifarnib is well tolerated, induces stabilization of disease, and inhibits farnesylation and oncogenic/tumor survival pathways in patients with advanced multiple myeloma. Blood. 2004;103(9):3271–3277. Anderson DM, et al. A homologue of the TNF receptor and its ligand enhance T-cell growth and dendritic-cell function. Nature. 1997;390:175–179. Attal M, et al. A prospective, randomized trial of autologous bone marrow transplantation and chemotherapy in multiple myeloma. Intergroupe Franc¸ais du My´elome. New England Journal of Medicine. 1996;335(2):91–97. Attal M, et al. Single versus double autologous stem-cell transplantation for multiple myeloma. New England Journal of Medicine. 2003;349(26):2495–2502. Attal M, et al. Maintenance treatment with thalidomide after autologous transplantation for myeloma: Þrst analysis of a prospective randomized study of the Intergroupe Francophone du My´elome (IFM 99 02). Proceedings of the American Society of Hematology. 2004. Abstract 535. Avigan D, et al. Dendritic cell myeloma fusions stimulate antitumor immunity: results from preclinical studies and a clinical trial. Proceedings of the American Society of Hematology. 2004. Abstract 751. Badros A, et al. Results of autologous stem-cell transplant in multiple myeloma patients with renal failure. British Journal of Haematology. 2001;114(4):822–829. Baris D, et al. Socioeconomic status and multiple myeloma among U.S. blacks and whites. American Journal of Public Health. 2000;90:1277–1281. Barlogie B, et al. Effective treatment of advanced multiple myeloma refractory to alkylating agents. New England Journal of Medicine. 1984;310:1353–1356.

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Barlogie B, et al. Superiority of tandem autologous transplantation over standard therapy for previously untreated multiple myeloma. Blood. 1997;89(3):789–793. Barlogie B, et al. Long-term follow-up after high-dose therapy for high-risk multiple myeloma. Bone Marrow Transplantation. 1998;21(11):1101–1107. Barlogie B, et al. Comparable survival in multiple myeloma (MM) with high-dose therapy (HDT) employing MEL 140 mg/m2 + TBI 12 Gy autotransplants versus standard dose therapy with VBMCP and no beneÞt from interferon (IFN) maintenance: results of intergroup trial S9321. Proceedings of the American Society of Hematology. 2003. Abstract 135. Barosi G. Management of multiple myeloma and related-disorders. Guidelines issued by the Italian Society of Hematology, the Italian Society of Experimental Hematology, and the Italian Group for Bone Marrow Transplantation. Haematologica. 2004;89:717–741. Bataille R, et al. Beta-2-microglobulin in myeloma: optimal use for staging, prognosis, and treatment—a prospective study of 160 patients. Blood. 1984;63(2):468–476. Bataille R, Harousseau J-L. Multiple myeloma. New England Journal of Medicine. 1997;336(23):1657–1664. Bensinger W, et al. Allogeneic marrow transplantation for multiple myeloma: an analysis of risk factors on outcome. Blood. 1996;88(7):2787–2793. Berenson JR, et al. EfÞcacy of pamidronate in reducing skeletal events in patients with advanced multiple myeloma. Myeloma Aredia Study Group. New England Journal of Medicine. 1996;334(8):488–493. Berenson JR, et al. Long-term pamidronate treatment of advanced multiple myeloma patients reduces skeletal events. Myeloma Aredia Study Group. Journal of Clinical Oncology. 1998;16(2):593–602. Berenson JR. New advances in the biology and treatment of myeloma bone disease. Seminars in Hematology. 2001;38(2 suppl 3):15–20. Berenson JR, et al. American Society of Clinical Oncology clinical practice guidelines: the role of bisphosphonates in multiple myeloma. Journal of Clinical Oncology. 2002;20: 3719–3736. Bezabeh S, et al. Does benzene cause multiple myeloma? An analysis of the published case-control literature. Environmental Health Perspectives. 1996;104(suppl 6): 1393–1398. Blad´e J, et al. Alternating combination VCMP/VBAP chemotherapy versus melphalan/ prednisone in the treatment of multiple myeloma: a randomized multicenter study of 487 patients. Journal of Clinical Oncology. 1993;11(6):1165–1171. Blad´e J, et al. Criteria for evaluating disease response and progression in patients with multiple myeloma treated by high-dose therapy and haemopoietic stem-cell transplantation. Myeloma Subcommittee of the EBMT. European Group for Blood and Marrow Transplant. British Journal of Haematology. 1998;102(5):1115–1123. Boffetta P, et al. A case-control study of multiple myeloma nested in the American Cancer Society Prospective Study. International Journal of Cancer. 1989;43:554–559. Borad M, et al. Melphalan, arsenic trioxide, and ascorbic acid (MAC) is effective in the treatment of refractory and relapsed multiple myeloma (MM). Proceedings of the American Society of Hematology. 2003. Abstract 827. Borrello I, et al. Autologous tumor combined with a GM-CSF-secreting cell line vaccine (GVAX) following autologous stem-cell transplant (ASCT) in multiple myeloma. Proceedings of the American Society of Hematology. 2004. Abstract 440.

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Non-Small-Cell Lung Cancer

ETIOLOGY AND PATHOPHYSIOLOGY Introduction Lung cancer is divided into two major types based on the microscopic appearance of the tumor cells: small-cell lung cancer (SCLC) and non-small-cell lung cancer (NSCLC). NSCLC is more common than SCLC and generally grows and spreads more slowly. NSCLC comprises three major histological subtypes: • • •

Adenocarcinoma. Squamous cell carcinoma (epidermoid). Large-cell carcinoma (undifferentiated).

Approximately 10% of all NSCLC cases involve two or more of these histological subtypes. Cigarette smoking is by far the most important risk factor for NSCLC: 85% of all lung cancer cases are attributable to this habit (Reddy AA, 2000). Although the link between NSCLC and smoking is well established, the genetic pathogenesis of NSCLC has yet to be fully elucidated, and researchers estimate that more than 20 genetic events occur during the development of lung cancer (Wistuba II, 2001). Determining the prognostic signiÞcance of each genetic alteration and designing new therapies that directly target these alterations continues to be a signiÞcant challenge for researchers interested in preventing NSCLC and improving treatment outcomes. Etiology Genetic Pathogenesis of Non-Small-Cell Lung Cancer. The genetic pathogenesis of NSCLC is very complex, and multiple genetic mutations are Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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present in both precancerous and cancerous cells (Wistuba II, 2001). The signiÞcance of each genetic mutation and the manner in which genetic mutations interact to cause NSCLC are poorly understood; these subjects are the focus of immense scientiÞc endeavor. Table 1 brießy describes genetic mutations implicated in NSCLC, together with their clinical signiÞcance, and the following subsections discuss several of the most interesting genetic mutations. p53. Mutations in the tumor suppressor gene (TSG) p53 have been linked to most cancers, including NSCLC, and are often important in the cascade of events that lead to the malignant phenotype. Researchers are beginning to unravel the clinical implications of these mutations. Studies have demonstrated that NSCLC tumors with high levels of p53 mutations are less responsive to both radiation and cisplatin-based chemotherapy. Researchers hypothesize that an intact p53 apoptosis-regulatory mechanism is required for the activity of many DNAdamaging cytotoxic drugs. Fragile Histidine Triad. The fragile histidine triad (FHIT ) is a candidate TSG that is deleted in more than 70% of NSCLC patients (Sozzi G, 1998). The abnormal expression of the FHIT gene is more frequent in squamous cell NSCLC than in the adenocarcinoma form of the disease and is associated with cigarette smoking (Tseng JE, 1999). Researchers hypothesize that FHIT deletion is one of the earliest genetic events in NSCLC pathogenesis (Fong KM, 1997) and that loss of the gene may be critical to the initial stages of NSCLC pathogenesis. The decrease in FHIT protein expression is probably followed by the accumulation of additional mutations during the neoplastic process that leads to invasive and metastatic cancer (Huebner K, 1999). Researchers have proved the importance of FHIT to maintenance of the apoptotic pathway (a pathway for programmed cell death). In lung cancer cells, the replacement of abnormal FHIT genes with normal FHIT genes causes apoptosis and cell-cycle arrest in vivo and in vitro (Siprashvili Z, 1997; Ji L, 1999). These results suggest that FHIT gene therapy is a potential therapeutic approach in the prevention of early lesions associated with lung carcinogenesis. K-ras. Among the oncogenes, the ras family is thought to be one of the most important in NSCLC. The ras oncogene family (K-ras, H-ras, and N-ras) encodes cell-membrane guanosine-triphosphate (GTP) proteins that are involved in cell growth. Mutations that activate the ras oncogene result in inappropriate, prolonged signaling for continued cell division. To be active in the cell, ras oncogenes must have a lipid modiÞcation (farnesylation), which is regulated by farnesyl transferase. Epidermal Growth Factor Receptor. Recent advances in understanding of the epidermal growth factor receptor (EGFR) family signaling system and its critical role in cancer have led to the development of new targeted therapies—speciÞcally, monoclonal antibodies and tyrosine kinase inhibitors. In general, the EGFR family is a group of four structurally similar growth factor receptors with tyrosine-kinase activity—HER-1/erb-B1, HER-2/erb-B2, HER-3/erb-B3 and

TABLE 1. Select Genetic Mutations in Non-Small-Cell Lung Cancer Gene/ Protein

Function

Tumor suppressor genes p53 The p53 protein normally prevents the propagation of DNA damage by causing cell arrest at the G1/S checkpoint and inducing apoptosis. Mutations in p53 remove this regulatory control, allowing mutation-carrying cells to progress through the cell cycle. An intact p53 regulatory mechanism is thought to be necessary for the activity of DNA-damaging cytotoxic agents. Rb The retinoblastoma tumor suppressor gene (Rb) encodes a protein (pRb) that acts downstream of p53, also causing cell arrest at the G1/S checkpoint. CCND1 The CCND1 gene encodes cyclin D1 protein, which inhibits the activity of pRb, stimulating its phosphorylation by the enzyme cyclin-dependent kinase 4 (CDK4). Overexpression of cyclin D1 disrupts the normal cell-cycle regulatory role of pRb. p16 The p16 gene regulates pRb function by inhibiting CDK4 enzyme activity. Inactivation of this gene therefore has the downstream effect of disrupting pRb-mediated cell-cycle control.

Comments

Therapeutic Agent(s) Targeting Genetic Alterations

Approximately 50% of NSCLC patients carry mutations in the p53 gene (Wistuba II, 2001).

Gene therapy (e.g., INGN-201, ONYX-015)

Approximately 15–40% of NSCLC cases carry mutations that disrupt normal pRb cell-cycle control (Roland M, 1998).

Gene therapy

Approximately 15% of patients overexpress the CCND1 protein; evidence suggests this overexpression may be an early premalignant mutation.

—

Mutated in 50% of NSCLC cases (Roland M, 1998). Along with deletions on chromosome 3, mutations of p16 are among the earliest mutations found in NSCLC.

—

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TABLE 1. (continued) Gene/ Protein FHIT

LRP-DIT (Lipoprotein receptor-related protein-deleted in tumors) BAX RASSFIA

PTEN/ MMAC1

Function The FHIT gene is a candidate tumor suppressor gene located at chromosome 3p14.2. FHIT encodes the enzyme dinucleotide hydrolase. Decreased function or loss of this enzyme is thought to result in accumulation of diadenosine tetraphosphate, which promotes DNA replication. A candidate tumor suppressor gene that mediates endocytosis of multiple proteins from the cell surface. LRP-DIT plays an important role in tumorigenesis. A protein that antagonizes the action of bcl-2 and promotes apoptosis. RASSFIA is a putative tumor suppressor gene on 3p21.3 and exhibits loss of heterozygosity in most lung cancers.

Phosphatase and tensin homologue deleted in chromosome 10 (PTEN), also called mutated in multiple advanced cancers (MMAC1), is a new TSG that has been located on chromosome 10q23.3 (Li J, 1997). PTEN phosphatase negatively regulates cell interactions with the extracellular matrix (Tamura M, 1998).

Comments

Therapeutic Agent(s) Targeting Genetic Alterations

Loss of protein expression occurs in 73% of NSCLC and is associated with smoking (Sozzi G, 1998).

—

Data suggest that LRP-DIT is inactivated in at least 40% of NSCLC lines (Liu CX, 2000).

—

—

—

Researchers have found hypermethylation of RASSFIA in approximately 40% of NSCLC (Wistuba II, 2001). RASSFIA, however, appears to be a key TSG for SCLC rather than NSCLC because it is inactivated by hypermethylation in more than 90% of SCLC cases. Loss of heterozygosity has been found in 41% of NSCLC cases (Virmani AK, 1998). Mutations of the PTEN/MMAC1 gene were found in 11% of all lung cancers, indicating that the gene may play a role in the pathogenesis of a small subset of lung cancers (Forgacs E, 1998).

—

—

TABLE 1. (continued) Gene/ Protein Oncogenes K-ras

Epidermal growth factor receptor (erb-B1) HER-2/neu (erb-B2)

Myc Fos

bcl-2

Therapeutic Agent(s) Targeting Genetic Alterations

Function

Comments

The ras family of genes encodes for the signal transduction proteins involved in transmitting growth-promoting signals. When mutated to the permanently active form, K-ras activates an intracellular cascade of enzymes that ultimately stimulates cell growth and division. EGFR is a tyrosine kinase receptor that, when bound by EGF or TGF-α, induces cell growth and division in epithelial cells.

Mutations of K-ras may be present in 30% of all adenocarcinomas but are seldom seen in other NSCLC subtypes (Van Zandwijk N, 1998). Mutations of this gene are common in many forms of cancer, including colon and pancreatic cancer (Wistuba II, 2001).

Farnesyl transferase inhibitors (e.g., SCH-66336, BMS-214662, tipifarnib)

EGFR is overexpressed in 40-80% of NSCLC.

EGFR inhibitors (e.g., gefitinib, erlotinib); monoclonal antibodies (e.g., ImClone’s cetuximab) Monoclonal antibody (e.g., trastuzumab [in patients with 2+ or 3+ overexpression of HER-2]).

Receptor tyrosine kinase that is involved with epithelial cell growth and division. Overexpression of the growth receptor on the surface of cells enhances metastatic potential. Nuclear transcription factor that activates genes that drive cell growth. Nuclear transcription factor that binds to DNA and allows transcription of growth-promoting genes. The bcl-2 (B-cell lymphoma/leukemia-2) gene encodes a protein that blocks apoptosis. Overproduction of bcl-2 allows cells to escape apoptosis.

Overexpressed in approximately 30% of NSCLC cases, with a higher preponderance in adenocarcinomas.

The myc protein is overexpressed in 8-20% of NSCLC patients (Wistuba II, 2001). The fos and fos-related genes are detected in approximately 50% of NSCLC cases (Van Zandwijk N, 1998). Present in 25–35% of squamous cell carcinoma cases and 10% of adenocarcinoma cases.

— —

Genasense (bcl-2 antisense oligonucleotide)

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8

TABLE 1. (continued) Gene/ Protein Survivin

Function

Comments

An apoptosis inhibitor that is expressed during fetal development but is normally undetectable in adulthood.

PCR of 83 NSCLC samples showed that 86% of tumors expressed survivin mRNA, compared with only 12% of normal tissue samples (Monzo M, 1999[a]).

Anti-survivin rabbit polyclonal antibody (available only for investigation; not approved for use in humans or clinical diagnosis).

—

—

Metastasis modulators Fucosyl-transferase Enzymes involved in the synthesis of the sialyl (FUC-T) II, IV, and Lewis-x antigen on the surface of tumor cells VII (associated with metastasis). KAI1/CD28 Transmembrane protein thought to modulate tumor progression and metastasis. Tumor resistors Beta-tubulin mutations

Multiple drug resistance (MDR)-1

Therapeutic Agent(s) Targeting Genetic Alterations

Beta-tubulin is a substrate for the formation of microtubules, one of the principal components of the cellular cytoskeleton and the mitosis spindle (the apparatus responsible for separation and organization of DNA during mitosis). The taxane class of chemotherapy agents targets tubulin as its main mechanism of action. Beta-tubulin mutations are thought to confer resistance to the taxanes. The MDR-1 gene encodes a glycoprotein drug-efflux pump (PGP) that is expressed on the surface of tumor cells and is normally present on epithelia of the renal and biliary tract. Expression of PGP allows tumor cells to expel chemotherapeutic agents and detoxify the intracellular environment.

Approximately 23% of NSCLC patients express KAI1/CD28.

—

In a study of 49 biopsy specimens from stage III and IV NSCLC patients, 33% had beta-tubulin mutations. None of the patients with the mutations responded to paclitaxel chemotherapy, whereas 39% of patients without beta-tubulin mutations responded to therapy. Median survival was three months for patients with the mutations and ten months for the controls (Monzo M, 1999[b]).

Taxanes (patients with these mutations may be resistant to taxane therapy).

Expression of PGP is thought to play a minor role in lung cancer resistance. Approximately 15–17% of NSCLC tumors may express PGP, but the level of expression is highly variable and does not correlate with clinical drug resistance.

—

TABLE 1. (continued) Gene/ Protein

Function

Other relevant genes/events in NSCLC Cyclin B1 Cyclin B1 is a regulatory subunit of cyclin-dependent kinase 1 (cdk1). Together, cdk1 and cyclin B1 (MPF) control entry into mitosis. Activation of cdk1/cyclin B1 and entry into mitosis require phosphorylation by cdk7, the catalytic subunit of cdk-activating-kinase CAK. E-cadherin (E-cad) The presence of E-cad and all catenins is and its essential for full adhesive function of the cell, associated and catenins α, β, and γ are often intracellular expressed in NSCLC, and significantly more molecules, in adenocarcinomas than in squamous cell catenins α, β, or large-cell carcinomas (Pirinen RT, 2001). and γ

Retinoic acid receptor beta (RAR-beta)

Retinoids are essential to the growth and differentiation of normal lung epithelial tissues. They interact with the retinoic acid receptors [RARs] and retinoid X receptors [RXRs], both of which are involved in regulating transcription of specific genes that, in turn, regulate cell differentiation, proliferation, and loss.

Comments Overexpression of cyclin B1 is more frequently observed in squamous cell tumors than in tumors of other histological cell types. Patients with high expression of cyclin B1 had a significantly shorter survival time than patients whose tumor expressed low levels of cyclin B1 (Soria JC, 2000). Tumor samples from 193 patients with stages I-III NSCLC showed absent or severely reduced membranous expression of E-cad, α, β, and γ catenins in 10%, 17%, 8%, and 31% of cases, respectively. This reduced expression was associated with tumor differentiation, invasiveness, metastasis, and reduced survival in NSCLC patients (Bremnes RM, 2002). RAR-beta mRNA is undetectable in 50% of NSCLCs (Khuri FR, 2000). Overall survival is significantly worse in patients with strongly positive RAR-beta expression compared with patients with weak or absent RAR-beta expression (Khuri FR, 2000).

Therapeutic Agent(s) Targeting Genetic Alterations —

—

—

9

10 TABLE 1. (continued) Gene/ Protein TNF-α/TGF-β

Function

Genes encoding deoxy-cytidine kinase (dCK) and 5-nucleotidase (5
-NT)

TGF-β plays an important role in controlling growth. TGF-β inhibits cell proliferation by repressing c-myc expression. It also inhibits the activity of cyclin-dependent kinases (CDKs), which leads to pRb hypophosphorylation and the arrest of the cell cycle. TNF-α is a cytokine that influences various types of cells. Nucleoside analogues, including gemcitabine, are phosphorylated to their active compounds by cellular dCK, while metabolites are inactivated by dephosphorylation through 5’-NT.

Matrix metalloproteinase (MMP)-9

An endopeptidase that degrades basement membrane type IV collagen and plays an important role in the angiogenic process.

Vascular endothelial growth factor (VEGF)

VEGF is essential to the regulation of endothelial cell proliferation and angiogenesis, both key contributors to the growth of cancer and vascular diseases.

Comments

Therapeutic Agent(s) Targeting Genetic Alterations

TNF-α and TGF-β mRNA are expressed in approximately 55% and 45% of tumors, respectively (Boldrini L, 2000).

dCK and 5
-NT were detected in seven of ten and eight of ten NSCLC samples respectively. The pattern of expression may have relevance in the chemotherapeutic activity of nucleoside analogues (Danesi R, 2001). Pretreatment serum levels of MMP-9 analyzed with enzyme-linked immunoassay in 118 NSCLC patients showed that MMP-9 was significantly associated with short survival time (Laack E, 2001). VEGF was found to significantly influence overall survival and recurrence-free survival in NSCLC. A significantly higher expression of VEGF in serum is observed in patients with advanced disease than in patients with early-stage disease (Gregorc V, 2001).

—

Gemcitabine

Matrix metalloproteinase inhibitors (e.g., Neovastat, BMS-275291)

VEGF inhibitors (rhuMab-VEGF)

TABLE 1. (continued) Gene/ Protein NQOR (NAD(P)H: quinone oxido-reductase)

Cyclo-oxygenase-2 (COX-2)

Telomerase

Death-associated protein kinase (DAPK)

Collapsin response mediator protein 1 (CRMP1)

Function

Comments

A flavoprotein that catalyzes the two-electron reduction of quinones and their derivatives. Quinones are present in cigarette smoke and are implicated in the pathogenesis of lung cancer. An enzyme that synthesizes prostaglandins. Tumors with up-regulation of COX-2 synthesize high levels of prostaglandin E2 (PGE2), a potent signaling molecule. An enzyme that extends the ends of telomeres that have been shortened with successive cell divisions. DAPK is a calmodulin-dependent serine/threonine kinase that contains a death domain and functions as a positive mediator of apoptosis induced by interferon γ . Recent research suggests that DAP-kinase may be important in the progression of NSCLC from early- to late-stage disease. CRMP1 is also a novel gene associated with microtubule-containing structures. Although the precise role of CRMP1 is not yet clear, it appears to be involved in cancer invasion and metastasis and may be an invasion-suppressor gene.

NQOR is overexpressed and is more frequently mutated in tumor samples than in normal lung tissue (Kolesar JM, 2000).

High PGE2 levels are associated with increased production of proangiogenic factors and enhanced metastatic potential. A high proportion (82%) of lung tumors showed positive telomerase activity ´ (Gonzalez-Quevedo R, 2002). Researchers have found methylation of DAPK in approximately 47 (25%) of 185 NSCLC tumors (Kim D-H, 2001). Methylation of DAPK was significantly associated with an advanced stage, increased tumor size, lymph node involvement, and distant metastasis (Kim D-H, 2001). Down-regulation has been linked to metastasis and shorter survival in NSCLC (Monzo M, 2002).

Therapeutic Agent(s) Targeting Genetic Alterations —

COX-2 inhibitors

—

—

—

11

DNA = Deoxyribonucleic acid; EGFR = Epidermal growth factor receptor; FHIT = Fragile histidine triad; mRNA = Messenger ribonucleic acid; NSCLC = Non-small-cell lung cancer; PCR = Polymerase chain reaction; SCLC = Small-cell lung cancer; TGF = Transforming growth factor; TSG = Tumor suppressor gene. Full source citations appear in ‘‘References.’’

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HER-4/erb-B4—that are composed of an extracellular ligand-binding domain, a transmembrane domain, and an intracellular tyrosine kinase domain (Pao W, 2005; Sridhar SS, 2003). These receptors are widely expressed on the cell surface and are implicated in the development and progression of epithelial tumors. Binding to the receptor results in tyrosine kinase activation and receptor autophosphorylation, which then initiate signal transduction cascades implicated in cell proliferation. Overexpression of EGFR correlates with a worse prognosis in several cancers, including NSCLC; EGFR is overexpressed in 40–80% of NSCLC (Sridhar SS, 2003). Researchers have found that speciÞc mutations in the tyrosine kinase domain of the EGFR gene strongly correlate with sensitivity to the tyrosine kinase inhibitors geÞtinib (AstraZeneca’s Iressa) and erlotinib (OSI/Genentech/ Roche’s Tarceva) (Pao W, 2005; Lynch TJ, 2004[a]). Japanese patients have a signiÞcantly higher response rate to both geÞtinib and erlotinib than non-Japanese patients. SigniÞcantly higher response rates have also been observed in patients with good performance status, patients with the adenocarcinoma histology subtype, patients who never smoked, and female patients (Pao W, 2005; Lynch TJ, 2004[a], Paez JG, 2004). Therefore, screening for such mutations may identify patients who might respond to geÞtinib or erlotinib. However, the incidence of mutations reported so far is low (12.5%), and does not account for all patients who beneÞt from either geÞtinib or erlotinib trials. Other factors that predict response to treatment have yet to be identiÞed (Dowell JE, 2004). Telomerase. Telomerase is an enzyme that increases the length of chromosome ends (telomeres); its activation probably plays a crucial role in tumorigenesis. Telomerase activity is suppressed in most normal cells but is expressed in many tumor cells. Researchers hypothesize that the telomere is shortened after each cell division and that this shortening eventually leads to cell death. When activated, telomerase prevents or reverses telomere shortening, an action that probably contributes to the immortalization of cancerous cells (Soria JC, 2001; William RN, 2000). Immortalization may also require other genetic alterations that stop normal differentiation, such as mutations in the TSG p16 (Farwell DG, 2000). More than 80% of NSCLC patients express telomerase (Gonzalez-Quevedo R, 2002), and a high level of telomerase activity in NSCLC is associated with increased cell proliferation rates and advanced pathological stage (Albanell J, 1997). Recent evidence suggests that telomerase activity occurs frequently in smokers, that this activity appears at a very early stage in lung cancer caused by smoking, and that its expression can be modulated by treatment with a retinoid (Soria JC, 2001). The detection of telomerase could potentially help to identify NSCLC patients with unfavorable pathological features. Research has shown that the effect of elevated telomerase activity may depend on the level of p16 expression. Patients with elevated telomerase activity but normal p16 expression have a good prognosis; patients who have elevated telomerase activity and loss of p16 expression have a poor prognosis (Gonz´alez-Quevedo R, 2002).

ETIOLOGY AND PATHOPHYSIOLOGY

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FIGURE 1. Contribution of risk factors to incidence of non-small-cell lung cancer in the major pharmaceutical markets.

Risk Factors for Non-Small-Cell Lung Cancer. Most known cancer risk factors can be categorized as processes that increase a person’s exposure to mutagens, interfere with or prevent normal DNA repair, and/or increase the proliferation rate of cells (e.g., as a result of repetitive injury and tissue repair). As shown in Figure 1, the chief known risk factors for NSCLC are related to mutagen or carcinogen exposure. Cigarette Smoking. Cigarette smoking is by far the most important risk factor for NSCLC: as stated earlier, 85% of all lung cancer cases are attributable to this habit (Reddy AA, 2000). Smoking can contribute to cancer development by impairing mucociliary clearance in the lungs, by decreasing immunological response, or by causing mutations in TSGs. Tobacco smoke is composed of two elements: the vapor phase and the particulate phase. The particulate phase, known as “tar,” contains most of the 55 known cigarette smoke carcinogens. These carcinogens require activation by carcinogen-metabolizing enzymes, principally cytochrome P450 enzymes. Approximately 20% of smokers develop lung cancer, and smokers are 13 times more likely to develop lung cancer than nonsmokers (Fauci AS, 1998). The major subtypes of lung cancer most closely linked to smoking are squamous cell and small-cell lung carcinomas (Barbone F, 1997; Khuder SA, 1998). In addition, large numbers of nonsmokers are at high risk of lung cancer via exposure to environmental tobacco smoke (ETS)—i.e., passive smoking (Jarvis MJ, 2001). The tobacco industry challenged this Þnding in an industry-funded

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study that concluded that ETS may not affect lung cancer risk. In May 2003, however, the American Cancer Society (ACS) condemned the tobacco industry study for inaccurate use of data and said that the study’s researchers used a ßawed methodology to reach this conclusion. Researchers from the International Agency for Research on Cancer conÞrm that ETS is a risk factor for lung cancer and other respiratory diseases, especially in ex-smokers (Vineis P, 2005). A Hong Kong study also found signiÞcant dose-dependent associations between passive smoking and mortality from lung cancer and other respiratory diseases (McGhee SM, 2005). Currently available data clearly show that ETS is associated with an increased risk of lung cancer; however, the association of ETS exposure with certain genetic factors is not so clear. R.J. Hung and colleagues found that the variant of CYP1A1 (a P450 gene) might play a role in lung carcinogenesis in Caucasian nonsmokers, possibly in combination with the GSTM1 (glutathione S-transferase) null genotype (Hung RJ, 2003). C. Kiyohara and colleagues found that an increased—but not signiÞcant—risk is associated with the GSTM1 null genotype in Japanese nonsmokers (Kiyohara C, 2003). Recent research shows that nonsmokers with the GSTM1 null genotype who had been exposed to 20 years or more of household ETS had a 2.3-fold increased risk of developing lung cancer (Wenzlaff AS, 2005). Furthermore, people in this high-risk group who had both the GSTM1 null and GSTP1 (also glutathione S-transferase) Val allele were at a more than four-fold increased risk of lung cancer (Wenzlaff AS, 2005). These recent Þndings conÞrm that in nonsmokers, the GSTM1 genotype, alone or in combination with GSTP1 genotype, in the presence of EST alters the risk of developing lung cancer. As the number of cigarettes smoked and the time spent smoking per day increases, lung cancer risk rises sharply. Also, studies have found that risk of lung cancer is related to the type of cigarette smoked: low-tar or Þltered cigarettes are associated with lower risk than high-tar or unÞltered cigarettes (Blot WJ, 1996). A more recent report from the National Cancer Institute (NCI) found no difference, however (NCI, 2003). Smokers who switch from regular cigarettes to low-tar or light cigarettes compensate for the lower nicotine level by inhaling more deeply; by taking larger, more rapid, or more frequent puffs; or by increasing the number of cigarettes smoked per day. Consequently, these smokers cancel out any potential beneÞt of smoking a “low-tar” cigarette. The report concluded that quitting is the only proven way to reduce the disease risks associated with smoking. Prior Nonmalignant Lung Disease. Lung cancer risk is high in people with a history of prior nonmalignant lung disease, most notably asthma, pneumonia, emphysema, and tuberculosis (Alavanja MC, 1992). Exposure to Carcinogens. Although signiÞcant progress has been made in recent years in elucidating the mechanisms by which certain tobacco carcinogens cause genetic mutations, the relationship between speciÞc carcinogens and speciÞc TSG or oncogene mutations remains speculative. The following chemicals are known or suspected carcinogens:

ETIOLOGY AND PATHOPHYSIOLOGY

•

• •

15

Benzopyrene, a known carcinogen found in tobacco smoke, is the subject of numerous studies and is thought to cause mutations to the p53 gene. (As noted earlier, p53 is a key TSG that controls tumor growth by triggering apoptosis in abnormal cells.) NNK, a nitrosamine, has been linked with mutations in the p53 and K-ras genes. Researchers have suggested that aßatoxins cause lung cancer. Aßatoxins are produced by fungi that invade agricultural commodities (such as tobacco) when these plants are stored in warm, damp conditions after harvesting. These products subsequently cause mutations in the K-ras and p53 genes (Lane KS, 1999). The presence of aßatoxins in foodstuffs such as peanuts, corn, and grains has been monitored by the FDA since 1966, but their contamination of tobacco is not regulated.

Once metabolized, carcinogens interact with DNA. If the by-products of this interaction escape the efforts of cellular repair mechanisms, they may cause a permanent mutation. Errors that occur in certain regions of an oncogene or TSG may ultimately result in cancer. Dietary Factors. Researchers have found that the risk of lung cancer is high in people with a high intake of food rich in fat and cholesterol, including whole milk and eggs (Jain M, 1990; Shekelle RB, 1991). Evidence obtained in North and South America also suggest that consumption of alcoholic beverages, mostly beer, may be associated with increased risk of developing lung cancer (Bandera EV, 1992; Potter JD, 1992). In the Bandera study, however, the association was limited to heavy smokers. Researchers later found that both alcohol consumption and tobacco use are associated with p53 mutations in NSCLC, which suggests that alcohol may increase the mutagenic effects of cigarette smoke in the lung (Ahrendt SA, 2000). Radon Exposure. An inert gas produced by the decay of radium-226, radon222 is naturally present in rock and soil. As radon-222 decays, it emits highenergy alpha particles. If inhaled, these particles can disrupt DNA in the cells of the bronchial lining. Most evidence that correlates radon with lung cancer pertains to mining environments, where levels of the gas are relatively high. In a 1995 meta-analysis of 11 studies, the NCI determined that 40% of lung cancer deaths of uranium miners were caused by radon exposure. The NCI estimates that residential radon exposure could account for 2–4% of lung cancer deaths. The combination of smoking and radon increases the risk of lung cancer to an extent that ranges between additive and multiplicative; U.S. and U.K. research indicates that 50–75% of radon-linked lung cancer mortality is attributable to the excess risk posed by smoking. Air Pollution. Because lung cancer incidence and mortality rates are highest in urban areas, researchers suspect that high levels of air pollution are a risk factor

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NON-SMALL-CELL LUNG CANCER

for lung cancer. The excess relative risks are generally on the order of 50% or lower (Pershagen G, 1990). Genetic Predisposition. Although the majority of lung cancer cases are due to cigarette smoking, epidemiological studies suggest that inherited predisposition may play an important role in lung cancer susceptibility—lung cancer does occur in nonsmokers, and Þrst-degree relatives of smokers have an increased risk of developing lung cancer at an early age (Forgacs E, 2001). Researchers remain unsure of the role played by genetic factors in the pathogenesis of lung cancer. Several human genes have been extensively studied as risk factors for development of lung cancer; because of the complexity of carcinogen metabolism, however, single genotyping approaches have borne-limited fruit. Some studies have examined multiple genotypes and found increased risk associated with variants of CYP1A1 in combination with GSTM1 genotypes. Additional studies are required to elucidate this putative connection. Examining the phenotypes of carcinogen metabolites may prove to be a more productive research approach than examining all genes that can potentially react with carcinogens because the metabolites provide a more speciÞc trail to relevant genes. Other Risk Factors. Lung cancer is one of the most common cancers affecting both men and women in the United States, and its incidence is growing throughout the world. Lung cancer rates rise progressively with age. Lung cancer is also one of the major effects of exposure to high doses of ionizing radiation. Because studies have found a signiÞcant incidence of lung cancer in patients receiving radiation therapy, especially smokers, it is clear that smoking and radiation have a combined effect on lung cancer. However, the precise manner in which these two factors interact is less clear. Risk Reduction Factors for Non-Small-Cell Lung Cancer. A diet that is high in fruits and vegetables may decrease the risk of developing lung cancer by imparting high levels of antioxidants such as vitamins C and E, carotenoids, and selenium. Such antioxidants are potent scavengers of DNA-damaging free radicals and may impede the effect of carcinogens. Researchers have found that patients exhibiting a speciÞc polymorphism in the promoter region of myeloperoxidase (MPO, an enzyme that generates reactive oxygen species and free radicals that damage DNA) have lower levels of MPO expression and a reduced risk of lung cancer (Kantarci OH, 2002). The greatest reduction in risk, however, comes from cessation of smoking, which results in signiÞcant decrease in all lung cancer types. A study of the effect of smoking cessation found that risk reduction was highest for small-cell lung carcinoma, followed by squamous cell carcinoma, and lowest for large-cell cancer and adenocarcinoma (Khuder SA, 2001). Heavy smokers, particularly women, beneÞt most from quitting. The percentage reduction in risk after quitting depends on the duration of time smoked: shorter-term and younger smokers experience larger relative decreases in risk. However, the chance that risk of lung cancers in former smokers will revert to the level of nonsmokers is small (Blot WJ, 1996).

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FIGURE 2. Natural history of non-small-cell lung cancer.

Pathophysiology In general, researchers accept the hypothesis that lung cancer develops over an extended time period, ranging from 20 to 30 years. Prior to World War I, when cigarette smoking rates were low, lung cancer was a relatively rare malignancy. By studying pathological specimens from patients who smoked for different durations of time, investigators have found that lung cancer develops in stages, beginning from the time that a patient starts smoking (Figure 2). These stages correlate with the accumulation of genetic mutations (Figure 3). Classification of Non-Small-Cell Lung Cancer. A variety of benign and malignant tumors can arise in the lungs. Of these tumors, 90–95% originate in the epithelial lining of the bronchial airways and are called bronchogenic carcinomas. (The remaining 5–10% of tumors, which originate in the outer lining of the lung [the pleura], are not discussed here.) Pathologists further classify bronchogenic

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NON-SMALL-CELL LUNG CANCER

TABLE 2. Lung Cancer Subtype Distribution by Gender and Geographic Region (Percentage) United States Subtype Squamous cell Adenocarcinoma Large-cell Small-cell Other/unspecified

Europe

Japan

Males

Females

Males

Females

Males

Females

31 31 10 17 11

19 38 10 21 12

44 17 7 15 17

24 29 8 17 22

34 34 7 16 9

16 55 4 12 13

FIGURE 3. Timing of genetic changes found in non-small-cell lung cancer.

cancers as small-cell or non-small-cell lung cancers based on the microscopic appearance of the tumor cells. The fact that approximately 10% of all NSCLC cases involve two or more of the histological subtypes has led some researchers to suggest that all three subtypes may arise from a common pluripotent stem cell in the bronchial epithelium that can differentiate along multiple pathways. These histological subtypes classically present in different regions of the lung, and their natural disease courses differ to some degree (Figure 4). In addition, the distribution of histological subtypes varies by gender and geographical region (Table 2). In the past 20 to 30 years, the histological distribution of NSCLC has shifted: the rate of adenocarcinoma has been increasing (especially in females), while the squamous cell subtype has generally stabilized or decreased. Epidemiologists and researchers attribute this shift both to improved ability to diagnose peripheral tumors (whose predominant histology is adenocarcinoma) and to changes in cigarette type and tar content.

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19

FIGURE 4. Anatomy and histology of non-small-cell lung cancer.

Staging. NSCLC is staged according to the internationally recognized primary tumor, regional nodes, and distant metastasis (TNM) classiÞcation system developed by the American Joint Committee on Cancer (AJCC) and the International Union Against Cancer (UICC) (Tables 3 and 4). Symptoms of Lung Cancer. The most common symptoms of NSCLC are a persistent cough and wheezing. Because of the nonspeciÞc nature of these symptoms, diagnosis of NSCLC is often delayed while other, more common diagnoses are investigated. The signs and severity of symptoms depend on the location of the tumor, the extent of locoregional spread, and the site of metastatic disease. Hypertrophic pulmonary osteoarthropathy (HPO) occurs frequently in NSCLC patients. The symptoms of HPO are bone and joint pain and digital clubbing (fattening of the ends of the Þngers and toes), but all symptoms are not always present. Many asymptomatic patients are diagnosed incidentally following a chest X-ray for an unrelated condition.

20

NON-SMALL-CELL LUNG CANCER

TABLE 3. TNM Classification System for Non-Small-Cell Lung Cancer Tumor (T) T0 No evidence of primary tumor. T1 Tumor is 3 cm or less in greatest dimension, surrounded by lung tissue, and without evidence of invasion to the main bronchus. T2 Tumor has any of the following features: Greater than 3 cm in greatest dimension. Involves the main bronchus but separated by 2 cm or more from the carina (confluence of the left and right main-stem bronchi). Invades the visceral pleura. Associated with obstructive pneumonitis or partial atelectasis (collapse) of lung tissue. T3 Tumor has any of the following features: Tumor of any size that directly invades the chest wall, diaphragm, mediastinal pleura, or pericardium (without involving the heart or great vessels). Tumor in the main bronchus less than 2 cm from the carina. Associated with obstructive pneumonitis or atelectasis of the entire lung. T4 A tumor of any size that invades critical structures in the mediastinum such as the heart, great vessels, trachea, esophagus, vertebral body, or carina. Presence of a malignant pleural effusion (collection of fluid in the pleural space with evidence of malignant cells). Nodal involvement (N) N0 No regional lymph node metastasis. N1 Metastasis or direct extension into lymph nodes in the ipsilateral (same side as the primary tumor), peribronchial, or hilar region, or both. N2 Metastasis to the lymph nodes in the ipsilateral mediastinum or subcarinal region. N3 Metastasis to the contralateral lymph nodes. Metastasis (M) M0 No distant metastasis. M1 Distant metastasis present. Note: This system was developed by the American Joint Committee on Cancer (AJCC) and the International Union Against Cancer (UICC).

TABLE 4. Stage Grouping for Non-Small-Cell Lung Cancer AJCC/UICC Stage

Definition

IA

T1, N0, M0

IB

T2, N0, M0

IIA

T1, N1, M0

IIB

T2, N1, M0 T3, N0, M0

IIIA

T3, N1, M0 T1-3, N2, M0

IIIB

T4, any N, M0 Any T, N3, M0

IV

Any T, any N, M1

AJCC = American Joint Committee on Cancer. UICC = International Union Against Cancer.

CURRENT THERAPIES

21

Less frequent symptoms of localized NSCLC include blood in the sputum (20–40% of cases), a consequence of tissue necroses, and pneumonia (up to 25% of cases). Patients ultimately experience chest pain if the tumor spreads locally into the chest wall. Common symptoms of metastatic disease are anorexia-cachexia (a lack of appetite and physical wasting), generalized fatigue, and weakness. In addition, patients experience symptoms related to the site of metastases—for example, patients with brain metastases experience headaches, seizures, and cognitive effects. Prognostic Factors in Non-Small-Cell Lung Cancer. Table 5 presents established and investigational prognostic factors in NSCLC. Table 6 discusses the prognostic signiÞcance of select genetic mutations seen in NSCLC.

CURRENT THERAPIES Researchers once believed that the use of cytotoxic agents provided little beneÞt in treatment of non-small-cell lung cancer (NSCLC), but positive clinical trial data have ensured that chemotherapeutic regimens are now well established, especially in late-stage disease. Most treatment strategies are platinum-based, and newer, dual-combination regimens containing third-generation agents are becoming more popular than older, triple-combination regimens. A growing body of clinical evidence is expanding the population of patients who are treated with chemotherapy. For example, evidence now exists that carboplatin-based regimens, with their favorable toxicity proÞle, can beneÞt elderly patients and patients with a poor Eastern Cooperative Oncology Group (ECOG) performance status (PS) (PS > 2). Perhaps the most striking trend, however, is in the treatment of less advanced disease. Resection is still the standard of care in early-stage or localized disease (tumor, node, metastasis [TNM] stages I, II, and IIIA), but impressive data from recent trials support the use of chemotherapy after resection (adjuvant chemotherapy) to increase the patient’s chance of continued remission and to delay relapse (Le Chevalier T, 2003; Strauss GM, 2004; Winton TL, 2004), or before surgery (neoadjuvant chemotherapy), where the aim is to downstage the tumor and increase the likelihood of surgical success (Choi IS, 2004; DePierre A, 2002). Treatment options in NSCLC continue to expand. Three agents from two new drug classes have been licensed: two epidermal growth factor receptor (EGFR) inhibitors, geÞtinib (AstraZeneca’s Iressa) and erlotinib (OSI/Genentech/Roche’s Tarceva), and the Þrst antifolate to be approved for NSCLC, pemetrexed (Eli Lilly’s Alimta). These agents are currently approved as monotherapy for patients who have failed conventional therapy; they remain in development, in combination with other agents, for Þrst-line therapy. Table 7 outlines the chemotherapy regimens most commonly used to treat NSCLC; these regimens are discussed in the following sections.

22

NON-SMALL-CELL LUNG CANCER

TABLE 5. Prognostic Factors in Non-Small-Cell Lung Cancer Established prognostic factors Karnofsky performance status Disease stage (TNM) History of weight loss within the previous six months Investigational prognostic factors Anatomical and pathological markers: Tumor size Histological subtype Gender (females fare better) Serum albumin level Lactate dehydrogenase (LDH) level in serum Degree of tumor differentiation Degree of inflammatory response Blood vessel invasion Lymphatic vessel invasion Mitotic index Presence/absence of tumor giant cells Molecular/genetic markers: K-ras erb-B2 (HER-2/neu) p53 Rb protein bcl-2 protein Carcinoembryonic antigen (CEA) Cyclin D1 (CCND1) MRP-1/CD9 KAI1/CD82 CYFRA 21-1 Tissue polypeptide-specific antigen (TPS) Squamous cell carcinoma antigen (SCC) Growth factors Vascular endothelial growth factor (VEGF) Platelet-derived endothelial cell growth factor (PD-ECGF) Differentiation markers: Tumor expression of blood-group antigens Lewis-group antigens Proliferation markers: DNA ploidy S-phase fraction Thymidine labeling index Nucleolar organizing regions Proliferating cell nuclear antigens Markers of metastatic propensity: Angiogenesis microvessel density Basement membrane deposition

TABLE 6. Prognostic Significance of Select Genetic Mutations in Non-Small-Cell Lung Cancer Gene/ Protein

Prognostic Significance

Tumor suppressor genes p53

Adverse

Rb

Adverse

p16

Adverse

Fragile histidine triad (FHIT)

Adverse

BAX

Adverse (with absence of bcl-2 overexpression)

Oncogenes K-ras

Adverse

HER-2/neu (erb-B2)

Adverse

Discussion Tumors with negative p53 expression are associated with high metastatic potential in stage III NSCLC (Fokkema E, 1999). The demonstration of p53 mutations or accumulation is associated with shorter survival periods in certain subgroups of NSCLC. Also, p53-overexpressing tumors are likely to be resistant to cisplatin therapy (Taron M, 2001). Altered Rb protein expression is an independent prognostic factor for reduced overall survival in early-stage NSCLC. An even poorer prognosis is observed when loss of Rb protein function and p53 protein mutations occur in the same tumor (Xu HJ, 1994). In stages I and II adenocarcinomas, cases with Rb-negative tumors had a significantly shorter survival than cases with Rb-positive tumors (Brambilla E, 1999). A strong association is observed between the reduction or loss of p16 expression and tumor recurrence (Gonzalez-Quevedo R, 2002). A strong association is found between loss of the protein expression and cigarette smoking as well as metaplastic changes in NSCLC (Tseng JE, 1999). The replacement of the protein in cancer cells suppresses tumorigenicity in nude mice (Siprashvili Z, 1997). Multivariate analysis shows that the presence of bax+/bcl-2-tumor expression has a significant negative influence on survival in stage I NSCLC patients. Bax expression alone has no influence on survival of stage I patients, but patients with bax+/bcl-2-tumors have the worst prognosis (Apolinario RM, 1997).

23

The relapse rate is higher in NSCLC patients with codon-12 mutations of the K-ras gene. The presence of K-ras mutations is associated with shorter survival time (Nasioulas G, 2001). Overexpression of the growth factor receptor on the surface of cells enhances metastatic potential. Together with erbB-2 overexpression, the immediate presence of K-ras mutation on codon 12 could be an additional negative prognostic effect (Kern JA, 1994).

24 TABLE 6. (continued) Gene/ Protein

Prognostic Significance

Epidermal growth factor-receptor (erb-B1)

Adverse

bcl-2

Controversial

Survivin (a novel apoptosis inhibitor)

Adverse

Discussion Specific mutations in the tyrosine kinase domain of the EGFR gene strongly correlate with sensitivity to tyrosine kinase inhibitors (erlotinib and gefitinib) (Pao W, 2005; Lynch TJ, 2004). Studies involving stages I to IIIA NSCLC patients show that the probability of relapse-free survival at six years is higher for patients with bcl-2 positive tumors than for those with bcl-2 negative tumors (Silvestrini R, 1998). However, multivariate analysis indicates that bcl-2 protein expression is not an independent prognostic factor in patients with potentially curable NSCLC. Patients without survivin-expression had significantly better overall survival than patients with survivin-expression.

Metastasis modulators Fucosyltransferase (FUC-T) II, IV, and VII KAI1/CD28

Adverse

—

Favorable

—

Tumor resistors Beta-tubulin mutations

Adverse

Multiple drug resistance (MDR)-1

Adverse

Other relevant genes/events in NSCLC Cyclins B1, D3 Adverse

Beta-tubulin mutations in 16 of 49 tumor samples (33%) were found to be associated with poor response to paclitaxel treatment and shorter overall survival (Monzo M, 1999). Tumors with certain beta tubulin (TUBB) mutations exhibit taxane resistance (Taron M, 2001). —

A significantly shorter survival time is observed in patients with a high expression of cyclin B1 than in patients whose tumors express a low level of cyclin B1 (Soria JC, 2000). Overexpression of cyclins D3 and B1 adversely affects survival in patients with advanced NSCLC (Xynogalos S, 2002).

TABLE 6. (continued) Gene/ Protein

Prognostic Significance

E-cadherin (E-cad) and its associated intracellular molecules, catenin α, β, and γ Retinoic acid receptor beta (RAR-beta)

Adverse for early-stage NSCLC

TNF-α/TGF-β

Favorable

Matrix metalloproteinase (MMP)-9 Cyclooxygenase-2 (COX-2)

Adverse

Telomerase

Adverse

Basic fibroblast growth factor (bFGF) Vascular endothelial growth factor (VEGF) Stathmin (an oncoprotein also known as Op18) SXR (steroid and xenobiotic receptor)

Adverse

E-cad and β-catenin are independent prognostic factors for NSCLC survival.

Adverse in stage I NSCLC, favorable in adenocarcinomas

Adverse — —

Discussion Previous studies found that reduced expression of β-catenin or γ -catenin is related to poor prognosis. Recent studies found that reduced expression of E-cad and catenins are associated with tumor differentiation, invasiveness, metastasis, and reduced survival in NSCLC (Bremnes RM, 2002; Kren L, 2003). Overall survival is significantly worse in patients with strongly positive RAR-beta expression compared with patients with weak or absent RAR-beta expression (Khuri FR, 2000). In a univariate analysis, patients without nodal metastatic involvement, with less advanced tumor stage, or TNF-α and TGF-β positive cancers showed a favorable prognosis in terms of overall survival (Boldrini L, 2000). Pretreatment serum levels of MMP-9 analyzed in 118 NSCLC patients showed that MMP-9 was significantly associated with short survival (Laack E, 2001). Previous data show that strong COX-2 expression is associated with a relatively poor overall survival rate (Khuri FR, 2001[a]). More recent data show that COX-2 expression is associated with a good prognosis in adenocarcinomas (Yamaguchi NH, 2003). Researchers have found that patients with hTERT (a protein subunit of telomerase that is closely associated with telomerase activity) expression have shorter survival than those who do not express hTERT (Komiya T, 2000). Marked expression of bFGF is associated with a shorter survival period in patients with NSCLC. A number of studies have found VEGF expression to be associated with poor prognosis in NSCLC. Overexpression of stathmin enhances vinorelbine but decreases paclitaxel activity (Rosell-Costa R, 2002). Overexpression of SXR can decrease paclitaxel cytotoxicity (Rosell-Costa R, 2002).

25

26 TABLE 6. (continued) Gene/ Protein ERCC1 (excision repair cross-complementing) gene

Ribonucleotide reductase gene XPD (originally called excision repair cross-complementing group 2) Integrins (a group of transmembrane glycoproteins consisting of α and β subunits)

Prognostic Significance Adverse

— —

Adverse

Full source citations appear in ‘‘References.’’

Discussion Overexpression of the ERCC1 gene confers resistance to cisplatin and oxaliplatin therapy (Taron M, 2001; Rosell-Costa R, 2002). Preliminary data from the Italian Lung Cancer Project (ILCP) confirm the negative impact of ERCC1 overexpression on survival (Rosell-Costa R, 2002). Tumors with deletions in the region containing the ribonucleotide reductase gene are unlikely to respond to gemcitabine therapy (Taron M, 2001) XPD genotype can predict response to cisplatin chemotherapy; a higher response rate is found in the 751 Lys/Gln and Gln/Gln subgroups than in patients with the Lys/Lys genotype (Taron M, 2002). Reduced expression of integrin-α3 is associated with poor prognosis in patients with adenocarcinomas (Adachi M, 1998). In node-negative NSCLC patients, overexpression of integrin-α5 may also be associated with a negative prognostic factor (Adachi M, 2000).

CURRENT THERAPIES

27

Gemcitabine/Cisplatin Overview. The combination regimen of gemcitabine (Eli Lilly’s Gemzar) and cisplatin (Bristol-Myers Squibb’s Platinol-AQ, generics) is one of the regimens most widely used for treatment of NSCLC. Gemcitabine/cisplatin is used mainly as Þrst-line treatment for patients with inoperable, locally advanced (TNM stages IIIA and IIIB), or metastatic (TNM stage IV) NSCLC, but its popularity as neoadjuvant therapy for surgical patients is growing. Mechanism of Action. The individual components of the gemcitabine/cisplatin regimen contribute the following mechanisms to achieve its overall activity: •

•

Gemcitabine (Figure 5) is an antimetabolite. Antimetabolites act by blocking normal DNA synthesis, thus stopping cell replication. Gemcitabine exhibits cell-phase speciÞcity, primarily killing cells that are undergoing DNA synthesis (S phase) and also blocking the progression of cells through the G1/S-phase boundary. Cisplatin (Figure 6) is a platinum agent. Platinum agents inhibit tumor-cell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that cisplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle.

Clinical Performance. Results from the Japanese Four Arm Cooperative Study (FACS) of 602 Þt, NSCLC stage IIIb or IV patients (PS < 2) show that, compared with irinotecan (PÞzer’s Camptosar, Yakult/PÞzer’s Campto, Daiichi’s Topotecin)/cisplatin (the control arm), paclitaxel (Bristol-Myers Squibb’s Taxol, generics)/carboplatin (Bristol-Myers Squibb’s Paraplatin, generics), vinorelbine

NH2 N HO

O O

OH

N F F

FIGURE 5. Structure of gemcitabine.

FIGURE 6. Structure of cisplatin.

28

TABLE 7. Current Regimens Used for Non-Small-Cell Lung Cancer, 2005 Regimen Components Regimen Gemcitabine/cisplatin

Paclitaxel/cisplatin

Irinotecan/cisplatin

Agent

Availability

Dosage mg/m2 /d

Gemcitabine (Eli Lilly’s Gemzar)

US, F, G, I, S, UK, J

Gemcitabine: 1,000 over 30 minutes on days 1, 8, and 15.

Cisplatin (Bristol-Myers Squibb’s Platinol-AQ, generics) Paclitaxel (Bristol-Myers Squibb’s Taxol, generics)

US, F, G, I, S, UK, J

Cisplatin: 100 mg/m2 /d on day 1. 28-day cycle.

US, F, G, I, S, UK, J

Paclitaxel: 135 mg/m2 /d over 24 hours or 175–225 mg/m2 /d over 3 hours on day 1.

Cisplatin (Bristol-Myers Squibb’s Platinol-AQ, generics) Irinotecan (Pfizer’s Camptosar, Yakult/Pfizer’s Campto, Daiichi’s Topotecin)

US, F, G, I, S, UK, J

Cisplatin: 75–80 mg/m2 /d on day 1. 21-day cycle.

US, F, G, I, S, UK, J

Irinotecan: 60 mg/m2 /d on days 1, 8, and 15.

Cisplatin (Bristol-Myers Squibb’s Platinol-AQ, generics)

US, F, G, I, S, UK, J

Cisplatin: 80 mg/m2 /d on day 1. 28-day cycle.

Common Toxicities • Thrombocytopenia • Fatigue • Nausea • Diarrhea • Emesis • Neuropathy • Renal toxicity

• Leukopenia • Neutropenia • Anemia • Peripheral neuropathy

• Nausea • Vomiting • Neutropenia • Febrile neutropenia • Diarrhea

TABLE 7. (continued) Regimen Components Regimen Vinorelbine/cisplatin

Etoposide/cisplatin

Docetaxel/cisplatin

Agent

Availability

Dosage

29

Vorelbine (GlaxoSmithKline/ Pierre Fabre’s Navelbine, generics)

US, F, G, I, S, UK, J

Vinorelbine: 25–30 mg/m2 once weekly.

Cisplatin (Bristol-Myers Squibb’s Platinol-AQ, generics)

US, F, G, I, S, UK, J

Etoposide (Bristol-Myers Squibb’s VePesid, Pfizer/Nippon Kayaku’s Lastet, generics)

US, F, G, I, S, UK, J

Cisplatin: 100–120 mg/m2 /d on days 1 and 29, then once every 6 weeks. 28-day cycle. Etoposide: 100 mg/m2 /d as a 1-hour infusion on days 1, 2, and 3.

Cisplatin (Bristol-Myers Squibb’s Platinol-AQ, generics) Docetaxel (Sanofi-Aventis’s Taxotere)

US, F, G, I, S, UK, J

Cisplatin (Bristol-Myers Squibb’s Platinol-AQ, generics)

US, F, G, I, S, UK, J

Common Toxicities • Fatigue • Nausea • Anorexia • Vomiting • Sensory neuropathy • Constipation

• Leukopenia • Granulocytopenia • Anemia • Infection

US, F, G, I, S, UK, J

Cisplatin: 75–120 mg/m2 /d on day 1 or 2. 21-day cycle. Docetaxel: 75 mg/m2 immediately followed by cisplatin.

Cisplatin: 75 mg/m2 . Both as 1-hour intravenous infusions on day 1, repeated every 3 weeks.

• Leukopenia • Neutropenia • Asthenia • Nausea

30

TABLE 7. (continued) Regimen Components Regimen

Agent

Availability

Dosage mg/m2 /d

Common Toxicities

Gemcitabine/carboplatin

Gemcitabine (Eli Lilly’s Gemzar)

US, F, G, I, S, UK, J

Gemcitabine: 1,000 days 1 and 8.

US, F, G, I, S, UK, J

Carboplatin: AUC of 5 mg/mLa . 21-day cycle.

Paclitaxel/carboplatin

Carboplatin (Bristol-Myers Squibb’s Paraplatin, generics) Paclitaxel (Bristol-Myers Squibb’s Taxol, generics)

US, F, G, I, S, UK, J

Gemcitabine: 1,000 mg/m2 /d on days 1 and 8.

US, F, G, I, S, UK, J

Carboplatin: AUC of 5 mg/mLa . 21-day cycle.

US, F, G, I, S, UK, J

Vinorelbine: 25–30 mg/m2 once weekly.

US, F, G, I, S, UK, J

Carboplatin: AUC of 6 mg/mLa . 28-day cycle.

US, F, G, I, S, UK, J

Docetaxel: 75–80 mg/m2 as a 1-hour infusion.

US, F, G, I, S, UK, J

Carboplatin: AUC of 6 mg/mLa . 21-day cycle.

Vinorelbine/carboplatin

Docetaxel/carboplatin

Carboplatin (Bristol-Myers Squibb’s Paraplatin, generics) Vinorelbine (GlaxoSmithKline/Pierre Fabre’s Navelbine, generics) Carboplatin (Bristol-Myers Squibb’s Paraplatin, generics) Docetaxel (Sanofi-Aventis’s Taxotere)

Carboplatin (Bristol-Myers Squibb’s Paraplatin, generics)

on

• Neutropenia • Thrombocytopenia • Anemia

• Neutropenia • Thrombocytopenia • Anemia

• Granulocytopenia • Myelosuppression • Neutropenia • Nausea • Vomiting

• Leukopenia • Neutropenia • Asthenia • Pulmonary • Diarrhea

TABLE 7. (continued) Regimen Components Regimen

Agent

Availability

Dosage

Mitomycin (generics)

US, F, G, I, S, UK, J

Mitomycin: 6–8 mg/m2 /d on day 1.

US, F, G, I, S, UK, J

Ifosfamide: 3,000–5,000 mg/m2 /d on day 1.

US, F, G, I, S, UK, J

Cisplatin: 50–100 mg/m2 /d on day 2.

Nedaplatin regimens

Ifosfamide (Bristol-Myers Squibb’s Ifex, Baxter’s Holoxan/ Mitoxana, Shionogi’s Ifomide, generics) Cisplatin (Bristol-Myers Squibb’s Platinol-AQ, generics) Nedaplatin (Shionogi’s Aqupla)

J

Docetaxel, single agent

Docetaxel (Sanofi-Aventis’ Taxotere)

US, F, G, I, S, UK, J

Nedaplatin: starting dose of 50 mg/m2 on day 1, escalated by 10 mg/m2 until the maximum tolerated dose is reached. The dose of 100 mg/m2 was recommended because that is the recommended dose of single-agent nedaplatin. Combination with Irinotecan: 60 mg/m2 on days 1 and 8; cycle repeated every 3 weeks. Docetaxel: 60–100 mg/m2 as a 1-hour infusion every three weeks.

Mitomycin/ ifosfamide/ cisplatin

Common Toxicities • Nausea • Vomiting • Constipation • Alopecia • Leukopenia

• Neutropenia • Anemia • Thrombocytopenia

• Neutropenia • Febrile neutropenia • Thrombocytopenia

31

• Asthenia • Pulmonary

32 TABLE 7. (continued) Regimen Components Regimen

Agent

Availability

Dosage

Common Toxicities

Gemcitabine, single agent

Gemcitabine (Eli Lilly’s Gemzar)

US, F, G, I, S, UK, J

Gemcitabine: 1,000 mg/m2 /d as a 30-minute infusion on days 1, 8, and 15 of a 28-day cycle.

• Neutropenia • Fatigue

Vinorelbine, single agent

US, F, G, I, S, UK, J

Vinorelbine: 30 mg/m2 as a weekly infusion for 4–5 weeks.

• Neutropenia • Fatigue

Gefitinib, single agent

Vinorelbine (GlaxoSmithKline/Pierre Fabre’s Navelbine, generics) Gefitinib (AstraZeneca’s Iressa)

US, J

Gefitinib: 250 mg/day once daily; oral.

• Diarrhea (mild) • Skin rash (mild) • Pruritus (mild) • Dry skin (mild)

Erlotinib, single agent

(OSI/Genentech/Roche’s Tarceva)

US

Erlotinib: 150 mg/day once daily; oral.

• Rash • Diarrhea

Pemetrexed

Pemetrexed (Eli Lilly’s Alimta)

US, F

Pemetrexed: 500 mg/m2 over 10 minutes on day 1 of a 21-day cycle.

• Neutropenia • Febrile neutropenia • Thrombocytopenia • Anemia

a The dose of carboplatin is calculated according to the Calvert formula: the product of the target area under the plasma concentration curve (AUC) (usually 5–6 mg/mL × number of minutes) and the glomerular filtration rate (GFR) (mL/min) plus 25. The dose is typically 300–500 mg/m2 , given over a one-hour period.

US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan.

CURRENT THERAPIES

33

(GlaxoSmithKline/Pierre Fabre’s Navelbine, generics)/cisplatin, and gemcitabine/cisplatin demonstrate no signiÞcant differences in response rate (31%, 32%, 33%, and 30%, respectively). One-year survival rates were 59%, 51%, 48%, and 60%, respectively; median months of survival were 14, 12, 15, and 11, respectively (Kubota K, 2004). Although, according to the investigators, the three experimental regimens “did not demonstrate noninferiority to the control arm in this analysis,” the irinotecan/cisplatin arm demonstrated an unexpectedly high one-year survival rate that was similar to that found with the gemcitabine/cisplatin arm. Neutropenia was less common in the gemcitabine/cisplatin arm (63%) than in the other three arms: irinotecan/cisplatin (84%), paclitaxel/carboplatin (88%), and vinorelbine/cisplatin (88%). However, symptoms of nausea and diarrhea were higher in the gemcitabine/cisplatin and irinotecan/cisplatin arms than in the other study arms. A landmark trial conducted by the ECOG (ECOG 1594) compared four commonly used third-generation chemotherapy regimens: a reference regimen of paclitaxel/cisplatin and three experimental regimens: gemcitabine/cisplatin, docetaxel (SanoÞ-Aventis’s Taxotere)/cisplatin, and paclitaxel/carboplatin (Schiller JH, 2002). The trial randomized 1,207 chemotherapy-n¨aive patients with advanced stage IIIB or IV disease and good PS (2 or better). The overall response rate for all 1,155 evaluable patients was 19%; median survival was 7.9 months, and one- and two-year survival rates were 33% and 11%, respectively. No signiÞcant differences in response rate or survival were observed between the three experimental arms. Median survival times were 8.1 months for gemcitabine/cisplatin, 7.4 months for docetaxel/cisplatin, and 8.1 months for paclitaxel/carboplatin; the survival rates for these three groups at one year were 36%, 31%, and 34%, respectively; survival rates at two years were 13%, 11%, and 11%, respectively. The median time to disease progression did not differ signiÞcantly in any of the four groups. Major toxicities (grades III and IV) occurred in all groups, particularly in patients with a poor PS. None of the four treatment arms demonstrated a signiÞcant advantage over the others in terms of survival, but the paclitaxel/carboplatin arm had a lower rate of toxic effects than the other arms. Based on the results of this study, the ECOG chose the paclitaxel/carboplatin regimen as the reference regimen for future studies. The recruitment of PS 2 patients into ECOG 1594 was suspended because of a perceived unacceptably high rate of toxicity, but further analysis revealed that overall toxicity was similar in PS 0, 1, and 2 patients (Sweeney CJ, 2001). Another trial that compared attenuated regimens of paclitaxel/carboplatin and gemcitabine/cisplatin in 98 NSCLC patients (PS 2) yielded a 16% response rate for the paclitaxel/carboplatin regimen and a 25% rate for gemcitabine/cisplatin. Survival results were similar for the two regimens. Paclitaxel/carboplatin caused more grades III and IV neutropenia and neuropathy; gemcitabine/cisplatin caused more grade III thrombocytopenia, fatigue, and nausea. The study concluded that appropriately attenuated dual combination regimens are appropriate for PS 2 patients (Tester WJ, 2004).

34

NON-SMALL-CELL LUNG CANCER

FIGURE 7. Structure of paclitaxel.

Paclitaxel/Cisplatin Overview. The combination of paclitaxel (Bristol-Myers Squibb’s Taxol, generics) and cisplatin (Bristol-Myers Squibb’s Platinol-AQ, generics) is indicated for Þrst-line treatment of NSCLC patients who are not candidates for potentially curative surgery and/or radiation therapy. Paclitaxel/cisplatin has produced good responses in both chemotherapy-naive and previously treated NSCLC patients. Cisplatin is sometimes replaced by carboplatin (Bristol-Myers Squibb’s Paraplatin, generics), but results (described in the following section, “Clinical Performance”) show that the cisplatin combination is superior. Mechanism of Action. The individual components of the paclitaxel/cisplatin regimen contribute the following mechanisms to achieve its overall activity: •

•

Paclitaxel (Figure 7) is a taxane. Taxanes act by inhibiting the dynamic reorganization of microtubule networks that is essential to vital interphase and mitotic cellular functions. Taxanes exhibit a unique mechanism of action. Paclitaxel, which achieves selective toxicity against rapidly proliferating cells, is mainly active in the G2/M phase of the cell cycle. Cisplatin (Figure 6) is a platinum agent. Platinum agents inhibit tumor-cell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that cisplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle.

Clinical Performance. A landmark trial conducted by the ECOG (ECOG 1594) compared four commonly used third-generation chemotherapy regimens: a reference regimen of paclitaxel/cisplatin and experimental regimens of gemcitabine/cisplatin, docetaxel/cisplatin, and paclitaxel/carboplatin (Schiller JH, 2002). The trial randomized 1,207 chemotherapy-naive patients with advanced stage IIIB or IV disease and good PS (2 or better). The overall response rate for all 1,155 evaluable patients was 19%; median survival was 7.9 months, and oneand two-year survival rates were 33% and 11%, respectively. Researchers found no signiÞcant differences in response rate or survival between the four arms. The median survival times were 7.8 months for paclitaxel/cisplatin, 8.1 months

CURRENT THERAPIES

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for gemcitabine/cisplatin, 7.4 months for docetaxel/cisplatin, and 8.1 months for paclitaxel/carboplatin. The survival rates for these four groups at one year were 31%, 36%, 31%, and 34%, respectively, and survival rates at two years were 10%, 13%, 11%, and 11%. The median time to disease progression ranged from 3.4 months to 4.2 months and did not differ signiÞcantly between the four groups. Major toxicities were similar in all groups and were found particularly in patients with poor PS. The paclitaxel/carboplatin regimen had a lower rate of toxic effects than the other regimens. Based on the results of this study, the ECOG chose the paclitaxel/carboplatin regimen as the reference regimen for future studies. In an international, randomized Phase III study, paclitaxel/cisplatin was found to be superior to paclitaxel/carboplatin for treatment of stages IIIB and IV NSCLC (Rossell R, 2002). From April 1996 to July 1997, 618 patients were randomized to the two treatment arms. The arms were well balanced with respect to gender (83% male in both groups), age (median 58 years), PS (0–1), stage (68% IV, 32% IIIB), and histology (38% squamous cell carcinoma). The response rate was 25% in the paclitaxel/carboplatin arm and 28% in the paclitaxel/cisplatin arm. Median survival was 8.5 months in the paclitaxel/carboplatin arm and 9.8 months in the paclitaxel/cisplatin arm; the one-year survival rates were 33% and 38%, respectively. Twenty-two months of additional follow-up yielded a median survival of 8.2 months in the paclitaxel/carboplatin arm and 9.8 months in the paclitaxel/cisplatin arm; the two-year survival rates were 9% and 15%, respectively. Excluding neutropenia and thrombocytopenia (more frequent in the paclitaxel/carboplatin arm) and nausea/vomiting and nephrotoxicity (more frequent in the paclitaxel/cisplatin arm), the rate of severe toxicities was generally low and comparable in the two arms. Overall quality of life (QOL) was also similar in the two arms. This randomized trial was the Þrst such trial to compare paclitaxel/cisplatin with paclitaxel/carboplatin in treatment of advanced NSCLC. Another randomized trial provides evidence for the role of the paclitaxel/cisplatin regimen in elderly patients (Langer CJ, 2002). The Phase III trial, called ECOG 5592, compared the results of using cisplatin with either etoposide or paclitaxel in Þt patients (PS 0 or 1) aged 70 or older and in younger patients. Toxic effects, response rates, and survival rates were compared between age-groups. A total of 574 patients enrolled were evaluable; 86 patients (15%) were age 70 or older. Older patients had a higher incidence of cardiovascular and respiratory comorbidities and nonanalgesic medication use. Leukopenia and neuropsychiatric toxicity were more common in elderly men than in younger men. Elderly women lost more weight than younger women. Other toxic effects were similar in older and in younger patients. The proportions of overall response rates were similar (21.5% in the older age-group versus 23.3% in the younger group) as were median time to progression (4.37 months versus 4.30 months) and survival distribution (median survival, 9.05 months versus 8.53 months; one-year survival, 38% versus 29%; and two-year survival, 14% versus 12%). Baseline QOL and treatment-outcome indices were similar. Declines in functional well-being over time were equivalent in both age-groups.

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A recent study has demonstrated that paclitaxel/cisplatin is an effective neoadjuvant chemotherapy in stage IIIA NSCLC. Thirty-four chemotherapy-naive surgical candidates with IIIA (N2) disease received two preoperative cycles of paclitaxel (175 mg/m2 ) and cisplatin (60 mg/m2 ) every three weeks and three postoperative cycles of paclitaxel (135 mg/m2 ) and carboplatin (area under curve [AUC] 5). The response rate was 65%. Mediastinal lymph nodes were tumor-free in 21% of patients. Grades III and IV hematologic toxicities and nonhematologic toxicities were uncommon. Grade III leukopenia occurred in 4% of the cycles, and grades I and II peripheral neuropathy occurred in 21% of the patients. At a median follow-up of 19.5 months, median time to progression was 12.1 months, and median overall survival was 23.6 months (Choi IS, 2004). Irinotecan/Cisplatin Overview. In Japan, the combination regimen of irinotecan (PÞzer’s Camptosar, Yakult/PÞzer’s Campto, Daiichi’s Topotecin) and cisplatin (Bristol-Myers Squibb’s Platinol-AQ, generics) is a Þrst-line treatment for locally advanced or metastatic NSCLC. Japanese preclinical studies conÞrm that irinotecan acts synergistically with cisplatin (Kudoh S, 1993). Elsewhere, irinotecan is approved but rarely used because non-Japanese physicians are less convinced of irinotecan’s efÞcacy in NSCLC and other cancer indications. Mechanism of Action. The individual components of the irinotecan/cisplatin regimen contribute the following mechanisms to achieve its overall activity: •

•

Irinotecan (Figure 8) is a topoisomerase I inhibitor. Topoisomerase I is a nuclear enzyme that causes reversible, single-strand breaks in DNA during mitosis. Topoisomerase I inhibitors bind to the topoisomerase I-DNA complex and prevent religation of the DNA strand, resulting in cell death. The drug is believed to exert its cytotoxic effects during the S phase of the cell cycle. Cisplatin (Figure 6) is a platinum agent. Platinum agents inhibit tumor-cell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that cisplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle.

Clinical Performance. A Phase III trial randomizing 398 patients with previously untreated, advanced NSCLC to irinotecan/cisplatin, vindesine (generics)/cisplatin, or irinotecan alone demonstrated that irinotecan/cisplatin is a valuable treatment option (Negoro S, 2003). The median survival times were 50 weeks for patients on irinotecan/cisplatin, 45.6 weeks for patients on vindesine/cisplatin, and 46 weeks for patients on irinotecan. Overall response rates were 43.7% for irinotecan/cisplatin, 31.7% for vindesine/cisplatin, and 20.5% for irinotecan. The response rate in the irinotecan/cisplatin arm was signiÞcantly higher than response rates in the other arms but did not translate to a signiÞcant difference in overall

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H3C O

O

O N

N N O N

HO H3C

O

FIGURE 8. Structure of irinotecan.

survival. Major adverse reactions were grade IV neutropenia (37% of patients on irinotecan/cisplatin, 54% on vindesine/cisplatin, and 8% on irinotecan) and grades III and IV diarrhea (12%, 3%, and 15% of patients, respectively). Based on the results of the Negoro study, the previously mentioned Japanese FACS study (see the “Gemcitabine/Cisplatin” section) was initiated to compare the irinotecan/cisplatin control arm with three newer regimens (Kubota K, 2004). This study of 602 Þt, NSCLC stage IIIB or IV patients (PS < 2) showed that, compared with irinotecan/cisplatin (the control arm), paclitaxel/carboplatin, vinorelbine/cisplatin, and gemcitabine/cisplatin demonstrate no signiÞcant differences in response rate (31%, 32%, 30%, and 33%, respectively). One-year survival rates were 59%, 51%, 48%, and 60%, respectively; median months of survival were 14, 12, 15, and 11, respectively (Kubota K, 2004). Although, according to the investigators, the three experimental regimens “did not demonstrate non-inferiority to the control arm in this analysis,” the irinotecan/cisplatin arm demonstrated an unexpectedly high one-year survival rate that was similar to that found with the gemcitabine/cisplatin arm. Neutropenia was less common in the gemcitabine/cisplatin arm (63%) than in the other three arms: irinotecan/cisplatin (84%), paclitaxel/carboplatin (88%), and vinorelbine/cisplatin (88%). However, symptoms of nausea and diarrhea were higher in the gemcitabine/cisplatin and irinotecan/cisplatin arms than in the other study arms. Vinorelbine/Cisplatin Overview. The combination regimen of vinorelbine (GlaxoSmithKline/Pierre Fabre’s Navelbine, generics) and cisplatin (Bristol-Myers Squibb’s Platinol-AQ, generics) is used as Þrst-line chemotherapy mainly in patients with advanced NSCLC (Fossella F, 2003). Mechanism of Action. The individual components of the vinorelbine/cisplatin regimen contribute the following mechanisms to achieve its overall activity:

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N

CH3 N

N H CH3O2C CH3O

CH3 N

OCOCH3 HO CO2CH3 CH3

FIGURE 9. Structure of vinorelbine.

•

•

Vinorelbine (Figure 9) is a vinca alkaloid. Vinca alkaloids act by binding with microtubular proteins of the mitotic spindle, leading to mitotic arrest or cell death. The drug achieves its selective toxicity against cancer cells by acting on proliferating cells. Cisplatin (Figure 6) is a platinum agent. Platinum agents inhibit tumor-cell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that cisplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle.

Clinical Performance. EfÞcacy data for the vinorelbine/cisplatin regimen are contradictory. The previously mentioned Japanese FACS study (see the “Gemcitabine/Cisplatin” section) was initiated to compare the irinotecan/cisplatin control arm with three newer regimens (Kubota K, 2004). This study of 602 Þt, NSCLC stage IIIB or IV patients (PS < 2) showed that, compared with irinotecan/cisplatin (the control arm), paclitaxel/carboplatin, vinorelbine/cisplatin, and gemcitabine/cisplatin demonstrate no signiÞcant differences in response rate (31%, 32%, 30%, and 33%, respectively). One-year survival rates were 59%, 51%, 48%, and 60%, respectively; median months of survival were 14, 12, 15, and 11, respectively (Kubota K, 2004). Although, according to the investigators, the three experimental regimens “did not demonstrate non-inferiority to the control arm in this analysis,” the irinotecan/cisplatin arm demonstrated an unexpectedly high one-year survival rate that was similar to that found with the gemcitabine/cisplatin arm. Neutropenia was less common in the gemcitabine/cisplatin arm (63%) than in the other three arms: irinotecan/cisplatin (84%), paclitaxel/carboplatin (88%), and vinorelbine/cisplatin (88%). However, symptoms of nausea and diarrhea were higher in the gemcitabine/cisplatin and irinotecan/cisplatin arms than in the other study arms. A trial conducted by the Italian Lung Cancer Group comparing the paclitaxel/carboplatin regimen with gemcitabine/cisplatin and vinorelbine/cisplatin demonstrated equal efÞcacy but some toxicity differences between the different regimens (Scagliotti GV, 2002). The trial randomized 612 patients with locally advanced (stage IIIB with either pleural effusion or N3 supraclavicular

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nodal disease), recurrent, and/or metastatic disease. After a median follow-up of approximately nine months, no signiÞcant differences in overall response rates were observed (vinorelbine/cisplatin, 30%; paclitaxel/carboplatin, 32%; gemcitabine/cisplatin, 30%). No signiÞcant differences were observed in the other end points: overall survival, time to disease progression, and time to treatment failure. Median survival times were 9.5 months for vinorelbine/cisplatin, 9.9 months for paclitaxel/carboplatin, and 9.8 months for gemcitabine/cisplatin. SigniÞcant differences were observed in toxicity proÞles (indicated by percentages of cycles with toxicities). Rates of grades III and IV thrombocytopenia were higher in the gemcitabine/cisplatin and paclitaxel/carboplatin arms (16% and 3%, respectively) than in the vinorelbine/cisplatin arm (0.1%). Rates of grades III and IV neutropenia and anemia were lower in the gemcitabine/cisplatin arm (neutropenia, 17%; anemia, 6%) and the paclitaxel/carboplatin arm (35% and 2%, respectively) than in the vinorelbine/cisplatin group (43% and 7%). A Phase III trial has demonstrated the efÞcacy of vinorelbine/cisplatin as adjuvant chemotherapy in completely resected NSCLC. A total of 482 patients with completely resected stage I or II NSCLC were randomized to receive four cycles of vinorelbine/cisplatin or follow-up only. For the vinorelbine/cisplatin patients, overall survival was signiÞcantly prolonged: 94 months versus 73 months (p = 0.0). Five-year survival was also improved: 69% versus 54%. The most common side effects in patients receiving vinorelbine/cisplatin were fatigue (77%), nausea (76%), anorexia (53%), vomiting (46%), sensory neuropathy (45%), and constipation (44%); two patients died of drug-related toxicities (Winton TL, 2004). This randomized clinical trial was the Þrst such trial to demonstrate that a thirdgeneration, platinum-based, dual combination regimen prolongs survival after surgery in early-stage NSCLC. Etoposide/Cisplatin Overview. The combination regimen of etoposide (Bristol-Myers Squibb’s VePesid, PÞzer/Nippon Kayaku’s Lastet, generics) and cisplatin (Bristol-Myers Squibb’s Platinol-AQ, generics) is active and well tolerated in patients with NSCLC and was considered a standard regimen throughout the 1980s and 1990s. Although still used by some oncologists (especially in combination with radiation) in Europe, this regimen is gradually being replaced by other regimens with lower toxicity and easier outpatient administration. Mechanism of Action. The individual components of the etoposide/cisplatin regimen contribute the following mechanisms to achieve its overall activity: •

Etoposide (Figure 10) is an epipodophyllotoxin. Epipodophyllotoxins act by causing single-strand breaks in DNA. Epipodophyllotoxins also inhibit DNA topoisomerase II to cause metaphase arrest, thereby inducing cytotoxic activity. Etoposide is cell-cycle-phase-speciÞc, with predominant activity occurring in late S phase and G2.

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R


O O HO

O O OH O O O O CH3O

OCH3 RO

FIGURE 10. Structure of etoposide (R = H, R1 = CH3 ).

•

Cisplatin (Figure 6) is a platinum agent. Platinum agents inhibit tumor-cell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that cisplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle.

Clinical Performance. The ECOG conducted a randomized Phase III trial that compared paclitaxel/cisplatin with a standard chemotherapy regimen of etoposide/cisplatin in PS 0 or 1, chemotherapy-n¨aive patients with advanced NSCLC (Bonomi P, 2000). A total of 574 patients with stage IIIB or IV NSCLC (enrolled August 1993 through December 1994) were randomized to receive either paclitaxel/cisplatin (two different arms: low-dose or high-dose paclitaxel) or etoposide/cisplatin. Paclitaxel was administered at different dose levels: 135 mg/m2 or 250 mg/m2 . All 574 patients were analyzed. The paclitaxel/cisplatin regimen was superior in terms of survival (median survival time, 9.9 months; one-year survival rate, 38.9%) compared with etoposide/cisplatin (median survival time, 7.6 months; one-year survival rate, 31.8%). The median survival duration for stage IIIB patients taking etoposide/cisplatin was 7.9 months; for all paclitaxel patients, median survival was 13.1 months. The median survival time for stage IV patients on the etoposide/cisplatin regimen was 7.6 months; for patients taking paclitaxel/cisplatin, median survival was 8.9 months. The objective response rate was 12.4% for etoposide/cisplatin, 27.7% for high-dose paclitaxel combined with cisplatin, and 25.3% for low-dose paclitaxel combined with cisplatin. The overall complete remission rate was 1.6%. Researchers found that paclitaxel combined with cisplatin produces a modest survival improvement, compared with etoposide/cisplatin, without producing negative effects on QOL. Based on the results of this study, paclitaxel/cisplatin replaced etoposide/cisplatin as the ECOG reference regimen for future studies. Docetaxel/Cisplatin Overview. Docetaxel (SanoÞ-Aventis’s Taxotere) in combination with cisplatin (Bristol-Myers Squibb’s Platinol-AQ, generics) was approved by the FDA in

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FIGURE 11. Structure of docetaxel.

November 2002 for Þrst-line treatment of patients with unresectable, locally advanced, or metastatic NSCLC who have not received prior chemotherapy. The regimen was approved in Europe in January 2003 but is not approved in Japan. In preclinical models, docetaxel was found to be synergistic with platinum-based compounds and with radiation (Davies AM, 2003). Mechanism of Action. The individual components of the docetaxel/cisplatin regimen contribute the following mechanisms to achieve its overall activity: •

•

Docetaxel (Figure 11) is a taxane. Taxanes inhibit the dynamic reorganization of microtubule networks that is essential to vital interphase and mitotic cellular functions. Docetaxel achieves selective toxicity against rapidly proliferating cells and is mainly active in the S phase of the cell cycle. Cisplatin (Figure 6) is a platinum agent. Platinum agents inhibit tumor-cell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that cisplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle.

Clinical Performance. Recent Þndings are encouraging for docetaxel/cisplatin when compared with those for some other regimens for Þrst-line treatment of advanced NSCLC. A multicenter, international, prospective, open-label, randomized Phase III trial compared the effects of docetaxel/cisplatin and docetaxel/carboplatin with those of the control regimen, vinorelbine/cisplatin, in chemotherapy-n¨aive patients with advanced NSCLC (Fossella F, 2003). Although the superiority of platinum plus docetaxel was not demonstrated (using a signiÞcance level of 0.028), survival estimates and overall response rates clearly favored docetaxel/cisplatin. Also, although no direct comparison of docetaxel/carboplatin against docetaxel/cisplatin was carried out, survival estimates and response rates were very similar (see the “Docetaxel/Carboplatin” section). In the Phase III trial, a total of 1,218 patients with locally advanced or recurrent (stage IIIB) or metastatic (stage IV) NSCLC were randomized to receive treatment with docetaxel/cisplatin, docetaxel/carboplatin, or vinorelbine/cisplatin (the control regimen). Median survival for vinorelbine/cisplatin was 10.1 months, compared with 11.3 months for docetaxel/cisplatin. One-year survival rates were

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41% for vinorelbine/cisplatin and 46% for docetaxel/cisplatin; two-year survival rates were 14% and 21%, respectively. Overall response rates were 24.5% for vinorelbine/cisplatin and 31.6% for docetaxel/cisplatin. Survival and overall response rates were similar for vinorelbine/cisplatin and docetaxel/carboplatin. Vinorelbine/cisplatin was associated with signiÞcantly more grades III and IV anemia, nausea, and vomiting than the docetaxel combinations. Grades III to IV diarrhea occurred more frequently in both the docetaxel combination regimens than in the vinorelbine/cisplatin regimen. Occurrence of infection and febrile neutropenia was similar in all three trial arms. Both docetaxel/cisplatin and docetaxel/carboplatin were better tolerated and consistently improved patient QOL compared with vinorelbine/cisplatin; investigators noted a better safety proÞle, improved QOL, higher two-year survival rate, and more convenient dosing schedule. Gemcitabine/Carboplatin Overview. Gemcitabine (Eli Lilly’s Gemzar) combined with carboplatin (Bristol-Myers Squibb’s Paraplatin, generics), like all carboplatin-containing regimens, is perceived to be better tolerated and easier to administer than its cisplatin-containing counterpart regimen. Phase II data that compare these two regimens directly prove that gemcitabine/carboplatin’s side-effect proÞle is more advantageous than that of gemcitabine/cisplatin, but Phase III data are required to address efÞcacy as well as toxicity differences. Mechanism of Action. The individual components of the gemcitabine/ carboplatin regimen contribute the following mechanisms to achieve its overall activity: •

•

Gemcitabine is an antimetabolite. Antimetabolites act by blocking normal DNA synthesis, thus stopping cell replication. Gemcitabine exhibits cellphase speciÞcity, primarily killing cells that are undergoing DNA synthesis (S phase) and also blocking the progression of cells through the G1/S-phase boundary. Carboplatin (Figure 12) is a platinum agent. Platinum agents inhibit tumorcell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that carboplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle.

Clinical Performance. Data on a Phase III trial comparing gemcitabine/carboplatin with gemcitabine/paclitaxel in the Þrst-line treatment of advanced NSCLC were presented at the 2005 American Society of Clinical Oncology (ASCO) meeting (Kosmidis PA, 2005). In 445 evaluable patients, after a median follow-up time 23 months, no signiÞcant differences in efÞcacy end points were observed. Overall survival and one-year survival rates were 10.5

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FIGURE 12. Structure of carboplatin.

months and 41% for gemcitabine/carboplatin versus 10 months and 43% for gemcitabine/paclitaxel. However, statistically signiÞcant differences were seen in grade III/IV toxicity, with gemcitabine/carboplatin exhibiting more neutropenia, thrombocytopenia, and anemia than gemcitabine/paclitaxel (28%, 25%, and 12.5%, versus 17%, 2.5% and 5%, respectively) but with gemcitabine/paclitaxel exhibiting more neurotoxicity (5% versus 0.5%). The superiority of gemcitabine/carboplatin over older triple-combination regimens has been demonstrated. In a Phase III trial, 422 patients with previously untreated stage IIIB or IV NSCLC were randomly assigned to treatment with either gemcitabine/carboplatin or mitomycin (generics)/ifosfamide (BristolMyers Squibb’s Ifex, Baxter’s Holoxan/Mitoxana, Shionogi’s Ifomide, generics)/cisplatin (MIC). Gemcitabine/carboplatin demonstrated a signiÞcant survival advantage compared with MIC (p = 0.008). Median survival was 10 months with gemcitabine/carboplatin and 7.6 months with MIC; one-year survival rates were 40% and 30%, respectively. Overall response rates were similar (42% versus 41%). More thrombocytopenia occurred with gemcitabine/carboplatin than with MIC, but less nausea, vomiting, constipation, and alopecia (Rudd RM, 2005). A Phase II randomized study assessed the efÞcacy (response was the primary end point) and the toxicity of gemcitabine/carboplatin compared with gemcitabine/cisplatin in 120 patients with NSCLC (Mazzanti P, 2003). A total of 533 cycles were delivered, with a median of four cycles per patient. The objective response rate was 41.9% for gemcitabine/cisplatin and 31% for gemcitabine/carboplatin. Investigators observed no signiÞcant differences in median survival (gemcitabine/carboplatin, 10.8 months; gemcitabine/cisplatin, 10.8 months) and median time to progression (gemcitabine/carboplatin, 5.1 months; gemcitabine/cisplatin, 5.4 months). Both regimens were very well tolerated, with no statistical differences between arms in grades III and IV toxicities. When all toxicity grades were combined, emesis, neuropathy, and renal toxicity occurred more frequently in the gemcitabine/cisplatin arm. Researchers concluded that gemcitabine/carboplatin did not provide a signiÞcant difference in response rate compared with gemcitabine/cisplatin but did offer better overall tolerability. Paclitaxel/Carboplatin Overview. The combination regimen of paclitaxel (Bristol-Myers Squibb’s Taxol, generics) and carboplatin (Bristol-Myers Squibb’s Paraplatin, generics) is

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commonly used, although some doubt exists about its efÞcacy relative to paclitaxel/cisplatin. Carboplatin is more expensive than cisplatin, which limits its use, especially in countries where drug use is inßuenced by budgetary constraints. Mechanism of Action. The individual components of the paclitaxel/ carboplatin regimen contribute the following mechanisms to achieve its overall activity: •

•

Paclitaxel (Figure 7) is a taxane. Taxanes act by inhibiting the dynamic reorganization of microtubule networks that is essential to vital interphase and mitotic cellular functions. Taxanes exhibit a unique mechanism of action. Paclitaxel, which achieves selective toxicity against rapidly proliferating cells, is mainly active in the G2/M phase of the cell cycle. Carboplatin (Figure 12) is a platinum agent. Platinum agents inhibit tumorcell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that carboplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle.

Clinical Performance. A trial conducted by the Italian Lung Cancer Group compared the paclitaxel/carboplatin regimen with gemcitabine/cisplatin and vinorelbine/cisplatin (Scagliotti GV, 2002). The trial randomized 612 patients with locally advanced (stage IIIB with either pleural effusion or N3 supraclavicular nodal disease), recurrent, and/or metastatic disease. After a median follow-up of approximately nine months, no signiÞcant difference in overall response rates was observed (paclitaxel/carboplatin, 32%; vinorelbine/cisplatin, 30%; gemcitabine/cisplatin, 30%). No signiÞcant differences were observed in the other end points: overall survival, time to disease progression, and time to treatment failure. Median survival times were paclitaxel/carboplatin, 9.9 months; vinorelbine/cisplatin, 9.5 months; and gemcitabine/cisplatin, 9.8 months. Some signiÞcant differences in toxicity proÞles were observed (indicated by percentages of cycles with toxicity). Rates of grades III and IV thrombocytopenia were higher in the gemcitabine/cisplatin and paclitaxel/carboplatin arms (16% and 3%, respectively) than in the vinorelbine/cisplatin arm (0.1%). Rates of grades III and IV neutropenia and anemia were lower in the gemcitabine/cisplatin (neutropenia, 17%; anemia, 6%) and paclitaxel/carboplatin arms (35% and 2%, respectively) than in the vinorelbine/cisplatin arm (43% and 7%, respectively). No serious hemorrhagic events were noted in any arm. The numbers of patients with febrile neutropenia were low across all treatment groups: one receiving gemcitabine/cisplatin, two receiving paclitaxel/carboplatin, and six receiving vinorelbine/cisplatin. A landmark trial conducted by the ECOG (ECOG 1594) compared four commonly used third-generation chemotherapy regimens: a reference regimen of paclitaxel/cisplatin and three experimental regimens: gemcitabine/cisplatin, docetaxel (SanoÞ-Aventis’s Taxotere)/cisplatin, and paclitaxel/carboplatin (Schiller JH, 2002). The trial randomized 1,207 chemotherapy-naive patients with

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advanced stage IIIB or IV disease and good PS (2 or better). The overall response rate for all 1,155 evaluable patients was 19%; median survival was 7.9 months, and one- and two-year survival rates were 33% and 11%, respectively. No signiÞcant differences in response rate or survival were observed between the three experimental arms. Median survival times were 8.1 months for gemcitabine/cisplatin, 7.4 months for docetaxel/cisplatin, and 8.1 months for paclitaxel/carboplatin; the survival rates for these three groups at one year were 36%, 31%, and 34%, respectively; survival rates at two years were 13%, 11%, and 11%, respectively. The median time to disease progression did not differ signiÞcantly in any of the four groups. Major toxicities (grades III and IV) occurred in all groups, particularly in patients with a poor PS. None of the four treatment arms demonstrated a signiÞcant advantage over the others in terms of survival, but the paclitaxel/carboplatin arm had a lower rate of toxic effects than the other arms. Based on the results of this study, the ECOG chose the paclitaxel/carboplatin regimen as the reference regimen for future studies. This combination is also the Þrst carboplatin-based dual combination regimen to demonstrate improved survival after resection. The Cancer and Leukemia Group B (CALGB) 9633 trial randomized 344 patients with fully resected stage IIB disease, within 4–8 weeks of resection, to either treatment with four cycles of paclitaxel/carboplatin or to observation only (Strauss GM, 2004). During a median follow-up of 34 months, there were 36 deaths from any cause among the 173 patients in the paclitaxel/carboplatin group and 52 deaths among the 171 patients in the observation group (p = 0.028). Survival rates at four years were 71% in the paclitaxel/carboplatin and 59% in the observation group. In addition, a signiÞcant advantage in failure-free survival favored the chemotherapy group (p = 0.035). Vinorelbine/Carboplatin Overview. Vinorelbine (GlaxoSmithKline/Pierre Fabre’s Navelbine, generics) combined with carboplatin (Bristol-Myers Squibb’s Paraplatin, generics) is a well-tolerated, efÞcacious Þrst-line regimen for inoperable NSCLC patients (Cremonesi M, 2003; Masters GA, 2003). Nevertheless, vinorelbine/carboplatin is one of the least used regimens in the major pharmaceutical markets, likely because of the lack of randomized data that show its efÞcacy to be superior to that of other regimens. Mechanism of Action. The individual components of the vinorelbine/ carboplatin regimen contribute the following mechanisms to achieve its overall activity: •

Vinorelbine (Figure 9) is a vinca alkaloid. Vinca alkaloids act by binding with microtubular proteins of the mitotic spindle, thus leading to mitotic arrest or cell death. The drug achieves its selective toxicity against cancer cells by acting on proliferating cells.

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•

Carboplatin (Figure 12) is a platinum agent. Platinum agents inhibit tumorcell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that carboplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle.

Clinical Performance. In a Phase II study, the efÞcacy of vinorelbine/ carboplatin was investigated in 52 previously untreated patients who had locally advanced and metastatic NSCLC (Cremonesi M, 2003). All 52 patients were treated with vinorelbine/carboplatin, and all patients were assessable. The response rate was 34.6%; 30.8% of the patients had stable disease, and 34.6% progressed while on treatment. The median time to progression and overall survival were, respectively, 7.5 months and 12.3 months. The most common grades III and IV toxicities were granulocytopenia, observed in 26 patients (50%); no neutropenic fever was reported. The treatment was generally very well tolerated. In a Phase III trial that compared platinum and non-platinum-based therapies, 316 patients with advanced, inoperable NSCLC were treated with either vinorelbine/carboplatin or vinorelbine/gemcitabine. Median survival was 8.6 months for vinorelbine/carboplatin and 11.5 months for vinorelbine/gemcitabine; oneyear survival was signiÞcantly higher in the vinorelbine/gemcitabine arm: 48.9% versus 34.4% (p = 0.01). The response rates were 32.4% in patients receiving vinorelbine/carboplatin and 40.9% in patients receiving vinorelbine/gemcitabine (p = 0.19). Moreover, higher rates of hematologic and nonhematologic side effects were reported in the group receiving carboplatin (Abratt RP, 2004). Docetaxel/Carboplatin Overview. Doubt exists as to whether the docetaxel (SanoÞ-Aventis’s Taxotere) and carboplatin (Bristol-Myers Squibb’s Paraplatin, generics) regimen is as efÞcacious as docetaxel/cisplatin. However, the carboplatin regimen is easier to administer than the cisplatin regimen. Mechanism of Action. The individual components of the docetaxel/ carboplatin regimen contribute the following mechanisms to achieve its overall activity: •

•

Docetaxel (Figure 11) is a taxane. Taxanes act by inhibiting the dynamic reorganization of microtubule networks that is essential to vital interphase and mitotic cellular functions. Docetaxel, which achieves selective toxicity against rapidly proliferating cells, is mainly active in the S phase of the cell cycle. Carboplatin (Figure 12) is a platinum agent. Platinum agents inhibit tumorcell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that carboplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle.

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Clinical Performance. A randomized trial compared docetaxel/cisplatin and docetaxel/carboplatin against the control regimen of vinorelbine/cisplatin in chemotherapy-naive patients with advanced NSCLC and demonstrated some efÞcacy advantages of the two docetaxel/platinum regimens versus vinorelbine/cisplatin (Fossella F, 2003). Although efÞcacy endpoints were slightly better for docetaxel/cisplatin than for docetaxel/carboplatin, the difference was not signiÞcant. Median overall survival times were 9.4 months for docetaxel/carboplatin and 11.3 months for docetaxel/cisplatin. One-year survival rates were 38% for docetaxel/carboplatin and 46% for docetaxel/cisplatin; two-year survival rates were 18% and 21%, respectively. Response rates for docetaxel/carboplatin and docetaxel/cisplatin were 23.9% and 31.6%, respectively; median times to progression were 20 weeks and 22 weeks, respectively. In the comparison of docetaxel/carboplatin and vinorelbine/cisplatin, median survival times were 9.4 months and 9.9 months, respectively. One-year survival rates were 38% for docetaxel/carboplatin and 40% for vinorelbine/cisplatin; twoyear survival rates were 18% and 14%, respectively. Response rates were 23.9% for docetaxel/carboplatin and 24.5% for vinorelbine/cisplatin; median times to disease progression were 20 weeks and 22 weeks, respectively. Although the efÞcacy results of this trial have not demonstrated a clear advantage for one regimen over the others, some toxicity and QOL differences were observed. Vinorelbine/cisplatin was associated with signiÞcantly more grades III and IV anemia, nausea, and vomiting than the docetaxel combinations. However, grade III or IV diarrhea occurred more frequently with both docetaxel combinations than with vinorelbine/cisplatin; grade III or IV nonhematologic toxicities occurred less frequently in the docetaxel/carboplatin arm. Both docetaxel regimens were better tolerated and consistently improved patient QOL. Mitomycin/Ifosfamide/Cisplatin Overview. Mitomycin (generics) in combination with ifosfamide (Bristol-Myers Squibb’s Ifex, Baxter’s Holoxan/Mitoxana, Shionogi’s Ifomide, generics) and cisplatin (Bristol-Myers Squibb’s Platinol-AQ, generics) (MIC) is one of the most active “old” regimens developed and used mainly in Europe. Other triple combination regimens that incorporate vinca alkaloids with cisplatin/ifosfamide are also used in NSCLC, albeit infrequently. In the United States, triple combination regimens are not used. Mechanism of Action. The individual components of the MIC regimen contribute the following mechanisms to achieve its overall activity: •

•

Ifosfamide (Figure 13) and mitomycin are alkylating agents, which act by cross-linking with DNA during cell reproduction, thereby preventing synthesis of second-generation DNA. Both agents are cell-cycle-phase nonspeciÞc. Cisplatin (Figure 6) is a platinum agent. Platinum agents inhibit tumor-cell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that cisplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle.

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FIGURE 13. Structure of ifosfamide.

Clinical Performance. Two large, randomized trials have been carried out to determine whether MIC inßuences duration and QOL in localized, unresectable, and advanced NSCLC (Cullen MH, 1999). In the Þrst trial, MIC1, 461 localizeddisease patients were randomized to receive up to four cycles of MIC followed by radical radiotherapy (MIC plus radiotherapy [RT]) or to receive RT alone (control arm). The second trial, MIC2, randomized 359 advanced-stage patients to MIC with palliative care (PC) (MIC + PC) or to PC alone (control arm). Objective response rates in the chemotherapy arms of MIC1 and MIC2 were 54% and 32%, respectively. Median survival times in MIC1 were 11.7 months (MIC + RT) and 9.7 months (RT); median survival times in MIC2 were 6.7 months (MIC + PC) and 4.8 months (PC). Median overall survival was signiÞcantly greater in the chemotherapy arm than in the control arm in MIC2 but not in MIC1. Overall, the positive effect of MIC on survival was signiÞcant. MIC has also shown equivalent efÞcacy to gemcitabine/cisplatin in a large, multicenter, randomized Phase III trial in advanced NSCLC. Gemcitabine/cisplatin and MIC were compared in 307 patients with stage IIIB or IV NSCLC (Crino L, 1999). The response rate was signiÞcantly higher in the gemcitabine/cisplatin arm (38%) than in the MIC arm (26%). The median duration of response was 8.7 months for gemcitabine/cisplatin and 8.2 months for MIC. In stage IV disease, the response rate was statistically superior in the gemcitabine/cisplatin arm (37% versus 23%). Researchers found no statistically signiÞcant difference in overall median survival times between the gemcitabine/cisplatin and MIC arms (8.6 months versus 9.6 months). The one-year survival rate was 33% in the gemcitabine/cisplatin arm and 34% in the MIC arm. No differences were found in median time to progression (gemcitabine/cisplatin, 5 months; MIC, 4.8 months) and median time to treatment failure (4 months and 3.7 months, respectively). Nedaplatin Regimens Overview. Nedaplatin (Shionogi’s Aqupla) is an effective compound against NSCLC. Nedaplatin is indicated in Japan for treatment of several cancers, including lung cancer, but it is not available in the United States or Europe. This agent is used in combination with a wide variety of chemotherapy agents but is most commonly used with gemcitabine (Eli Lilly’s Gemzar), irinotecan (PÞzer’s

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Camptosar, Yakult/PÞzer’s Campto, Daiichi’s Topotecin), or vinorelbine (GlaxoSmithKline/Pierre Fabre’s Navelbine, generics). In preclinical studies, nedaplatin has been shown to have a synergistic effect with gemcitabine (Kurata T, 2003). Mechanism of Action. Nedaplatin is a platinum agent. Platinum agents inhibit tumor-cell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that nedaplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle. Clinical Performance. Recent studies have investigated combinations of nedaplatin with irinotecan or gemcitabine. A study by Oshita and colleagues was split into two parts; a dose-ranging study (Phase I) and a Phase II study at the maximum tolerated dose (MTD) (Oshita F, 2003). In the Phase I study, 42 patients were treated with irinotecan (60 mg/m2 ) and nedaplatin starting at 50 mg/m2 and increasing by 10 mg/m2 until the MTD was reached. A nedaplatin dose of 100 mg/m2 was chosen for the Phase II dose. A total of 16 patients were entered into this study; seven patients (43.8%) had an objective response. In the 42 cycles administered, the adverse events were grade III or IV neutropenia (50% of cycles), grade III anemia (12% of cycles), and grade III or IV thrombocytopenia (7% of cycles). The overall response rate was 31%, and median survival time was 341 days; the one-year survival rate was 45.2%. Nineteen patients had stable disease, and seven had progressive disease. In a Phase I study of nedaplatin combined with gemcitabine in 12 evaluable patients with advanced NSCLC, researchers observed 1 partial response and 8 stable diseases (Kurata T, 2003). Adverse advents were grade III or IV neutropenia (41%) and grade III thrombocytopenia (17%). This study found that nedaplatin demonstrates promising antitumor activity against NSCLC with lower nephrotoxicity than cisplatin.

Docetaxel, Single Agent Overview. Docetaxel (SanoÞ-Aventis’ Taxotere) is the standard second-line treatment for patients with advanced/refractory NSCLC owing to its proven activity and documented survival beneÞt (Kosmas C, 2002). Single-agent docetaxel was established as a standard second-line treatment following two Phase III trials that demonstrated improved survival and QOL (see the following “Clinical Performance” section). Mechanism of Action. Docetaxel (Figure 11) is a taxane. Taxanes act by inhibiting the dynamic reorganization of microtubule networks that is essential to vital interphase and mitotic cellular functions. Docetaxel, which achieves selective toxicity against rapidly proliferating cells, is mainly active in the S phase of the cell cycle.

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Clinical Performance. Two Phase III trials of second-line chemotherapy for patients with NSCLC have shown signiÞcant advantages in docetaxel treatment compared with best supportive care, in terms of survival and QOL, and superior efÞcacy compared with other single agents. In the TAX 317 trial, 204 patients were randomized to receive either docetaxel (100 mg/m2 or 75 mg/m2 ) or best supportive care (Fossella FV, 2002; Shepherd FA, 2000). Median survival was 7.5 months with low-dose docetaxel and 4.6 months with best supportive care; oneyear survival rates were 37% and 11%, respectively. Partial response to both doses of docetaxel was observed in 6% of patients; 40% of patients had stable disease. The median time to disease progression was 12.3 weeks with low-dose docetaxel and 7 weeks with best supportive care. A statistically signiÞcant improvement in disease-related symptoms was observed in patients taking docetaxel compared with patients receiving best supportive care. In the TAX 320 study, 373 patients were randomized to receive docetaxel at 100 mg/m2 or 75 mg/m2 or to receive either vinorelbine or ifosfamide (control arm) (Fossella FV, 2000). Partial response rates were 6.7%, 10.8%, and 0.8%, respectively, for low-dose docetaxel, high-dose docetaxel, and the control arm. Approximately one-third of patients in each group had stable disease. The median durations of response were 9.1 months in the low-dose docetaxel arm and 7.5 months in the high-dose arm. The one-year survival rates were 32% for low-dose docetaxel, 21% for high-dose docetaxel, and 19% for the control arm; median survival times were 5.7 months, 5.5 months, and 5.6 months, respectively. Median time to disease progression was about 8 weeks in each of the three groups; 26-week, progression-free survival rates in the low-dose docetaxel, high-dose docetaxel, and control arms were 17%, 19%, and 8%, respectively. In a second survival data analysis of the TAX 320 study, response rates to docetaxel were equivalent in patients who had received prior paclitaxel (10.5%) and patients who had not received prior paclitaxel (8.5%) (Fossella FV, 2002). One-year survival rates for patients with no prior paclitaxel therapy were 32% in the low-dose docetaxel arm, 21% in the high-dose docetaxel arm, and 19% in the control arm; one-year survival rates were 30% (low-dose docetaxel arm) and 17% (control arm). Grade IV neutropenia and febrile neutropenia occurred more frequently in both docetaxel groups, but infection, grade IV thrombocytopenia, and severe, nonhematologic side effects occurred equally across all three arms. Recent data presented at the 2004 American Society of Clinical Oncology (ASCO) meeting are likely to support docetaxel monotherapy in second-line treatment of NSCLC. Phase II data demonstrate that the addition of irinotecan or gemcitabine to docetaxel in second-line chemotherapy produces more toxicity without any increase in response rate or survival times (Pectasides D, 2004; Takeda K, 2004). Gemcitabine, Single Agent Overview. Single-agent gemcitabine (Eli Lilly’s Gemzar) is rarely used as Þrstline treatment of NSCLC, but like most single agents, it is generally used for

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elderly or poor PS patients who cannot tolerate combination therapy or as a second- or third-line treatment. Mechanism of Action. Gemcitabine is an antimetabolite. Antimetabolites act by blocking normal DNA synthesis, thus stopping cell replication. Gemcitabine exhibits cell-phase speciÞcity, primarily killing cells undergoing DNA synthesis (S phase) and also blocking the progression of cells through the G1/S-phase boundary. Clinical Performance. Single-agent gemcitabine has been studied extensively. In a majority of Phase II trials involving more than 400 patients with NSCLC, the response rate was found to be approximately 20% (Manegold C, 2000). Some studies show that gemcitabine is effective and well tolerated in elderly patients. In the open-label, randomized Multicenter Italian Lung Cancer in the Elderly Study (MILES) Phase III trial, the effectiveness and toxicity of the vinorelbine/gemcitabine combination were compared with the effectiveness and toxicity of each drug given as a single agent (Gridelli C, 2003). Results show that the combination does not improve survival and is more toxic than either drug given alone. Six hundred and ninety-eight patients aged 70 or older were randomized to receive vinorelbine/gemcitabine or either drug as a single agent. Median survival times for patients in the single-agent vinorelbine and gemcitabine arms were 36 weeks and 28 weeks, respectively; median survival time for patients in the combination arm was 30 weeks. The difference in survival was not statistically signiÞcant. Median times to disease progression were 18 weeks for vinorelbine, 17 weeks for gemcitabine, and 19 weeks for the combination; response rates were 18%, 16%, and 21%, respectively. Vinorelbine/gemcitabine caused more neutropenia, vomiting, fatigue, cardiac toxicity, and constipation than single-agent gemcitabine.

Vinorelbine, Single Agent Overview. Vinorelbine (GlaxoSmithKline/Pierre Fabre’s Navelbine, generics) is effective as a single agent in inoperable or advanced NSCLC and is also used in other cancers, such as breast and ovarian cancers. Although it is approved as a monotherapy in Þrst-line therapy, it is rarely used in this setting; it is occasionally used as a second- or third-line agent in elderly patients or in patients who are failing Þrst-line chemotherapy. Single-agent vinorelbine is known to prolong survival and improve QOL in elderly patients with advanced NSCLC. Mechanism of Action. Vinorelbine (Figure 9) is a vinca alkaloid. Vinca alkaloids act by binding with microtubular proteins of the mitotic spindle, thereby leading to mitotic arrest or cell death. The drug achieves its selective toxicity against cancer cells by acting on proliferating cells.

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Clinical Performance. Because of its favorable toxicity proÞle, single-agent vinorelbine has been investigated as a treatment for elderly patients with advanced NSCLC; vinorelbine is known to prolong survival and improve QOL in these patients. In the open-label, randomized MILES Phase III trial, the effectiveness and toxicity of the combination of vinorelbine/gemcitabine were compared with the effectiveness and toxicity of each drug given as a single agent (Gridelli C, 2003). Results show that the combination does not improve survival and is more toxic than either drug given alone. Six hundred and ninety-eight patients aged 70 or older were randomized to receive vinorelbine/gemcitabine or either drug as a single agent. Median survival times for patients in the single-agent vinorelbine and gemcitabine arms were 36 weeks and 28 weeks, respectively; the median survival time for patients in the combination arm was 30 weeks. The difference in survival was not statistically signiÞcant. Median times to disease progression were 18 weeks for vinorelbine, 17 weeks for gemcitabine, and 19 weeks for the combination; response rates were 18%, 16%, and 21%, respectively. Vinorelbine/gemcitabine caused more thrombocytopenia and hepatic toxicity than single-agent vinorelbine. Gefitinib, Single Agent Overview. GeÞtinib (AstraZeneca’s Iressa) has been withdrawn by the time of composing this reference. However, it is of some historical and educational interest to note its stages of progressive development. It was the Þrst compound to be launched in a new class of targeted anticancer drugs that inhibit the tyrosine kinase activity of the epidermal growth factor receptor (EGFR). Following Japanese approval in 2002, geÞtinib was launched in October 2003 in the United States for third-line treatment of advanced NSCLC after gaining accelerated FDA approval. However, after a follow-up Phase III trial failed to show an overall survival advantage in patients receiving geÞtinib (AstraZeneca, press release, 2004), the future of the molecule seems doubtful. Soon after results of the Iressa Survival Evaluation in Lung (ISEL) cancer trial were announced in December 2004, the FDA issued a statement informing physicians and patients that “alternative therapies are available.” European approvals for geÞtinib are currently on hold, and the FDA’s accelerated approval program authorizes the agency to remove a drug from the market if a postmarketing clinical study fails to verify clinical beneÞt. The FDA’s Oncology Drugs Advisory Committee (ODAC) discussed the future of Iressa in March 2005. As mentioned above, GeÞtinib has been withdrawn by the time of writing. Mechanism of Action. GeÞtinib is a small-molecule inhibitor that speciÞcally inhibits EGFR tyrosine kinase (TK). The drug’s precise mechanism of action is unknown, but the drug appears to impede cell-cell signaling pathways that have been implicated in rapid cell division and survival. Overactivation of these pathways is thought to be central to tumor growth and metastasis. Researchers

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initially thought that because EGFR is expressed more in cancerous than in healthy tissue, geÞtinib’s EGFR TK inhibitory activity is directly related to its anticancer activity. However, it is now evident that some EGFR inhibitors are active in EGFR-negative patients as well as in EGFR-positive patients and that EGFR expression does not correlate with response to geÞtinib. Further research is required to fully understand geÞtinib’s mechanism of action. Clinical Performance. GeÞtinib’s approval in the third-line setting was based on data from two randomized, double-blind U.S. Phase II trials (Iressa Dose Evaluation in Advanced Lung Cancer [IDEAL-1 and IDEAL-2]). In IDEAL-2, 216 patients with locally advanced or metastatic NSCLC who had failed platinumbased or docetaxel chemotherapies were randomized to receive oral geÞtinib at 250 mg per day or 500 mg per day (Kris MG, 2002). Treatment was continued until disease progression or unacceptable toxicity occurred. In the 142 evaluable patients, the objective tumor response rates for the 250 mg/day and 500 mg/day doses were 11.8% and 8.8%, respectively. The duration of tumor response ranged from three to seven months. Stable disease was observed in 31% of the 250 mg/day patients and in 27% of the 500 mg/day patients. Symptom response rates were 43% and 35%, respectively; the duration of response ranged from 1 to 7.4 months. Median survival was 6.1 months in the 250 mg/day dose group and 6 months in the 500 mg/day group. The 500 mg/day dose caused greater toxicity, as measured by the incidence of adverse events and withdrawals due to drug-related adverse events. The majority of drug-related adverse events were generally mild, consisting of grade I or II diarrhea and skin rash. Researchers concluded that both doses of geÞtinib showed clinically signiÞcant antitumor activity with acceptable tolerability. The IDEAL-1 trial was a large, international (Europe, Australia, South Africa, Japan), double-blind trial that evaluated geÞtinib 250 mg/day and 500 mg/day in 210 patients with advanced NSCLC who had received one or two prior chemotherapy regimens (at least one platinum-based regimen) (Fukuoka M, 2002). In the 208 evaluable patients, researchers found no difference between the two doses in terms of objective response, disease control rate, overall survival, and progression-free survival. The objective response rates were 18.4% for the 250 mg/day dose and 19% for the 500 mg/day dose; overall response rates were 54.4% and 51.4%, respectively. Overall survival times were 7.6 months for the 250 mg/day dose and 8.1 months for the 500 mg/day dose; progression-free survival times were 2.7 and 2.8 months, respectively. As it did in IDEAL-2, the 500 mg/day dose in IDEAL-1 caused greater toxicity and more withdrawals than the 250 mg/day dose. Adverse events were generally mild; the most common were grade I or II rash, diarrhea, pruritus, and dry skin. Patients receiving geÞtinib 250 mg/day suffered fewer grades III and IV adverse events than patients receiving 500 mg/day. Oral geÞtinib 250 mg/day demonstrated clinically signiÞcant antitumor activity with a favorable tolerability proÞle. In December 2004, AstraZeneca reported disappointing results from the ISEL trial. This study had enrolled 1,692 patients with advanced NSCLC who had

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progressed or were no longer able to tolerate chemotherapy. Despite a statistically signiÞcant improvement in tumor shrinkage, geÞtinib monotherapy failed to prolong survival in the overall population compared with placebo (median survival was 5.6 versus 5.1 months; p = 0.11), or in patients with adenocarcinoma (median survival was 6.3 versus 5.4 months; p = 0.07) (AstraZeneca, press release, 2004). Lacking any obvious reason why these results differ from the positive data generated in the recent BR21 trial of the EGFR inhibitor erlotinib in a similar patient population (see the “Erlotinib” section), the future of geÞtinib looks uncertain. The reasons behind these and other negative results remain unclear. Analysis of tumor biopsies from the Iressa NSCLC Trial Assessing Combination Treatment (INTACT) trials (discussed in the following paragraph) suggests that there is no correlation between EGFR expression and response to geÞtinib therapy (Bailey LR, 2004), while statistical analysis of the IDEAL-1 trial showed that certain NSCLC subpopulations respond better to geÞtinib than others (Fukuoka M, 2003). Response rates were higher for Japanese patients than for non-Japanese patients (27.5% versus 10.4%). A pharmacokinetic analysis of geÞtinib plasma concentrations did not reveal any differences between Japanese and non-Japanese patients that might explain the difference in response. Further investigation revealed that the odds in favor of response were more than six times higher for PS 0 or 1 patients and patients who had received prior immuno/hormonal treatment than for PS 2 patients and patients without prior immuno/hormonal treatment. Response rates appeared to be higher in females than in males (17.5% versus 5.1%), higher in nonsmokers than in previous or current smokers (29.4% versus 4.6%), and higher in adenocarcinomas than in other histologies (12.4% versus 6.7%). Data presented at the 2005 ASCO meeting shows that patients with certain mutations respond better than others (Lynch TJ, 2005). Tumor samples were obtained from patients enrolled in the IDEAL and INTACT studies. Selected samples were analyzed for ras and p53 mutations. Correlations were made between mutational status, therapies, and outcome measures including tumor response and survival. Among 416 IDEAL patients, 119 had available tumor samples, and 78 were fully evaluable. Fourteen mutations were detected. Of the 14 patients with mutations, 6 had objective tumor responses. Among 2,130 INTACT patients, there were 672 available tumor samples. Of 312 evaluable samples to date, 32 mutations have been detected. Of 23 patients with mutations, 15 had objective tumor responses to treatment. GeÞtinib has been evaluated in combination with platinum-based chemotherapy regimens for Þrst-line treatment for advanced NSCLC; results so far have been negative. In the INTACT-1 and INTACT-2 trials, 2,130 chemotherapynaive patients with advanced NSCLC were randomized to a standard platinum-based chemotherapy regimen (gemcitabine/cisplatin for INTACT-1, paclitaxel/carboplatin for INTACT-2) or to the same chemotherapy regimen combined with geÞtinib. Results from both trials showed no beneÞt from the addition of geÞtinib to chemotherapy in the treatment of advanced NSCLC. The addition of geÞtinib did not demonstrate any increase, or trend toward such an increase,

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in tumor response rates, time to progression, or overall survival (Giaccone G, 2002; Johnson DH, 2002). As a result, geÞtinib is not indicated for use in this setting. Erlotinib, Single Agent Overview. Erlotinib (OSI/Genentech/Roche’s Tarceva) is an orally active EGFR inhibitor. In November 2004, after review under the FDA’s accelerated approval program, erlotinib was approved and launched in the United States for treatment of patients with locally advanced or metastatic NSCLC after failure of at least one prior chemotherapy regimen. Erlotinib was also granted fast-track status for treatment of patients with chemotherapy-naive stage III and IV NSCLC. In March 2005, erlotinib received approval in Switzerland for treatment of locally advanced or metastatic NSCLC after the failure of at least one prior chemotherapy regimen. The drug has in fact been launched in Europe in October 2005. Approval in Japan is expected as well. Mechanism of Action. Erlotinib is a small-molecule inhibitor that speciÞcally inhibits EGFR tyrosine kinase (TK). The drug’s precise mechanism of action is unknown, but it appears to impede cell-cell signaling pathways that have been implicated in rapid cell division and survival. Overactivation of these pathways is thought to be central to tumor growth and metastasis. Researchers initially thought that because EGFR is expressed more in cancerous than in healthy tissue, erlotinib’s EGFR TK inhibitory activity is directly related to its anticancer activity. However, it is now evident that some EGFR inhibitors are active in EGFR-negative as well as in EGFR-positive patients and that EGFR expression does not correlate with response to erlotinib. Further research is required to fully understand erlotinib’s mechanism of action. Clinical Performance. Erlotinib is an oral, once-daily treatment that has been used mainly in stage IIIB and IV patients, but it is likely to also show effectiveness in patients with early-stage disease and low tumor burden. It may also require chronic administration for effective control of tumor cell growth, much as tamoxifen therapy is administered to breast cancer patients. The FDA approval of erlotinib for second-line monotherapy was based primarily on the Phase III CAN-NCIC-BR21 (BR21) study. This trial randomized 731 patients with IIB or IV NSCLC and PS 0–3, who had previously undergone one to two chemotherapy regimens, to treatment with erlotinib monotherapy (150 mg/day) or placebo for a median duration of 34 weeks. Overall survival was signiÞcantly longer in patients receiving erlotinib therapy (6.7 versus 4.7 months, p = 0.001) as was progression-free survival (2.23 versus 1.84 months, p < 0.001) (Shepherd FA, 2004). QOL measures, such as time to deterioration of patient-reported symptoms, also showed statistically and clinically meaningful beneÞt for patients receiving erlotinib. The most common adverse events were rash and diarrhea, but dropout due to toxicity was low: 5% of patients receiving

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erlotinib versus 2% of placebo patients. Interestingly, patients with rashes have been shown to have signiÞcantly longer survival times than patients without rashes, indicating that this side effect is a clinically important pharmacodynamic marker of EGFR inhibitor activity (Clark GM, 2003). Two Þrst-line Phase III trials have also investigated the combination of erlotinib with chemotherapy in advanced-stage NSCLC patients, with disappointing results. The trials, known as TRIBUTE in the United States and as the Tarceva Lung Cancer Investigation Trial (TALENT) in Europe, were placebo-controlled studies that randomized previously untreated patients with advanced NSCLC to treatment with erlotinib (150 mg/day) or placebo with six cycles of paclitaxel/carboplatin (TRIBUTE) or six cycles of gemcitabine/cisplatin (TALENT), followed by erlotinib maintenance monotherapy. In the 1,059-patient TRIBUTE study, no difference in median survival time or overall response rate was observed between patients treated with erlotinib and paclitaxel/carboplatin and patients treated with placebo and paclitaxel/carboplatin: survival times were and overall response rates were 10.8 versus 10.6 months and 21.5% versus 19.3%, respectively (Herbst RS, 2004). Similar results were obtained in the 1,172-patient TALENT study, which combined erlotinib with a gemcitabine/cisplatin regimen; no signiÞcant differences between the erlotinib and placebo groups were noted in overall survival or in time to progression, which were 301 days versus 309 days and 167 days versus 179 days, respectively (Gatzemeier U, 2004). The toxicity proÞle of erlotinib, combined with either standard chemotherapy regimen, was similar to that of the chemotherapy regimen alone, with the exception of increased incidence of grades III and IV diarrhea and grades III and IV skin rash. The investigators concluded that erlotinib in combination with gemcitabine/cisplatin did not improve survival or other treatment outcomes in patients with advanced NSCLC. In addition to these Phase III trials, erlotinib is being assessed in multiple Phase I and II trials in the neoadjuvant setting, in chemoradiotherapy (CRT) regimens, and in combination with other cytotoxic and biological agents such as bevacizumab (Genentech’s Avastin), the antivascular endothelial growth factor (VEGF) monoclonal antibody being developed by Genentech and Roche. Phase I/II data on erlotinib/bevacizumab in 40 patients, reported at the 2004 ASCO meeting, indicate that this combination is well tolerated with minimal drug interactions. Preliminary data suggest that this combination has potent antitumor activity: median overall survival and time to progression were 9.3 and 4.6 months, respectively (Sandler AB, 2004). Erlotinib has also shown efÞcacy in bronchoalveolar cell carcinoma (BAC), a subtype of NSCLC (Miller VA, 2003). In a Phase II trial, 69 patients with BAC received erlotinib (150 mg/day) therapy. Partial responses occurred in 15 of 59 evaluated patients; response rates negatively correlated with smoking habits (Kris MG, 2004). These results may suggest that tobacco-related genetic changes predispose resistance to this drug, but this relationship needs to be investigated in a Phase III trial. As with geÞtinib, genetic studies have been conducted with erlotinib in the hope of identifying subgroups that are more likely to beneÞt. Data

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FIGURE 14. Structure of pemetrexed.

were disclosed at the ASCO meeting in 2005 (Tsao MS, 2005). Erlotinib appeared to have a greater effect on survival in patients with EGFR protein expression. In a small, preliminary subset, EGFR ampliÞcation appeared to be a better predictor of response than the EGFR mutation. Pemetrexed Overview. Eli Lilly has developed and launched pemetrexed (Alimta), a multitargeted antifolate. In August 2004, the FDA approved pemetrexed as monotherapy for the treatment of locally advanced or metastatic NSCLC in patients who had failed at least one other chemotherapy regimen; in September 2004, pemetrexed was approved in Europe for the same indication. Pemetrexed is also approved in combination with cisplatin for Þrst-line treatment of unresectable mesothelioma (not covered in this report). Mechanism of Action . Pemetrexed (Figure 14) is an antifolate. The folate pathway is involved in the metabolism and synthesis of the DNA precursors (purines); antifolates inhibit this pathway and interfere with DNA synthesis, causing tumor cell death. Previous generations of antifolates were directed principally at the enzyme thymidylate synthase, a key enzyme in the folate pathway. Pemetrexed also inhibits the enzymes dihydrofolate reductase and glycinamideribonucleotide (GAR) formyl-transferase. Eli Lilly hopes that the multitargeted approach will overcome resistance acquired through overexpression of any single enzyme. Clinical Performance. A pivotal, multicenter, Phase III trial randomizing 571 previously treated patients with recurrent NSCLC showed that pemetrexed has equivalent efÞcacy to, and a better toxicity proÞle than, docetaxel. Ninety percent of the patients had prior platinum therapy and 28% had prior taxane therapy. Median survival was similar in both arms of the study: 8.3 months for pemetrexed and 7.9 months for docetaxel. However, the incidence of severe neutropenia was only 5% in patients treated with pemetrexed compared with 40% in patients treated with docetaxel. In addition, there were signiÞcant differences in the incidence of neutropenic fever and subsequent hospitalizations (2% for pemetrexed, 13% for docetaxel) and the incidence of drug-related, serious adverse events (10% for pemetrexed, 24% for docetaxel) (Hanna NH, 2003).

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Pemetrexed was also investigated in combination with either carboplatin (arm A) or oxaliplatin (arm B) as Þrst-line therapy in 80 patients with locally advanced or metastatic NSCLC. The conÞrmed response rate in arm A was 33% with 41% stable disease, versus 27% and 44%, respectively, in arm B. Median time to progression was 5.7 months in arm A and 5.5 months in arm B. Median overall survival time was 10.5 months in both arms; one-year survival was 43.9% in patients receiving pemetrexed/carboplatin and 49.9% in patients receiving pemetrexed/oxaliplatin. Grades III and IV hematologic toxicities in arm A included neutropenia (26%), febrile neutropenia (3%), thrombocytopenia (18%), and anemia (8%). In arm B, no grade IV toxicities were recorded, but grade III neutropenia (5%), thrombocytopenia (2%), and anemia (2%) were reported. Toxicity was low compared with other dual combination regimens, and response rates were similar, suggesting that one of these two combinations may provide a preferable risk-beneÞt proÞle for treatment of NSCLC patients (Scagliotti GV, 2005). Previous Phase II trials reported the beneÞts of combining pemetrexed with cisplatin in Þrst-line therapy for patients with stage IIIB or IV NSCLC: response rates were 39–45%, and one-year survival was approximately 50%. A Phase II trial presented at ASCO in 2003 sought to maintain efÞcacy but reduce the toxicity proÞle by substituting carboplatin for cisplatin; the overall response rate was 32%, and both hematologic and nonhematologic toxicities were minimal, which suggests that this regimen is promising (Zinner R, 2003). Preliminary data from a Phase II trial of pemetrexed and gemcitabine in the Þrst-line treatment of stages IIIB and IV NSCLC patients were presented at the 2004 ASCO meeting (Adjei AA, 2004). This study compared three different dosing regimens: pemetrexed then gemcitabine on day 1, followed by gemcitabine on day 8 (arm A); gemcitabine then pemetrexed on day 1, followed by gemcitabine on day 8 (arm B); or gemcitabine on day 1, followed by pemetrexed then gemcitabine on day 8 (arm C). Arm B was terminated early because of poor efÞcacy. A partial response rate of 29% was achieved in arm A compared with 17% in arm C. Febrile neutropenia was experienced by 5% of patients in arms A and C and 20% in arm B. Patients in arm A had less severe toxicities than those in arm C (grade III or higher events, 86% versus 93%; grade IV or higher events, 40% versus 50%). The authors concluded that the regimen used in arm A may be the most favorable in this combination. Other studies of pemetrexed in combination with gemcitabine have demonstrated a toxicity proÞle comparable to those of other dual combination regimens, and despite a low overall response rate, time to progression and median one-year survival were higher than reported with other two-drug combinations in cooperative group studies (Ettinger DS, 2002; Monnerat C, 2002). Trials are ongoing to investigate this combination further. Nonpharmacological Approaches Unlike chemotherapy, which is used systemically, surgery and radiotherapy are local treatments. Surgery offers the best chance of long-term curative treatment

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for patients with early-stage NSCLC. Surgical resection of the tumor is indicated for patients who are physically Þt enough to tolerate both the procedure and the anesthesia when complete resection of the tumor is anticipated. Usually, these are T1-4, NO-1 patients, and selected N2 patients—that is, stages I and II patients and selected stage IIIA patients (see the “Etiology and Pathophysiology” section for tumor classiÞcations and stage deÞnitions). To assess a patient’s operability, a surgeon conducts preoperative tests of cardiovascular and respiratory functions; these tests may include a stress test, an echocardiogram, and pulmonary function tests. Standard procedures for surgical removal of lung tumors include lobectomy (a whole lobe of the lung is removed), pneumonectomy (the whole lung is removed), and segmentectomy (wedge resection; only a small part of the lung is removed). In NSCLC, radiotherapy is most commonly administered via external-beam radiation. Because the delivery of radiation focuses on one region or Þeld of the body (the thorax), radiotherapy is considered a localized treatment modality. Radiotherapy is used to treat NSCLC in various settings. In early-stage NSCLC, radiotherapy is used in patients who are not surgical candidates, usually because of poor PS or poor cardiovascular function. In these patients, radical high-dose radiotherapy (with chemotherapy if the patient is deemed Þt enough to tolerate it) is administered with curative intent. Postsurgical adjuvant radiation therapy has become rare in stages I and II disease since a meta-analysis demonstrated a 7% reduction in overall two-year survival rates in patients who received adjuvant radiation (PORT Meta-Analysis Trialists Group, 1998), presumably due to the detrimental effects of radiation. Radiotherapy is commonly used in stage III disease, usually with sequential or concurrent chemotherapy. In patients with stage IIIA NSCLC, neoadjuvant radiation (with or without chemotherapy) may shrink tumors and increase the likelihood of surgical resectability. Numerous studies have shown that in stage IIIB NSCLC, the combination of chemotherapy and radiation increases survival rates compared with either treatment administered alone. In addition, radiation therapy may be used as an adjuvant to surgery to prevent or delay recurrence in stage IIIA disease, particularly when a tumor is upstaged during surgery. In patients with metastatic disease, radiation therapy is used primarily for palliation of speciÞc symptoms, such as bone pain due to tumor metastasis.

EMERGING THERAPIES The pipeline of drugs in development for non-small-cell lung cancer (NSCLC) has suffered recently from the withdrawal or failure of several drugs in late-stage clinical trials. A notable recent disappointment was Schering-Plough’s orally active farnesyl transferase inhibitor lonafarnib (Sarasar), which was in Phase III trials for NSCLC in the United States in combination with taxanes. The trials planned to enroll 800 patients, but enrollment was suspended after an interim data analysis in early 2004 revealed that the trials would not provide sufÞcient

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evidence of efÞcacy to warrant further development. Disappointing results from Phase III trials have also halted development of Isis Pharmaceuticals/Eli Lilly’s antisense RNA, ISIS-3521 (AfÞnitak), and SanoÞ-Aventis’ bioreductive agent, tirapazamine (Tirazone). The future of the COX-2 class of agents was cast into serious doubt by the withdrawal of valdecoxib (PÞzer’s Bextra) after an FDA review concluded that this agent poses unacceptable cardiovascular risks and the risk of potentially life-threatening, sometimes fatal skin reactions, but offers “no demonstrated advantage compared with other NSAIDs.” The FDA has also requested that all prescription NSAIDS marketed in the United States, including celecoxib, add a boxed warning to their drug labels that highlights the potentially increased risk of adverse cardiovascular events and gastrointestinal hemorrhage. These setbacks occurred after another COX-2 inhibitor, rofecoxib (Merck’s Vioxx), was found to increase the risk of cardiovascular events in patients with a history of colorectal adenomas. Despite some encouraging data, the high thromboembolic risk conferred by malignant diseases may prevent the development of COX-2 inhibitors for treatment of NSCLC. However, these disappointments have been offset by several recent successful launches; for example, the epidermal growth factor receptor (EGFR) inhibitors erlotinib (OSI/Genentech/Roche’s Tarceva) and geÞtinib (AstraZeneca’s Iressa) and the antifolate pemetrexed (Eli Lilly’s Alimta) have all received recent approval for NSCLC. These three agents are discussed in the “Current Therapies” section. The majority of emerging therapies that remain in development for NSCLC are biologics with varying mechanisms of action and targets. Current data suggest that any successful new drugs will not replace current therapies but rather will be used in addition to established chemotherapy regimens for Þrst- and second-line treatments and as stand-alone drugs in third-line treatment. In many cases, their clean toxicity proÞles and differing mechanisms of action suggest the potential to add more than one biologic to a chemotherapy regimen. Table 8 lists emerging therapies currently in development for NSCLC. Epidermal Growth Factor Receptor Inhibitors Overview. The epidermal growth factor receptor (EGFR) family is a group of four structurally similar growth factor receptors with tyrosine-kinase activity (HER-1/erb-B1, HER-2/erb-B2, HER-3/erb-B3, and HER-4/erb-B4) that is overexpressed in approximately 40–80% of NSCLC tumors (Sridhar SS, 2003; Raben D, 2005). Two EGFR inhibitors have been successfully launched for treatment of NSCLC: geÞtinib (AstraZeneca’s Iressa) and erlotinib (OSI/Genentech/Roche’s Tarceva). Of the agents remaining in clinical development, cetuximab (ImClone Systems/Merck KGaA/Bristol-Myers Squibb’s [BMS’s] Erbitux), which is in Phase III trials, is the most advanced. Trastuzumab (Genentech’s Herceptin) has been investigated in Phase II trials in NSCLC, but experts are unenthusiastic, given the low expression of its target, the human epidermal growth factor2 (HER-2) protein, on NSCLC cells. ABX-EGF (Abgenix/Amgen) remains in

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TABLE 8. Emerging Therapies in Development for Non-Small-Cell Lung Cancer, 2005 Compound

Development Phase

Epidermal growth factor receptor inhibitors Cetuximab (Erbitux) United States II Europe III Japan — Angiogenesis inhibitors Bevacizumab (Avastin) United States Europe Japan AE-941 (Neovastat) United States Europe Japan Thalidomide (Thalomid) United States Europe Japan Antisense oligonucleotides Oblimersen (Genasense) United States Europe Japan Modified cytotoxics PG-TXL United States Europe Japan Vinflunine (Javlor) United States Europe Japan Oral topotecan (Hycamtin) United States Europe Japan

Marketing Company

Bristol-Myers Squibb/ImClone Systems Merck KGaA —

III PC —

Genentech Roche —

III — —

AEterna Laboratories — —

III III —

Celgene Celgene —

III — —

Genta — —

III III I

Cell Therapeutics Cell Therapeutics Chugai

— III —

— Pierre Fabre —

III III —

GlaxoSmithKline GlaxoSmithKline/Merck Nihon Kayaku

Selective apoptotic antineoplastic drugs Exisulind (Aptosyn) United States II/III Europe — Japan —

OSI Pharmaceuticals — —

Retenoid X receptor agonists Bexarotene (Targretin [LGD-1069]) United States III Europe III Japan —

Ligand Pharmaceuticals Ligand Pharmaceuticals —

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TABLE 8. (continued) Compound

Development Phase

Marketing Company

Radiosensitizers RSR13 (Efaproxiral/Efaproxyn/Revaproxyn) United States III Europe — Japan —

Allos Therapeutics — —

Vaccines IGN-101(Edrecolomab) United States Europe Japan

— III —

— Aphton —

Novel alkylating agents TLK-286 (Telcyta) United States Europe Japan

III — —

Telik — —

PC = Preclinical (includes discovery).

Phase II trials for NSCLC, as does AstraZeneca’s ZD-6474. Amgen and Abgenix also have an anti-EGFR monoclonal antibody (MAb), panitumumab, in Phase II development for NSCLC, but lack of data precludes its discussion here. Mechanism of Action. Upon binding to a cognate ligand (e.g., epidermal growth factor [EGF] or transforming growth factor [TGF]-alpha, both of which may be overexpressed in some tumors), an EGFR undergoes dimerization, internalization of the receptor-ligand complex, and tyrosine autophosphorylation (Arteaga CL, 2002). These events ultimately trigger a cascade of intracellular responses that augment cell proliferation and survival, inhibit apoptosis, and potentially promote metastasis. Evidence suggests that overexpression of EGFR and overactivity of its downstream signal transduction pathway are signiÞcant factors in the growth of tumors in a subset of NSCLC patients. EGFR inhibitors effectively compete with cognate ligands for binding of an EGFR, thereby preventing tyrosine kinase activation and inhibiting tumor growth. Cetuximab. Cetuximab (ImClone Systems/Merck KGaA/BMS’ Erbitux) is a chimeric (part human/part mouse) anti-EGFR IgG (immunoglobulin G) approved for treatment of colorectal carcinoma in the United States and the United Kingdom and in Phase III trials for Þrst-line treatment of NSCLC in Europe. Plans for Phase III trials for second-line treatment of NSCLC in the United States were announced in late 2004. Cetuximab competitively binds to the extracellular portion of the EGFR, thereby preventing tyrosine kinase activation and inhibiting the growth of cancer cells that express this receptor. To date, Phase II data on cetuximab’s efÞcacy in NSCLC have been not been convincing enough to warrant its entry into the NSCLC market. The Lung

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Cancer Cetuximab Study (LUCAS) evaluated cisplatin and vinorelbine, with or without cetuximab, in Þrst-line treatment of 61 chemotherapy-naive patients with advanced NSCLC (Gatzemeier U, 2003). Inclusion of cetuximab increased tumor response rates signiÞcantly: 53% response with cetuximab versus 32.3% without. A further study, presented at the 2004 American Society of Clinical Oncology (ASCO) meeting, randomized 86 chemotherapy-naive patients with late-stage NSCLC (92% in stage IV) who were previously screened for EGFR expression, to treatment with vinorelbine/cisplatin with or without cetuximab (Rosell R, 2004). Preliminary assessment of the data revealed that the difference in response rates between treatment arms was not signiÞcant: 31.7% in the vinorelbine/cisplatin/cetuximab arm, compared with 20% in the vinorelbine/cisplatin arm. Time to progression also was not signiÞcantly different: 4.7 and 4.2 months, respectively. Patients receiving cetuximab demonstrated more asthenia and fatigue and higher incidence of infection and acneiform rash, but there was no signiÞcant difference between the two arms in gastrointestinal toxicity or leukopenia. Cetuximab is also being used in combination with gemcitabine and carboplatin in Þrst-line treatment of chemotherapy-naive stage IV patients; in one trial, 10 of 35 patients treated achieved a partial response (2 unconÞrmed) and 14 achieved stable disease, with a median time-to-progression of 166 days and median survival of 277 days (Robert F, 2003). SigniÞcant treatment-related adverse events were anemia (grade III, 11%); thrombocytopenia (grades III and IV, 54%); leukopenia (grade IV, 20%); and infection (grade III, 9%). Cetuximab-related toxicities included an acneiform rash (grades I and II, 80%; grade III, 20%); dry skin (grades I and II, 40%; grade III, 9%); fatigue/malaise (grades I and II, 20%; grade III, 6%); and one grade III allergic reaction. Cetuximab has been investigated as a second-line therapy, in combination with docetaxel, in chemotherapy-refractory patients with advanced NSCLC. In a Phase II trial in 47 patients, 28% demonstrated partial response, and 17% had stable disease; time to progression was 89 days (Kim ES, 2003). Grade III toxicities included infection (21%), fatigue (21%), and acneiform rash (19%); four patients experienced an allergic reaction that required discontinuation. Final analysis of duration of response and survival data is ongoing. The BMS/ImClone/Merck Phase III program supporting cetuximab in NSCLC is extensive. It includes the EMR-046 study, which plans to enroll 1,050 patients with metastatic NSCLC in Europe, Asia PaciÞc, and Latin America for Þrstline treatment with a platin/vinorelbine combination, with or without cetuximab. Another planned U.S. trial aims to treat 800 patients with recurrent NSCLC with a standard chemotherapy regimen plus docetaxel, pemetrexed, or cetuximab. ImClone and BMS are also enrolling patients in a Phase II trial to assess the efÞcacy and safety of cetuximab as a single-agent therapy for patients with recurrent or progressive NSCLC who have failed one or more chemotherapy regimens, including platinum-based combinations. Results presented at the 2005 ASCO meeting revealed a response rate of 3.3% and a disease control rate of

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28%. Median time to progression was 2.3 months and median survival was 8.1 months. The 6-month survival rate was 63% and the one-year survival was 43%. The most commonly reported toxicities were rash in 77% of patients; grade III/IV toxicities included dyspnea (15%), fatigue (13.6%), infection (9%), and gastrointestinal side effects (6%) (Lynch TJ, 2004[b]; Lilenbaum P, 2005). If launched for second-line therapy, cetuximab will face stiff competition from the small-molecule EGFR inhibitor erlotinib (see the “Current Therapies” section), which is now supported by impressive Phase III data. If launched for Þrst-line therapy, competition would come from the anti-VEGF monoclonal antibody bevacizumab (discussed in a later section). Cetuximab’s and bevacizumab’s mechanisms of action have the potential to be synergistic, but the high cost of this combination is likely to severely limit patient share. Angiogenesis Inhibitors Overview. Angiogenesis (the formation of new blood vessels) is critical to the survival and metastasis of tumors; it allows tumors to develop their own vascular supply, thus enabling them to grow and invade healthy tissue. Numerous studies have demonstrated that higher degrees of tumor vascularization portend poorer prognosis in NSCLC and other cancers. Anti-angiogenic compounds in late-stage development for advanced NSCLC include matrix metalloproteinase (MMP) inhibitors, MAbs, and thalidomide. Calcium-channel blockers, such as L-651582, also remain under investigation for their anti-angiogenic qualities. However, although L-651582 has been in Phase III trials for NSCLC since 1999, no clinical data on its efÞcacy in this indication have been released. The MMP inhibitor prinomastat (PÞzer) failed to show an efÞcacy beneÞt when added to conventional treatment in Phase III NSCLC trials (Bissett D, 2005; Leighl NB, 2004). Several anti-angiogenic molecules with various mechanisms of action remain in Phase II development—for example, squalamene (Genaera), which has anti-angiogenic, pro-apoptotic and VEGF antagonistic properties. Novartis has announced a Phase II trial of its VEGF receptor tyrosine kinase inhibitor vatalanib. Bayer’s sorafenib remains in Phase II development for NSCLC, as does Abbott’s ABT-510. Lack of data precludes discussion of these molecules at this stage of their development. Mechanism of Action. Angiogenesis inhibitors reduce the neovascularization of tumors, thereby depriving tumors of the nutrients required to sustain uncontrolled growth. Tumor cells may continue to show high rates of replication, but many more cells die in the nonvascularized, hypoxic state than in a tumor with abundant vascularization. Bevacizumab. Bevacizumab (Genentech/ Roche’s Avastin) is in Phase III trials for treatment of NSCLC in the United States, where it was launched in February 2004 for use in combination with 5-ßuorouracil (5-FU)-based chemotherapy as a

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treatment for patients with Þrst-line metastatic cancer of the colon. Bevacizumab is also approved for treatment of patients with previously untreated metastatic colorectal cancer in Europe, where it remains in the discovery phase for NSCLC with parent company Roche. Bevacizumab is a chimeric, antivascular endothelial growth factor (AVEGF) MAb. VEGF is a multifunctional cytokine released by tumors that plays a central role in tumor angiogenesis. It is a potent instigator of endothelial cell migration from neighboring capillaries to the tumor site, where the cells replicate to form new blood vessels. This growth factor also increases vascular permeability, facilitating migration to the tumor site of other cells that support the formation of tumor connective tissue and thereby foster tumor growth. In March 2005, Genentech released data from a Phase II/III trial of bevacizumab in combination with carboplatin/paclitaxel compared with carboplatin/paclitaxel alone. Conducted in collaboration with the Eastern Cooperative Oncology Group (ECOG), this study enrolled 878 patients with previously untreated, advanced NSCLC. Interim analysis revealed that patients in the bevacizumab-containing arm had a signiÞcantly longer median overall survival (12.5 months) than patients receiving the standard regimen (10.2 months) (Genentech, 2005). Although patients with any prior signiÞcant pulmonary bleeding and patients with squamous-cell carcinoma were excluded from the trial, fatal pulmonary bleeding did occur in more patients receiving bevacizumab than in patients receiving the standard regimen. In a second planned interim analysis announced at the 2005 ASCO meeting (Sandler AB, 2005), the bevacuzimab/paclitaxel/carboplatin regime was signiÞcantly superior to paclitaxel/carboplatin alone. The response rate was 10% for the standard regimen compared to 27% in the bevacizumab-containing regimen; progression-free survival was 4.5 months versus 6.4 months, respectively, and median survival 10.2 months versus 12.5 months, respectively. The bevacizumab-containing arm led to nine treatment-related deaths, Þve due to hemoptysis. There were two treatmentrelated deaths in the standard regimen arm. These positive results led to the ECOG adopting bevacuzimab/paclitaxel/carboplatin as the new treatment standard in this patient population. Previous data from a Phase II trial of 99 treatment-naive patients with stage IIIB or IV NSCLC, who were randomized to treatment with one of two dose levels of bevacizumab in combination with carboplatin/paclitaxel or to treatment with chemotherapy alone, reported four hemoptysis-related deaths (DeVore RF, 2000). This phenomenon appeared to be associated with squamous cell cancers located near large blood vessels. However, a subsequent analysis that measured survival in the advanced non-squamous-cell lung cancer patients found that 53 patients who had received either low- or high-dose bevacizumab in combination with carboplatin/paclitaxel achieved a median survival of 76 weeks, a signiÞcant improvement on the median survival of 34–53 weeks achieved by chemotherapy alone (Johnson DH, 2001). Bevacizumab is under evaluation for early-stage NSCLC in a neoadjuvant setting. A trial sponsored by the National Cancer Institute (NCI) is investigating

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neoadjuvant bevacizumab, paclitaxel, and carboplatin in patients with stage IB, II, or IIIA resectable NSCLC. In addition, bevacizumab is being tested in combination with erlotinib, an EGFR tyrosine kinase inhibitor. A Phase I/II study enrolled 40 patients with either stage IIIB, stage IV, or recurrent NSCLC who had been treated with at least one prior chemotherapy regimen. A dose-ranging period established erlotinib (150 mg/day) plus bevacizumab (15 mg/kg) as a safe dose; 34 patients were then treated with this regimen. Preliminary data indicate no pharmacokinetic interaction between erlotinib and bevacizumab. The median overall survival and time-to-progression were 9.3 and 4.6 months, respectively; 17.5% of patients had partial responses, 5% had minor responses, and 35% had stable disease. Mild to moderate rash, diarrhea, and proteinuria were the most commonly reported side effects (Sandler AB, 2004). When bevacizumab is launched for NSCLC, it is likely to have a restricted label. Patients with prior bleeding complications, central lesions, or squamous cell histology (approximately one-third of the NSCLC population) will be ineligible for treatment with this drug. AE-941. AE-941 (Neovastat) is a matrix metalloproteinase inhibitor (MMPI) under development by AEterna Laboratories. Originally one of many MMPIs in late-stage development for NSCLC, AE-941 is the only one that remains in Phase III trials in the United States and Canada. MMPs are a family of naturally occurring enzymes that are critical in angiogenesis and metastasis because they degrade components of the extracellular matrix, such as collagen, and allow cell migration into tissues. MMP degradation products of collagen may also trigger cell proliferation, leading to tumorigenesis; therefore, inhibiting the activity of MMPs may help to control tumor metastasis. AE-941 is an extract from shark cartilage that has activity against MMP-2 and MMP-12, commonly expressed in lung cancer. In January 2000, AEterna initiated a Phase III clinical trial of AE-941 in combination with chemoradiotherapy (CRT) to treat stages IIIA and IIIB NSCLC; patients are to receive platinum-based induction chemotherapy (IC) followed by concurrent CRT with or without AE-941. This trial, sponsored by the NCI and in collaboration with the Radiation Therapy Oncology Group (RTOG) and the Community Clinical Oncology Program (CCOP), aims to enroll 760 patients over Þve years, with Þnal results expected in 2006. Patients with unresectable stage III NSCLC (PS < 2) are being randomized to receive carboplatin (AUC 6) and paclitaxel (200 mg/m2 ), or cisplatin (75 mg/m2 ) and vinorelbine (30 mg/m2 ), followed by chemoradiation therapy with AE-941 (120 mL bid) or placebo. Accrual to this study continues and the data have not yet been unblinded, but blinded toxicity data were made available on 179 patients during the chemotherapy cycle and 172 subjects during radiation therapy and AE-941 or placebo at the 2005 ASCO meeting (Lu C, 2005). The most common grade III/IV hematologic toxicities were granulocytopenia (in 22% and 1% of patients, respectively), leukopenia (in 2%/4%), thrombocytopenia (1%/3%), anemia (1%/0%), and neutropenic fever

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(5%/2%). The most common grade III or IV nonhematologic toxicities during chemotherapy were fatigue (4%), hyperglycemia (3%), and dyspnea (2%). The most common grade III or IV nonhematologic toxicities during chemoradiation therapy were esophagitis (17%) and dyspnea/pneumonitis (5%). A combined partial and complete response rate of 35% after chemotherapy and 39% after radiotherapy was observed. Stable disease was observed in 57% and 48% of patients, respectively, and progressive disease in 8% and 13%, respectively. A retrospective analysis of a subgroup of 47 patients with unresectable NSCLC who were treated with oral AE-941 at doses higher than 2.63 mL/kg/day found that this group achieved a signiÞcant increase in survival time: 6.15 months versus 4.17 months (Franquois B, 2001). The most frequent adverse events were nausea (7%), vomiting (3%), dyspepsia (2%), and anorexia (2%). Thalidomide. Thalidomide (Celgene’s Thalomid), originally developed as an oral sedative, has enjoyed renewed interest since the scientiÞc community discovered that this drug has anti-angiogenic properties. Thalidomide is approved for treatment of cutaneous manifestations of moderate-to-severe erythema nodosum leprosum (ENL) in the United States and for multiple myeloma in Australia; it is currently in Phase III trials for NSCLC in the United States and Europe. Development is supported by Celgene, with additional support from various national cancer agencies, such as the NCI and London Cancer Group. Although the precise mechanism of action is poorly understood, thalidomide is thought to inhibit angiogenesis by downregulating pro-angiogenic factors such as basic Þbroblast growth factor (bFGF) and VEGF (Li X, 2003). Other mechanisms have also been suggested, such as the induction of peroxisome proliferatoractivated receptor gamma (PPAR-gamma) (DeCicco KL, 2004). Thalidomide has demonstrated activity against various other advanced malignancies, including breast, prostate, melanoma, colon, and prostate cancers (Hirschfeld SH, 2000). Thalidomide is being assessed in NSCLC as part of various Þrst-line combination regimens for nonresectable stage IIIA, stage IIIB, and stage IV disease. ECOG is currently conducting Phase III trials of thalidomide in nonresectable stage IIIB NSCLC patients. The trial is comparing carboplatin/paclitaxel and radiotherapy with or without thalidomide. The investigators plan to enroll a total of 588 patients in this study. In a pilot and safety study of carboplatin/paclitaxel and thalidomide in nine patients with stage IIIA, IIIB, or stage IV NSCLC, some of whom had received previous chemotherapy, no responses were observed, although six patients achieved stable disease (Merchant JJ, 2000). Another small-dose escalation trial has evaluated the combination of docetaxel (75 mg/m2 ) with thalidomide in 14 patients from a planned accrual of 35. The dose of thalidomide was increased from 50 mg/day to a maximum of 200 mg/day. Patients also received warfarin or another anticoagulant. Stable disease was reported in 21.3% of patients; median time to progression was 11.5 months. Grade III or IV neutropenia was observed in 58% of patients. Grade I and II toxicities included fatigue, constipation, and alopecia (Seidler CW, 2004).

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In addition, several Phase II trials are currently investigating the use of carboplatin, irinotecan, and thalidomide in stage IIIB and stage IV NSCLC patients. Preliminary results from 34 patients, presented at the 2004 ASCO meeting, were rather disappointing: of the 23 patients who were evaluable, 22% displayed a partial response, and 52% had stable disease. Median time-to-progression was 3.6 months, and median survival time was 7.3 months. The most frequent toxicities were neutropenia, fatigue/malaise, and nausea/vomiting (Miller AA, 2004). Recruitment is ongoing. Another trial in chemotherapy-naive patients with stage IIIB or IV NSCLC that was reported at ASCO 2004 showed that of eight evaluable patients, two had experienced thromboembolic complications, and one had experienced therapy-related pneumonitis that required removal from the trial. Median time to progression was 4.3 months. One patient achieved a partial response, and three had stable disease (Flora DB, 2004). The side effects of thalidomide therapy are generally manageable; however, potentially irreversible peripheral neuropathy can occur with prolonged treatment. Pregnancy tests are required before and during treatment, and birth control is mandatory for both men and women who are taking this drug. As part of its safety monitoring program, Celgene requires physicians to participate in a registration program before prescribing thalidomide. Antisense Oligonucleotides Overview. Physicians once believed that antisense oligonucleotides held promise for the treatment of NSCLC; however, disappointing trial results have dampened enthusiasm considerably in recent years. Isis Pharmaceuticals/Eli Lilly’s ISIS-3521 (AfÞnitak/Aprinocarsen) was in Phase III development; after disappointing data from two major trials of the agent in combination with carboplatin/paclitaxel and cisplatin/gemcitabine, Isis issued a statement in March 2005 that it would not invest further in developing this agent for treatment of NSCLC. Currently, one antisense agent remains in Phase III development for NSCLC—Genta’s oblimersen (Genasense). Lorus Therapeutics’ GTI-2040, an anti-R2 ribonucleotide reductase mRNA, is listed as being in Phase II development for NSCLC, but it is not discussed in detail here owing to a lack of data. Mechanism of Action. Antisense oligonucleotides are recombinant RNA molecules that bind to complementary messenger RNA (mRNA) sequences within the cytoplasm of cells. A target gene is effectively “silenced” by blocking the translation of the mRNA and generation of the corresponding protein. In this way, antisense molecules can block the expression of undesirable genes and their proteins. Oblimersen. Oblimersen (Genasense) is being developed by Genta and is in Phase III trials in the United States for treatment of NSCLC. The drug was formerly being codeveloped with SanoÞ-Aventis; in May 2004, however, following disappointing results in trials for metastatic melanoma, Genta withdrew its

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new drug application (NDA) for the use of oblimersen plus dacarbazine in the treatment of patients with advanced melanoma or chronic lymphocytic leukemia (CLL). SanoÞ-Aventis announced that it would terminate its involvement in May 2005. Oblimersen is an antisense oligonucleotide complementary to BCL-2, an antiapoptosis protein that is overexpressed in multiple cancers, including NSCLC. A randomized Phase III trial was initiated in November 2001 to compare oblimersen plus docetaxel versus docetaxel alone in patients with recurrent stage IIIB or stage IV NSCLC who had failed previous chemotherapy. In August 2004, Genta announced that enrollment of 298 patients was complete. The study will evaluate whether the combination of oblimersen and docetaxel is superior to docetaxel alone for increasing length of survival, anticancer response, and quality of life. Final results are expected in late 2006. Preclinical data released in April 2003 demonstrate that oblimersen is active when used alone or in combination with vinorelbine. Both drugs show antitumor activity and incremental improvements in survival when administered alone, but they show signiÞcantly greater antitumor response rate and lower incidence of distant metastases when used in combination. Additional data presented at the 94th meeting of the American Association for Cancer Research (AACR) in July 2003 demonstrate that the oblimersen/vinorelbine combination delayed the establishment of tumors to a greater extent than either compound alone and that growth of H460 tumors was inhibited more by combination treatment than by monotherapy (Hu YP, 2003). Modified Cytotoxics Overview. Cytotoxic chemotherapeutic drugs remain the most efÞcacious method of debulking tumors regardless of their origin. However, these drugs are highly toxic, and their use is limited in patients unable to tolerate adverse effects—for example, patients with advanced disease, the elderly, or patients with poor performance status (PS). Reducing the toxicity proÞle of wellestablished agents by chemical modiÞcation may increase the treatable patient population and the maximum tolerated dose. ModiÞed cytotoxic agents promise advantages over currently available agents through improved pharmacokinetics: drug targeting enhances antitumor activity, while systemic toxicity is reduced. Novel formulations of cytotoxics in Phase II development for NSCLC—but not discussed here because of a lack of information—include a liposomal formulation of cisplatin in development by Regulon and an intravenous (IV) emulsion formulation of paclitaxel being developed by Sonus Pharmaceuticals. Mechanism of Action. Cytotoxic drugs exert their activity by targeting processes within a cell that are crucial for survival, such as DNA synthesis, replication, and repair. Their mechanism of action is not tumor-speciÞc but preferentially targets cells with high proliferative rates, such as malignant cells.

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This section focuses on novel formulations of paclitaxel and vinßunine. Therefore, the discussion here is limited to the mechanisms of action of these two drugs. Paclitaxel is the prototype of the taxane family of antitumor drugs and the Þrst natural product shown to stabilize microtubules formed during mitosis. Suppression of microtubule dynamics and disassembly blocks mitosis at the metaphase/anaphase transition and induces cell death. Vinßunine is a vinca alkaloid that acts by binding to microtubular proteins of the mitotic spindle, leading to mitotic arrest or cell death.

PG-TXL. PG-TXL (Xyotax, paclitaxel poliglumex, CT-2103, CHC12103), a poly-(L-glutamic acid)-conjugate of paclitaxel, is being developed by Cell Therapeutics under license from the M.D. Anderson Cancer Center and in collaboration with Chugai. The agent is currently under investigation in Phase III trials in the United States and Europe and has completed Phase I trials in Japan. However, preliminary analyses of the key Phase III trials in NSCLC, released in March and May 2005, indicate that all three trials missed their primary end points. Paclitaxel is highly successful in the treatment of a range of solid tumors. For this reason, there has been much interest in developing new formulations and analogues to address the problems of paclitaxel administration. The presence of the poly-glutamic acid (PG; a biodegradable, water-soluble polymer of glutamic acid) in PG-TXL makes the conjugate 80,000 times more water soluble than native paclitaxel. This enhanced aqueous solubility eliminates the need for cremophor (the solubilizing agent required for paclitaxel administration) and results in a convenient ten-minute infusion with fewer allergic reactions. Cell Therapeutics reports that because paclitaxel is so tightly bound to the PG backbone, little free drug is found in the circulation, potentially reducing the harmful side effects on normal tissues. The maximum tolerated dose of PG-TXL is therefore twice that of standard paclitaxel. PG-TXL also bypasses the multidrug-resistance pump (MDR) and is engulfed by tumor cells via the endocytic pathway; it has proved effective on tumor cells that are resistant to standard paclitaxel. Phase I and II data showed good tolerability for this drug and encouraging responses in NSCLC patients, including patients with poor PS and elderly patients (Bolton MG, 2003; Norton MS, 2003). PG-TXL is under investigation for NSCLC in three major Phase III trials. One trial, Selective Targeted EfÞcacy in Lung Cancer, Lower Adverse Reactions 3 (STELLAR 3), is comparing the use of carboplatin in combination with either PG-TXL or paclitaxel in Þrst-line treatment of 400 poor PS patients (PS 2). Although the primary endpoint of superiority over paclitaxel/carboplatin in terms of one-year survival was not met, an intention-to-treat analysis presented in March 2005 revealed that PG-TXL/carboplatin was “statistically non-inferior” to paclitaxel/carboplatin. Survival data presented at the 2005 ASCO meeting survival were not signiÞcantly different between treatment arms, although patients who received PG-TXL/carboplatin had signiÞcantly less neutropenia, hair loss, and cardiac events, and a reduction in other side effects, including muscle and

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joint pain, and an overall reduction in neurological toxicities, compared with patients who received the paclitaxel/carboplatin regimen (Langer CJ, 2005). In April 2005, Cell Therapeutics announced that PG-TXL failed to show improved survival, compared with conventional chemotherapy regimes, in the STELLAR 4 study. This randomized Phase III trial is comparing PG-TXL with gemcitabine or vinorelbine in chemotherapy-naive patients who have stage IIIB, stage IV, or recurrent NSCLC. Results indicate that median survival of poor-PS patients who were receiving PG-TXL was 7.3 months, compared with 6.6 months for patients receiving either gemcitabine or vinorelbine (p = nonsigniÞcant). Two-year survival was 15% in the PG-TXL arm and 10% in the gemcitabine and vinorelbine arms (p = nonsigniÞcant). However, PG-TXL was associated with signiÞcant reductions in severe hematologic toxicities, compared with the other regimens, including reductions in anemia, neutropenia, and thrombocytopenia. GI side effects, including nausea and vomiting, were also reduced, but the incidence of grades III and IV neuropathy increased in patients receiving PG-TXL. The dose of PG-TXL used in this study, which originally aimed to recruit approximately 370 patients, was lowered from 235 mg/m2 to 175 mg/m2 in October 2003, after an analysis by an independent data-monitoring committee found that some patients had developed early neutropenia-related toxicities. PG-TXL is also under investigation as a second-line therapy versus docetaxel in patients with progressive NSCLC. STELLAR 2, a Phase III trial that was initiated in October 2002, aims to accrue 840 patients, and is examining overall survival, safety, and response. Preliminary analysis of the data reveal that patients in both arms had a median survival of 6.9 months. However, signiÞcantly fewer hematologic side effects were observed in the PG-TXL arm; other side effects that were also reduced included hair loss, fatigue, asthenia, mucositis, and ocular toxicity. Vinflunine. Vinßunine (Javlor) (Figure 15), which is being developed by Pierre Fabre in collaboration with Bristol-Myers Squibb (BMS), is in Phase III clinical trials for NSCLC in Europe, South Africa, Singapore, and Mexico. BMS will be responsible for the development and marketing of vinßunine in the United States and Japan; Pierre Fabre retains marketing rights in Europe. Vinßunine is a semisynthetic vinca alkaloid obtained through selective introduction of two ßuorine atoms at the 20’ position of vinorelbine. In animal models, vinßunine has greater antitumor activity than vinorelbine (BonÞl RD, 2002). In June 2003, Pierre Fabre launched Phase III trials of vinßunine in NSCLC and in bladder cancer. The company aimed to recruit 550 NSCLC patients to a randomized trial to investigate single-agent vinßunine in the second-line setting. The primary objective is to compare progression-free survival following vinßunine treatment with survival following docetaxel treatment. In a Phase II study of vinßunine monotherapy (320 mg/m2 every 3 weeks) in 63 patients with advanced NSCLC who had failed a previous platinum-based regimen, 8% of patients displayed a partial response lasting 5.8 months, and 53% displayed stabilization. Progression-free survival was 2.6 months. Grades III and IV neutropenia were

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N

CF2CH3

N N H CH3O2C CH3O

CH3 N

OCOCH3 HO CO2CH3 CH3

FIGURE 15. Structure of vinflunine.

observed in 25.4% and 23.8% of patients, respectively; grades III and IV anemia and thrombocytopenia were rare. Grade III or greater nonhematologic toxicities were myalgia (15.9%), fatigue (6.4%), stomatitis (4.8%), and constipation (7.9%) (Bennouna J, 2004). In a Phase II trial in bladder cancer, patients received 350 mg/m2 vinßunine every 21 days; the dose was decreased to 320 mg/m2 following an episode of fatal neutropenic sepsis. More fatal episodes of neutropenic sepsis occurred at this dose, and further dose reductions were made. Median dose intensity was 101.9 mg/m2 /week. Among 51 evaluable patients, there were 9 partial responses, 26 cases of stable disease, and 16 cases of disease progression (Bui B, 2003). At the 320 mg/m2 dose, grades III to IV neutropenia, leukopenia, anemia, thrombocytopenia, constipation, myalgia, and vomiting occurred in 67%, 45%, 14%, 6%, 12%, 4%, and 6% of patients, respectively. In a Phase I/II study of an experimental vinßunine/cisplatin combination, presented at the 2004 ASCO meeting, 16 of 49 evaluable patients had conÞrmed responses and 22 had stabilizations. Grades III and IV neutropenia were reported in 43% of the 237 cycles analyzed, and anemia was reported in 7%. Febrile neutropenia was reported in 5 patients, and there were two neutropenic infections. The most common grades III and IV nonhematologic toxicities were fatigue (15.1% of patients), abdominal pain (9.4%), nausea (3.8%), vomiting (3.8%), and constipation (5.7%) (Ramlau R, 2004). Oral Topotecan. Topotecan (GlaxoSmithKline/Merck/Nihon Kayaku’s Hycamtin) (Figure 16) is a water-soluble, semisynthetic analogue of camptothecin. An intravenous formulation is approved in the United States and Japan as monotherapy for patients with recurrent small-cell lung cancer (SCLC). An oral formulation of topotecan is under Phase III investigation for NSCLC in both the Þrst- and second-line settings in the United States and Europe. Topotecan is a topoisomerase I inhibitor that binds to the topoisomerase I-DNA complex and prevents religation of DNA single-strand breaks that occur naturally during DNA synthesis. A Phase II trial investigated the effects of oral topotecan (2.3 mg/m2 /day), for 5 days every 21 days for up to six cycles, in 30 chemotherapy-na¨õve patients

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CH3 N CH3 HO

O N N O HO H3C

O

FIGURE 16. Structure of topotecan.

with stage III or IV NSCLC (PS 0–2). No patients achieved complete or partial responses to treatment. 43% of patients had stable disease, with a median survival of 39.9 weeks and a one-year survival of 33.3%. Median time to progression was 12.3 weeks. Forty percent of patients experienced grade III/IV neutropenia and 16.6% had grade III/IV anemia. There were two episodes of grade III/IV thrombocytopenia (White SC, 2000). Results from an open-label, Phase III study comparing oral topotecan to intravenous docetaxel in 829 patients with recurrent stage III/IV NSCLC were reported at the 2005 ASCO meeting (Ramlau R, 2005). Patients were treated with either oral topotecan (2.3 mg/m2 /day on days 1–5 for 21 days) or IV docetaxel (75 mg/m2 /day every 21 days). Median survival and one-year survival for the topotecan and docetaxel groups were 27.9 versus 30.7 weeks and 25.1% versus 28.7%, respectively (p = nonsigniÞcant). Median time to progression was 11.3 versus 13.1 weeks, respectively (p = 0.02). The most common grade III/IV hematologic toxicities were neutropenia (23%/25% versus 24%/36%), thrombocytopenia (21%/4% versus 3%/4%) and anemia (18/7 versus 5/5). Selective Apoptotic Antineoplastic Drugs Overview. OSI Pharmaceuticals (formerly Cell Pathways) is pioneering a new approach to treatment and prevention of cancer with the development of agents known as selective apoptotic antineoplastic drugs (SAANDs). These drugs aim to induce apoptosis (programmed cell death) selectively in precancerous and cancerous cells. Exisulind (Aptosyn) is OSI’s Þrst-generation compound in this new class of drugs, but a more potent, second-generation SAAND is in development. In addition to inducing apoptosis, this more potent agent, an analogue of exisulind known as CP-461, has antiproliferative activity mediated through microtubule disruption and spindle formation. Preclinical data presented at the AACR meeting in April 2002 showed that in a rat model of human NSCLC, oral CP-461 achieved an improvement in survival equivalent to survival achieved by docetaxel (Whitehead CM, 2002). CP-461 is in Phase I/II investigations for

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CLL, prostate cancer, renal cancer, and melanoma but is not currently in clinical development for NSCLC. Other pro-apoptotic drugs in development for NSCLC are Millennium’s proteasome inhibitor bortezomib, which is approved for second-line treatment of multiple myeloma and is in Phase II development for NSCLC. Cambridge Antibody Technology and Human Genome Sciences have a pro-apoptotic monoclonal antibody, mapatumumab, in Phase II development for NSCLC. However, very few data on these two agents in NSCLC are available at present. Mechanism of Action. SAANDs exert their apoptotic activity by inhibiting cyclic GMP phosphodiesterase (cGMP PDE), an enzyme that is overexpressed in precancerous and cancerous cells. Inhibition results in an increase in cellular cGMP, a second messenger molecule that can induce apoptosis through a series of steps involving the activation of protein kinase G, decreased β-catenin, and activation of caspases. Exisulind. Exisulind (Aptosyn), under development by OSI Pharmaceuticals, is in Phase III trials in NSCLC in the United States in combination with a variety of chemotherapeutic agents. Preclinical research indicates that exisulind’s activity is directed at cGMPPDE and is independent of other apoptosis pathways, such as p53, Bcl2, BAX (a proapoptotic member of the Bcl-2 family), and the cyclooxygenase (COX) enzymes. In June 2004, OSI announced disappointing results from a Phase III trial investigating the combination of exisulind and docetaxel versus docetaxel and placebo in 610 previously treated patients with metastatic disease. The study did not meet its primary end point of improving overall survival, nor did it meet its secondary end points of improvements in one-year survival, progression-free survival, or response rate; median survival and the one-year survival rate were 6.9 months and 30.7%, respectively, in the exisulind/docetaxel arm and 6.9 months and 29.5%, respectively, in the placebo/docetaxel arm. More disappointing results for exisulind combination therapy were presented at the 2004 ASCO meeting. In an NCI-sponsored Phase II study, 58 patients with advanced NSCLC (grade IIIB with plural effusion or grade IV, PS 0–2) were treated with exisulind in combination with carboplatin and gemcitabine (Masters GA, 2004). Of the evaluable patients, 2 had complete responses and 6 had partial responses; the overall response rate was 17%. Median progression-free survival was 4.7 months, and median overall survival was 7.1 months. Principal toxicities of grade III or above were neutropenia (56% of patients), thrombocytopenia (30%), nausea (14%), and fatigue (18%). In a Phase I/II trial, 15 patients were treated in a dose-escalation phase with exisulind (from 125 mg bid up to 250 mg bid) in combination with gemcitabine (1,250 mg/m2 ). Phase II plans are to recruit 24 patients for treatment with 250 mg bid exisulind. In Phase I, the drug was well tolerated; dose-limiting toxicities were grade III fatigue and grade III constipation (one patient each). Median and

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one-year survival were ten months and 40%, respectively. Myelotoxicities were mild. Of the 18 patients evaluable in Phase II, 9% had a partial response and 44% had stable disease (Hoang T, 2004). The trial is ongoing. A similar study is assessing a combination of exisulind and vinorelbine in elderly patients with advanced NSCLC (Traynor AM, 2004). In the Phase I arm, presented at the 2004 ASCO meeting, dose-limiting toxicities included grade III constipation, grade III diarrhea, and grade IV neutropenic fever (one patient each). In the 15 currently evaluable patients from the Phase II arm, investigators observed grades III and IV treatment-related toxicities, including neutropenia (5 patients), leukopenia (4 patients), anemia (1 patient), fatigue (3 patients), dyspnea (2 patients), and constipation (1 patient). Clinical data have also been presented from Phase I/II trials investigating the response rate, survival rate, and toxicity of exisulind in combination with paclitaxel and carboplatin in previously untreated stages IIIB and IV NSCLC patients. The regimen consisted of up to six cycles of paclitaxel, carboplatin, and daily oral exisulind. Seven partial responses, ten stable diseases, and eight disease progressions were observed. Median time-to-progression was 166 days, median survival was 345 days, and one-year survival was 46%. Toxicity data from this study suggest an increased incidence of thromboembolic events with this three-drug combination, which had not previously been reported. Thromboembolic disease occurred in six patients, and there was one sudden death (no post mortem was carried out). Other grades III and IV toxicities included elevated liver enzymes (GGT)—which resolved after discontinuation of exisulind—neutropenia, neuropathy, and nausea/vomiting (Kang C, 2002; Villaßor VM, 2003). Retinoid X Receptor Agonists Overview. Retinoid X receptors (RXRs) are nuclear receptors that target and regulate multiple signaling pathways that have effects on metabolic function, cell differentiation, and cell death. Retinoids, or vitamin A analogues, are the natural ligands of RXRs; several natural and synthetic retinoids are being developed for the treatment of various diseases, including diabetes, psoriasis, and cancer. Mechanism of Action. RXRs are ligand-activated transcription factors that control expression of genes that modulate processes such as apoptosis, cell growth, and differentiation. Agents targeted against these receptors (either agonists or antagonists) can therefore impact downstream gene expression. RXRs have demonstrated an inhibitory effect on NSCLC cell growth in vitro (Brabender J, 2002). Bexarotene. Bexarotene (Targretin [LGD-1069]) (Figure 17), an oral RXR selective agonist, is being developed by Ligand Pharmaceuticals and is currently in Phase III clinical trials in the United States and Europe for NSCLC. The agent is approved for use in cutaneous T-cell lymphoma. Bexarotene is a synthetic retinoic acid derivative that binds to its target RXR with high afÞnity, showing little afÞnity for the retinoic acid receptor.

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CH2 H3C

CH3

C OH

H3C

CH3

CH3

C O

FIGURE 17. Structure of bexarotene.

Ligand is conducting two Phase III randomized clinical trials (Studies Providing Investigational Research in Targretin [SPIRIT I and SPIRIT II]) that are investigating the combination of bexarotene with chemotherapy in Þrst-line therapy in advanced-stage NSCLC. SPIRIT I is analyzing the outcomes of patients who receive cisplatin/vinorelbine with or without bexarotene; SPIRIT II is studying bexarotene in combination with carboplatin and paclitaxel. In September 2003, enrollment for both trials was complete and results were expected within one year. However, in March 2004, Ligand announced that survival data analysis would take 18 months, or 456 deaths. In March 2005, Ligand announced that both SPIRIT I and SPIRIT II failed to show a signiÞcant improvement in overall survival or projected two-year survival, the major end points of the studies. The data were also disclosed at the 2005 ASCO meeting (Blumenscheri GR, 2005; Jassem J, 2005). In SPIRIT II, median overall survival (254 days versus 277 days) and projected two-year survival (12.4% and 16.3%) was not statistically different between the bexarotene plus chemotherapy treatment arm and chemotherapy alone arm, respectively. Similar survival data and lack of efÞcacy of bexarotene were evident in SPIRIT I. In March 2005, Ligand also conÞrmed that bexarotene was in trials for secondline treatment of NSCLC as a monotherapy and in combination with erlotinib. Several Phase I/II trials have studied the effects of adding bexarotene to conventional dual combination chemotherapy regimens. In one trial, patients with stages IIIB or IV (PS 0–2) NSCLC were treated with a bexarotene (400 mg/m 2 )/ carboplatin/gemcitabine combination. Atorvastatin was also administered to prevent hypertriglyceridemia. The 41 evaluable patients had an overall response rate of 21%; 56% of patients were free from progression at six months (compared with 27% of historical controls; p = 0.03). Adverse events were grades III and IV hypertriglyceridemia in 44% of patients. Neutropenia was observed in 46% of patients and thrombocytopenia in 44% (Edelman MJ, 2004). Results from a Phase I/II evaluation of bexarotene in combination with carboplatin/paclitaxel in chemotherapy-naive, advanced-stage NSCLC patients revealed that of the 15 evaluable patients, 1 achieved complete response, 5 achieved partial responses, 5 achieved stable disease, and 4 experienced disease progression. The overall response rate of 40% was an improvement, compared with published data on carboplatin/paclitaxel alone. Phase I studies conÞrmed that bexarotene does not affect paclitaxel’s pharmacokinetics and is well tolerated in

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FIGURE 18. Structure of efaproxiral.

combination with paclitaxel/carboplatin in patients with advanced stage NSCLC (Jacobs CD, 2003). Bexarotene was also combined with vinorelbine/cisplatin in a Phase I/II study of 43 patients with advanced NSCLC. Median survival was 14 months in Phase II, with 32% of patients still alive after 24 months; the overall one-year survival rate was 63% (Khuri FR, 2001[b]). Radiosensitizers Overview. Radiosensitizers are used in conjunction with radiotherapy to increase its effectiveness. Several trials for these drugs are ongoing for various cancers. Mechanism of Action. Radiosensitizers are chemicals that potentiate the effect of ionizing radiation, often by restoring normal oxygen tension to a hypoxic tissue, such as a tumor. Because tissues with low oxygen content are less susceptible to damage, this increase in oxygen allows radiotherapy to be more effective. Efaproxiral. Efaproxiral (RSR-13, Efaproxyn/Revaproxyn) (Figure 18) from Allos Therapeutics is currently in Phase III trials in the United States as a chemoradiosensitizer in primary NSCLC. However, enrollment is currently suspended because Allos is focusing on development in other indications, such as treatment of brain metastases from breast cancer, for which the drug is preregistered in the United States and Europe. Efaproxiral is an allosteric modiÞer of hemoglobin that enhances the diffusion of oxygen to hypoxic tumor tissues, thereby making radiation therapy and some chemotherapeutic drugs more effective. Allos had initiated the Phase III Enhanced Lung cancer treatment with Induction chemotherapy and Thoracic radiation and Efaproxiral (ELITE); the trial aimed to recruit 600 patients to study the effectiveness of chemotherapy plus radiotherapy with or without efaproxiral. However, in March 2003, enrollment was suspended until Allos could secure funding for these trials. In March 2005, the company secured $50 million funding in private equity, but stated that it plans to use this capital to fund ongoing Phase III trials of efaproxiral in its primary indication, the treatment of brain metastases from breast cancer. Interim analysis of a Phase II study in which locally advanced, unresectable NSCLC patients were treated with induction chemotherapy followed by thoracic radiotherapy with concurrent efaproxiral was presented at the ASCO meeting in 2002 (Nabid A, 2003). One- and two-year survival rates were 67% and 40%,

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respectively, similar to rates observed in induction chemotherapy followed by concurrent chemoradiotherapy in nonresectable, stage III NSCLC patients (Vokes EE, 2002). Vaccines Overview. Preclinical research into speciÞc targets for vaccine design in cancer is abundant. However, there has been relatively poor success in this area in terms of clinical application. In NSCLC, there is little activity in this area, perhaps because of the lack of an appropriate target. Aphton’s IGN-101 is directed against the epithelial cell adhesion molecule (Ep-CAM) and is the most advanced vaccine in trials for NSCLC. Other vaccines in Phase II development for NSCLC but not discussed here due to limited data include Biomira/Merck KGaA’s BLP-25, a liposomal MUC-1 peptide-based vaccine; Antigenics’ HSPPC-96, a peptidic heat shock protein (HSP) vaccine; and a telomerase peptidic vaccine from GemVax. Cell Genesys has GVAX, a GM-CSF-producing autologous cell vaccine, in Phase II development, but the company is likely to pursue development for its primary indication of prostate cancer. Mechanism of Action. Cancer vaccines aim to enhance a host immune response against a tumor-speciÞc target, thereby triggering the cell-mediated killing of tumor cells. One of the major drawbacks of this approach is that cancer patients tend to be immunocompromised, owing to the cancer itself or to damage sustained by healthy cells during chemotherapy. Moreover, the immunogenicity of recombinant vaccines is often insufÞcient to illicit a protective response even in an intact immune system. IGN-101. IGN-101 (Edrecolomab) was formerly in development by Igeneon for treatment of epithelial cancers. In December 2004, Aphton, a biotechnology company based in the United States, announced that it would acquire Igeneon in an all-stock deal, and IGN-101 appears to be a key player in the merged company’s pipeline. Igeneon had previously progressed IGN-101 to Phase III trials for NSCLC in Europe. The IGN-101 vaccine consists of a murine MAb that is structurally similar to Ep-CAM, a membrane protein that is often overexpressed on epithelial cells. Vaccination with this recombinant molecule results in the production of host antibodies directed against Ep-Cam, thereby leading to immune destruction of cells that express Ep-Cam. Igeneon initiated Phase II/III trials of IGN-101 that aimed to enroll 420 patients with operable NSCLC to receive either IGN-101 or placebo immediately after successful surgery for a primary tumor. Preliminary results announced in a January 2002 press release revealed that the vaccine was well tolerated in the 28 patients treated thus far, induced a strong immune response in all patients, and reduced numbers of blood micrometastases. No further results have been announced.

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In November 2004, Igeneon presented data from a 239-patient, Moroccan Phase II trial of IGN-101 in metastatic rectal cancer patients. The trial included patients with colorectal cancer or upper gastrointestinal (GI) tract cancer as well as patients with NSCLC or liver or bile duct carcinoma. No difference in overall survival of the overall population was seen, but the median survival of rectal cancer patients treated with IGN-101 was 407 days, compared with 253 days for placebo (Igeneon, press release, November 2004). In 2001, a Phase I trial of IGN-101 was carried out in Austria in 18 patients with different epithelial cancers. Results showed that IGN-101 was well tolerated and induced production of antibodies in all patients, resulting in a signiÞcant reduction of circulating tumor cells (Loibner H, 2002; Samonigg H, 2002). A subsequent Phase II trial investigated the IGN-101 vaccination with various concomitant chemotherapy regimens (Samonigg H, 2003). Results showed the presence of speciÞc immunogenicity; the investigators concluded that the vaccine warranted further testing (Himmler G, 2003). Novel Alkylating Agents Overview. A nitrogen mustard alkylating agent was the Þrst nonhormonal chemical to demonstrate signiÞcant antitumor activity. Subsequently, less toxic and more clinically effective alkylating agents have been developed—for example, cyclophosphamide, ifosfamide, melphalan, mitomycin C, and nitrosourea derivatives. Despite the large number of agents available, an improvement in toxicity is still required. Mechanism of Action. Alkylating agents exert their cytotoxic action by alkylation of DNA bases. Alkylating agents may be mono- or bifunctional (the latter induces DNA cross-links). Unless repaired, the alkylated lesions will prevent the cell from replicating effectively. One mechanism of resistance to alkylating agents is DNA repair enzymes such as O6-alkylguanine-alkyltransferase, which removes the alkyl group from the O6 position in guanine. TLK-286. TLK-286 (Telcyta) is under development by Telik for treatment of a variety of cancers and is in Phase III trials for NSCLC in the United States. In December 2003, the FDA granted TLK-286 fast-track status for third-line therapy of locally advanced or metastatic NSCLC. TLK-286 is a pro-drug alkylating agent activated by glutathione S-transferase (GST) P1-1. Because this enzyme is commonly overexpressed in human cancers and correlated with chemotherapy resistance, Telik hopes that TLK-286 will be preferentially activated in malignant tissue. When cleaved by GST P1-1, TLK286 is converted to the active form, which initiates various cellular processes, such as induction of apoptosis. Preliminary results of a Phase I/II dose ranging trial of TLK-286 in combination with carboplatin and paclitaxel as Þrst-line therapy in patients with advanced NSCLC were disclosed at the 2005 ASCO meeting (Sequist SV, 2005). The doses of TLK-286 being used are 400, 500, 750, and 1,000 mg/m2 . Interim analysis of the Þrst nine treated patients revealed no dose-limiting toxicities. The most

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common toxicities observed were grade I/II alopecia, anemia, thrombocytopenia, and myalgia. Of the nine patients evaluable, Þve partial responses (63%) and two cases of stable disease were observed. Accrual at the 1,000 mg/m2 dose is continuing. In March 2004, Telik initiated the Phase III Assessment of Survival In Solid Tumors-2 (ASSIST-2) study, which was to enroll 500 patients with platinumrefractory or platinum-resistant NSCLC to receive either TLK-286 or geÞtinib for third-line treatment of NSCLC. The company has also initiated trials of TLK-286 in combination with carboplatin/paclitaxel and cisplatin for Þrst-line treatment of stage IIIB or IV NSCLC. Interim analysis from a Phase II study of weekly TLK-286 in NSCLC patients who had failed platinum-based chemotherapy revealed that partial response occurred in 1 of 12 patients, minor responses in 2 of 12, and stable disease in 4 of 12. The overall disease control rate was 58%. Therapy was well tolerated, with infrequent grade III and no grade IV adverse events (Papadimitrakopoulou V, 2003). Additional data from this trial were presented at the 10th World Conference on Lung Cancer in Vancouver in August 2003. Of 19 evaluable patients, 69% achieved disease stabilization, and the response rate was 11% (Telik, press release, August 2003). TLK-286 is also being investigated (up to a maximum dose of 960 mg/m2 ) in combination with docetaxel in patients with advanced-stage NSCLC who have failed platinum-based chemotherapy. In an interim analysis of 24 evaluable patients, grades III and IV neutropenia were observed in 15% of patients (febrile neutropenia in 7%), leukopenia in 10%, and fatigue in 15%. No cumulative toxicity has been observed. Six patients demonstrated partial response and 11 had stable disease (Papadimitrakopoulou V, 2004). The investigators concluded that a 960 mg/m2 dose of TLK-286 should be used in further studies. REFERENCES Abratt RP. Vinorelbine (NVB)-carboplatin (CBDCA) vs. non-platinum doublets in inoperable non-small-cell lung cancer (NSCLC) patients (pts)-Þnal results of the Glob 2 Phase III with patient beneÞt analysis. Proceedings of the American Society of Clinical Oncology. 2004. Abstract 7016. Adachi M, et al. Reduced integrin α3 expression as a factor of poor prognosis of patients with adenocarcinoma of the lung. Journal of Clinical Oncology. 1998;16:1060–1067. Adachi M, et al. SigniÞcance of integrin α5 gene as a prognostic factor in node-negative non-small cell lung cancer. Clinical Cancer Research. 2000;6:96–101. Adjei AA, et al. Pemetrexed (Pem)/gemcitabine (Gem) as front-line therapy for advanced NSCLC: A randomized, Phase II trial of three schedules. Proceedings of the American Society of Clinical Oncology. 2004. Abstract 7070. Ahrendt SA, et al. Alcohol consumption and cigarette smoking increase the frequency of p53 mutations in non-small cell lung cancer. Cancer Research. 2000;60(12): 3155–3159. Akerley W, et al. Induction chemotherapy with paclitaxel (9P) and carboplatin (C) followed by concurrent thoracic radiation and weekly PC for patients with unresectable

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Telik. Telik announces positive follow-up results from Phase II trial of Telcya in advanced non-small-cell lung cancer. Press release, August 2003. www.telik.com. Accessed May 2005. Tester WJ, et al. ECOG 1599: Randomized phase II study of paclitaxel/carboplatin or gemcitabine/cisplatin in performance status (PS) 2 patients with advanced non-smallcell lung cancer (NSCLC). clinical outcome of gemcitabine/cisplatin-treated advanced non-small-cell lung cancer patients. Proceedings of the American Society of Clinical Oncology. 2004. Abstract 7055. Tishler RB, et al. Taxol sensitizes human astrocytoma cells to radiation. Cancer Research. 1992;52:3495–3497. Tran HT, et al. Herceptin in combination with cisplatin and gemcitabine in patients (pts) with HER2 overexpressing, untreated, advanced, non-small-cell lung cancer (NSCLC): Þnal report of a Phase II trial. Proceedings of the American Society of Clinical Oncology. 2002. Abstract 1226. Travis WD, et al. United States lung carcinoma incidence trends. Cancer. 1996;77: 2464–2470. Treat J, et al. A randomized Phase III trial of gemcitabine (G) in combination with carboplatin (C) or paclitaxel (P) versus paclitaxel plus carboplatin in advanced (stage IIIB, IV) non-small cell lung cancer (NSCLC): update of the Alpha Oncology trial (A199002L). Proceedings of the American Society of Clinical Oncology. 2005. Abstract LBA7025. Tsai C, et al. Mutation in the tyrosine kinase domain of epidermal growth factor receptor is a predictive and prognostic factor for geÞtinib treatment in patients with non-small-cell lung cancer. Proceedings of the American Society of Clinical Oncology. 2005. Abstract 7086. Tsao MS, et al. Molecular analysis of the epidermal growth factor receptor (EGFR) gene and protein expression in patients treated with erlotinib in National Cancer Institute of Canada Clinical Trials Group (NCIC CTG) trial BR.21. Proceedings of the American Society of Clinical Oncology. 2005. Abstract 7007. Tseng JE, et al. Loss of FHIT is frequent in stage I non-small-cell lung cancer and in lungs of chronic smokers. Cancer Research. 1999;59(19):4798–4803. U.S. FDA ofÞcial hints at reprieve for Iressa. Scrip. PJB Publications Ltd; April 12, 2005;3040:14. Vansteenkiste JF, et al. Clinical beneÞt response in advanced non-small-cell lung cancer: a multi-center prospective randomized Phase III study of single agent gemcitabine versus cisplatin-vindesine. Annals of Oncology. 2001;12:1221–1230 Van Zandwijk N, Van’t Veer LJ. The role of prognostic factors and oncogenes in the detection and management of non-small-cell lung cancer. Oncology. 1998;12(1) (suppl 2):55–59. Villaßor VM, et al. Potential thromboembolic(TE) complications in patients with nonsmall-cell lung cancer (NSCLC) treated with paclitaxel(P), carboplatin(C), and exisulind(E). Proceedings of the American Society of Clinical Oncology. 2003. Abstract 2839. Vineis P, et al. Environmental tobacco smoke and risk of respiratory cancer and chronic obstructive pulmonary disease in former smokers and never smokers in the EPIC prospective study. British Medical Journal. 2005;330:277–287.

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Virmani AK, et al. Allelotyping demonstrates common and distinct patterns of chromosomal loss in human lung cancer types. Genes, Chromosomes, Cancers. 1998;21:308–319. Vokes EE, et al. Gemcitabine and radiation therapy for non-small-cell lung cancer. Seminars in Oncology. 1998;25:66–69. Vokes EE, et al. Randomized Phase II study of cisplatin with gemcitabine or paclitaxel or vinorelbine as induction chemotherapy followed by concomitant chemoradiotherapy for stage IIIB non-small-cell lung cancer: cancer and leukemia group B study 9431. Journal of Clinical Oncology. 2002; 20(20):4191–4198. Von Pawel J, et al. A tolerability study of tirapazamine, vinorelbine, and cisplatin combination therapy in subjects with advanced (stage III and IV) non-small-cell lung cancer. Proceedings of the American Society of Clinical Oncology. 2001. Abstract 2737. Wenzlaff AS, et al. GSTM, GSTT1 and GSTP1 polymorphisms, environmental tobacco smoke exposure and risk of lung cancer among never smokers: a population-based study. Carcinogenesis. 2005;26(2):395–401. West H, et al. Pemetrexed (P) plus gemcitabine (G) as front-line chemotherapy for patients (Pts) with locally advanced or metastatic non-small-cell lung cancer (NSCLC): a Phase II clinical trial. Proceedings of the American Society of Clinical Oncology. 2005. Abstract 7117. White SC, et al. Phase II study of oral topotecan in advanced non-small-cell lung cancer. Clinical Cancer Research. 2000; 6(3):868–873. Whitehead CM, et al. Cp461 in an orthotopic human NSCLC rat model involves phosphodiesterase targeting, apoptosis induction, g2/m block and anti-proliferation. Proceedings of the American Association of Cancer Research. 2002. Abstract 4577. William RN, et al. State of the art molecular and genetic aspects of lung cancer. American Journal of Respiratory and Critical Care Medicine. 2000;161:1355–1367. Williamson SK, et al. Paclitaxel/carboplatin (PC) v PC + tirapazamine (PCT) in advanced non-small-cell lung cancer (NSCLC). A Phase III Southwest Oncology Group (SWOG) Trial. Proceedings of the American Society of Clinical Oncology. 2003. Abstract 2502. Wingo PA, et al. Annual report to the nation on the status of cancer, 1973–1996, with a special section on lung cancer and tobacco smoking. Journal of the National Cancer Institute. 1999;91:675–690. Winton TL. A prospective randomized trial of adjuvant vinorelbine (VIN) and cisplatin (CIS) in completely resected stage 1B and II non-small-cell lung cancer (NSCLC) Intergroup JBR.10. Proceedings of the American Society of Clinical Oncology. 2004. Abstract 7018. Wistuba II, et al. Molecular genetics of small-cell lung carcinoma. Seminars in Oncology. 2001;28(2 suppl 4):3–13. Woods RL, et al. A randomized trial of cisplatin and vindesine versus supportive care only in advanced non-small-cell lung cancer. British Journal of Cancer. 1990;61:608–611. Xu HJ, et al. Altered retinoblastoma protein expression and prognosis in early-stage non-small-cell lung carcinoma. Journal of the National Cancer Institute. 1994;86(9): 695–699. Xynogalos S, et al. Expression of cyclins D3, B1, oncoprotein Rb and ki67 proliferation index in non-small-cell lung cancer. Proceedings of the American Society of Clinical Oncology. 2002. Abstract 1278. Yamaguchi NH, et al. COX-2 and MMP-9 as prognostic markers for lung adenocarcinoma. Proceedings of the American Society of Clinical Oncology. 2003. Abstract 2844.

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Ovarian Cancer

ETIOLOGY AND PATHOPHYSIOLOGY Introduction. The normal function of the human ovary (Figure 1) is the storage and release of unfertilized eggs and the secretion of sex hormones. Ovarian malignancies result from genetic mutations that occur during cell reproduction. These mutations alter or block mechanisms that regulate the cell-growth cycle, causing uncontrolled cell growth. This uncontrolled proliferation of cells can result in a cancerous tumor. Tumors of the ovarian epithelial cells make up 90% of ovarian cancers (CaOs). These tumors arise from the germinal epithelium of the ovary (the epithelium develops from the Mellerian duct and is closely related to the cells lining the peritoneal cavity). Tumors of the sex-cord stroma, tumors of germline cells, and tumors of undetermined histology make up approximately 6%, 3%, and 1%, respectively, of the remaining 10% of cases (Totolero-Luna G, 1995). Because most CaOs involve epithelial tumors, this section focuses on epithelial types of CaO. Epithelial CaO is the leading cause of gynecologic cancer deaths in the Western world and the Þfth-most-frequent cause of cancer death in women (after lung, breast, colorectal, and pancreatic cancer); 50% of all cases occur in women over age 65 (Yancik R, 1993). The etiology of CaO is poorly understood. However, epidemiological studies have identiÞed genetic, reproductive, and environmental factors that can affect a woman’s risk of developing CaO (Tiedemann D, 2000; Riman T, 2002). A reduced risk of CaO is associated with increased parity, oral Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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FIGURE 1. Female reproductive anatomy.

contraceptive use, and breast-feeding. Risk is increased in women with a family history of CaO and among women who have used fertility drugs. CaO is often referred to as the “silent killer.” Its high mortality rate is usually ascribed to late diagnosis because epithelial CaO usually lacks early warning symptoms. Furthermore, CaOs often lack deÞnite precursor lesions and are quite heterogeneous, and the molecular pathways underlying their progression are elusive, making diagnostic tests more difÞcult to establish. More than 70% of patients have stage III or IV disease at the time of diagnosis, a fact that translates into a poor Þve-year survival of 20%. By contrast, approximately 90% of women diagnosed with early-stage disease (conÞned to the ovary) survive for more than Þve years. Thus, early detection is essential for improved survival. Currently, no organization recommends routine CaO screening of average-risk women, and there is conßicting evidence for screening women with two or more Þrst-degree relatives with CaO. The lack of routine screening reßects the risk of false-positives and false-negatives from currently available screening methods, which include pelvic examination, CA-125 serum tumor marker, and transvaginal ultrasound (TVU). Therefore, great efforts are being made using genomic and proteomic technologies to identify new diagnostic markers of CaO that might be useful for screening and prognostic purposes. Etiology Genetic Risk Factors. The vast majority of CaOs are of unknown origin (sporadic); less than 10% of cases are hereditary (Richards WE, 1999). However, the ability to recognize the genetic component of CaO is becoming more reliable, with the result that it may be possible to identify populations at risk for developing CaO or populations that may be at greater risk for disease progression.

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Some mutations are associated with particularly aggressive or rapidly developing cancers; others affect the tumor’s susceptibility to different forms of treatment, such as chemotherapy or radiotherapy. Genetic mutations associated with CaO are detailed in the following sections. The identiÞcation of genes causing the cancer provides targets for therapy—in the case of tumor suppressor genes, turning them back on, and in the case of oncogenes, turning them off. Inherited Genomic Factors. Epidemiological studies have indicated that after controlling for age, the strongest risk factor for CaO is a family history of the disease. In the United States, compared with a lifetime risk of 1.6% for women in the general public, women who have a Þrst-degree relative (e.g., mother, sister, daughter) with CaO have a 5% risk, and women with two Þrst-degree relatives have a 7% risk (Werness B, 2001). To identify a familial cancer, a DNA sample from a cancer patient is analyzed to Þnd any speciÞc mutation in genes known to be involved in hereditary cancer. This laborious process can take as long as a year. If such a mutation is identiÞed, then other family members without cancer can be tested to see whether they possess this mutation and thus have an increased risk of developing a related cancer; this process is called predictive testing and is generally offered only when a relative has already been identiÞed as having a speciÞc mutation associated with hereditary cancers. Analyses of familial CaO pedigrees and other epidemiological studies have identiÞed three distinct ways in which CaO can be inherited: •

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Hereditary breast and ovarian cancer (HBOC) syndrome applies to women who have Þrst- and second-degree relatives with breast cancers and/or ovarian cancers. Approximately 10% of women with CaO are carriers of a breast and ovarian cancer susceptibility gene. The proportion of cases of CaO owing to such a gene declines with age: it is estimated to be 14% for women diagnosed in their fourth decade and 7% for women diagnosed in their sixth decade. BRCA1 and BRCA2 (breast cancer susceptibility genes 1 and 2, respectively) together account for 85% of HBOC syndrome. Hereditary nonpolyposis colorectal cancer (HNPCC), also known as Lynch syndrome II or family cancer syndrome, is a hereditary syndrome most commonly characterized by an increased risk for colorectal cancer. The lifetime risk of colorectal cancer is 80% and is typically diagnosed when the patient is in his or her mid-40s. The risk of endometrial (uterine) cancer associated with HNPCC is approximately 40%, and the risk of CaO is 10%. Other associated cancers include those of the stomach, small bowel, urinary tract, and biliary tract. Site-speciÞc CaO syndrome is a rare syndrome affecting women with two or more Þrst-degree or Þrst- and second-degree relatives who have had CaO. Multiple female family members are affected by the development of CaO only. Some researchers have suggested that site-speciÞc CaO syndrome

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represents a variant of HBOC in which early-onset breast cancer is not present (Boyd J, 1998). Mutations in the tumor suppressor genes BRCA1 or BRCA2 together account for an estimated 90% of hereditary CaO; two-thirds of those cases are linked to BRCA1 mutations and one-third are associated with BRCA2 (Frank TS, 1998). Women with BRCA1 and BRCA2 mutations are ten times more likely to develop CaO than are women without these mutations (Boyd J, 1998). Certain ethnic groups, such as Ashkenazi Jews, have an increased probability of harboring germline BRCA1 or BRCA2 mutations. The three most common BRCA mutations that occur in healthy Ashkenazi Jewish women are the 185delAG mutation in BRCA1 , the 5382insC mutation in BRCA1, and the 6174delT mutation in BRCA2, with a collective prevalence of approximately 2.5% (Moslehi R, 2000). Among Ashkenazi Jewish women with a known diagnosis of CaO, however, the frequency of such mutations has been reported to be as high as 26–41% (Ben David Y, 2002). Women with a germline mutation in BRCA1 are reported to have a lifetime risk of CaO ranging from 16% to 44% and a lifetime risk of breast cancer ranging from 56% to 87%. CaO may develop at an earlier age in women with germline BRCA1 mutations than in those with the sporadic form of the disease, although it is important to recognize that CaO may occur at any age in mutation carriers. Like BRCA1, the BRCA2 protein is localized in the nucleus and is involved in DNA repair through its association with the protein RAD51. Women with germline mutations in BRCA2 have a lifetime risk of breast cancer that is similar to that for carriers of the BRCA1 mutation, and their lifetime risk of CaO is approximately 10% (Struewing JP, 1997). Although little evidence supports the idea of mutational hot spots or clustering on BRCA1 and BRCA2 genes except in certain populations (e.g., Ashkenazi Jews), a deÞned repertoire of mutations has been detected. Research has suggested that the location of BRCA mutations is associated with different CaO risks. Mutations at the C-terminal of BRCA1 protein appear to be associated with breast cancer, whereas mutations at the N-terminal of the protein are more strongly associated with CaO (Gayther SA, 1995). The RING-Þnger domain of the N-terminus of BRCA1 protein appears to play a role in the antiapoptotic function in ovarian surface epithelium (OSE) cells (Johnson NC, 2002). Among BRCA2 mutation carriers, the risk of CaO is greatest for women with mutations clustered in a region of 3.3 kb in exon 11 (Gayther SA, 1997). The natural history of CaO that develops in the setting of BRCA1 or BRCA2 germline mutations appears to be characterized by a more indolent course than that of sporadic disease. The majority of BRCA1 -associated familial CaOs are serous adenocarcinomas in which the tumor appears as solid masses and cysts. The mean age at diagnosis of CaO in women with BRCA1 mutations (54 years) is younger than that of women with sporadic CaO (63 years). Furthermore, BRCA1 -associated familial CaO has a longer disease-free interval following primary chemotherapy in comparison with sporadic CaO, with a median time to recurrence of 14 months and 7 months, respectively (Boyd J, 2000).

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Lynch syndrome II probably accounts for the remaining 10% of hereditary CaOs (Boyd J, 1998). Lynch syndrome II cases are associated with germline mutations in the DNA mismatch repair genes hMLH1 and hMSH2 . The mutations cause widespread genomic instability, or a hypermutable state, which provides the background for an accelerated accumulation of mutations. Acquired Genomic and Proteomic Factors. Most DNA mutations related to CaO are acquired, rather than inherited. These mutations of oncogenes and/or tumor suppressor genes may result from radiation or cancer-causing chemicals. So far, studies have not been able to speciÞcally link any single chemical in the environment or in our diets to mutations that cause CaO. Thus, the cause of most acquired mutations remains unknown. Most sporadic CaOs have several acquired gene mutations. Research suggests that tests identifying acquired changes of certain genes, such as the p53 tumor suppressor gene or the HER2 oncogene, in CaOs may help predict a woman’s prognosis. These genetic changes can also be used to identify patients suitable for particular targeted therapies. Gene expression arrays are being used to identify novel serum tumor markers, and researchers have suggested that a number of markers in combination might identify more than 99% of CaO cases despite the heterogeneity of the disease (Lu KH, 2004). The true effectors of cancer are the proteins produced by the genetic changes. These associated proteins may provide useful biomarkers to detect CaO in the early stages and serve as prognostic markers that would enable physicians to better manage their patients’ disease. Emerging proteomic technologies that can detect CaO-associated proteins are paving the way for serum proteomic diagnostics and the use of proteomics to monitor molecular-targeted treatments (Posadas EM, 2004). Molecular-targeted therapy involves the characterization of promising targets within tumor cells and the development of substances that can speciÞcally attack these targets. The following sections describe the genes and their associated proteins that have frequently been observed in sporadic CaO. •

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BRCA1 and BRCA2. Although mutations of BRCA genes are rare in sporadic ovarian tumors, loss of function of these proteins may play a role in tumor development (Foster KA, 1996). BRCA1 and BRCA2 are tumor suppressor genes that, when functioning normally, help prevent cells from embarking on uncontrolled proliferation. In sporadic tumors, researchers have noted high rates of loss of heterozygosity at the BRCA1 and BRCA2 loci and/or reduced expression (Buller RE, 2001). Hence, manipulation of the BRCA1 and BRCA2 genes may hold therapeutic potential for CaO. HER2/neu. Human epidermal growth factor receptor 2 (HER2/neu, also known as ErbB-2 ) is one of the most widely studied oncogenes. It plays a major role in breast cancer, in which it is overexpressed in up to 30% of cases; by contrast, it is overexpressed in only 11% of CaO cases (Bookman MA, 2003). HER2 overexpression is associated with a greater likelihood of

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residual and recurrent disease after treatment, reduced sensitivity to certain types of drug treatment, poor prognosis, and shorter survival times (Tong X, 1999). Overexpression of this gene is thought to increase cell proliferation, possibly by making cells more responsive to normal levels of epidermal and/or epithelial growth factors (Boyd J, 1998). Hence, down-regulation of HER2 overexpression in tumor cells may be an attractive therapeutic approach. p53. The p53 tumor suppressor gene normally regulates apoptosis (programmed cell death). Mutations of p53 occur in 30–80% of epithelial-type CaO cases and cause the gene to lose its ability to direct cell death (Perego P, 2001). p53 alterations are also associated with chemo-resistance and aggressive tumor growth (Tong X, 1999). In laboratory studies of cultured epithelial tumor cells, restoration of the p53 gene function often halted or inhibited the growth of aberrant cells (Nielsen L, 1998). These experiments, although preliminary, suggest that restoring the expression of tumor suppressor genes may provide the basis for treating human cancers. EGF. Epidermal growth factor (EGF), a complex protein that triggers the growth of epithelial cells, is overexpressed in 20–30% of CaOs. For CaO, overexpression of the EGF receptors ErbB and ErbB-4 is associated with a poor prognosis. Many primary (as opposed to metastatic) CaO tumors express ErbB-4, and research suggests that levels of this receptor may be increased in advanced disease (Tong X, 1999). PDGFR. Platelet-derived growth factor (PDGF) expression is found in 75% of primary ovarian tumors but its expression is undetectable in benign tumors or normal ovaries (Versnel MA, 1994). In addition, approximately 73% of CaOs express the PDGF receptor (PDGFR) (a type III receptor tyrosine kinase) and at least 50% of these cases coexpress PDGFR-alpha (Henriksen R, 1993). Furthermore, CaO patients expressing PDGFR demonstrate an overall shorter survival time compared with those whose tumors do not express the receptor. Cyclin D1. Overexpression of the substance cyclin D1 is correlated with CaO. Cyclin D1 is a regulator of early-stage cell growth and may play a role in the pathogenesis of several cancers. Researchers believe that cyclin D1 probably plays a role in the early development of ovarian malignancy (Worsley SD, 1997; Buller RE, 2001). Telomerase. The ribonucleoprotein telomerase—produced by germ cells and cancerous cells—interferes with apoptosis and therefore contributes to cell proliferation (Meyerson M, 2000). This enzyme acts on the telomeres, which cap the ends of chromosomes, keeping them from unraveling or being cut off during gene replication. Under normal circumstances, telomeres become ever shorter with each replication. Eventually, they become so short that apoptosis is triggered, killing the cell containing the shortened chromosome. Cancerous cells, however, can produce telomerase, an ability

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that helps the telomeres maintain their length and thus contributes to cellular immortality. Overexpression of telomerase is common in high-grade (poorly differentiated), aggressive tumors, including ovarian tumors. Oncology specialists believe that this enzyme is a promising new target for cancer screening (Blake P, 2000). K-ras. Mutations in the K-ras gene are frequently found in malignant neoplasms. K-ras-activating mutations lead to constitutive activation of the protein by increasing guanine diphosphate/guanine triphosphate (GDP/GTP) exchange or reducing GTPase activity of the protein, thus leading to increased cell proliferation. K-ras mutation frequencies seem to be highly related to tumor histology; in CaO, K-ras mutations occur signiÞcantly more frequently in mucinous tumors than in serous carcinomas and all nonmucinous types of epithelial ovarian tumors combined (Enomoto T, 1991). C-myc. C-myc ampliÞcation is a common Þnding in advanced-stage CaO. A recent study found that expression of the c-myc gene was 1.78-fold higher in platinum-resistant versus platinum-sensitive CaO cells (Elahi A, 2005). Thus, c-myc expression may represent a novel targeted molecular therapeutic approach to patients with platinum-resistant CaO. OPCML. Researchers found that in 90% of CaO cases, the opioid-binding protein/cell adhesion molecule-like (OPCML, also called OBCAM ) gene was switched off (Sellar GC, 2003). Furthermore, experiments show that when a functioning OPCML is inserted into CaO tissue, the gene greatly suppresses cancer growth, indicating that it may be a key tumor suppressor gene in CaO. OPCML is a member of the IgLON family of immunoglobulin (Ig) domain-containing glycosylphosphatidylinositol (GPI)-anchored cell adhesion molecules. EEF1A2. The gene encoding protein elongation factor EEF1A2 (also known as eEF-1 alpha 2 ) is ampliÞed in 25% of primary ovarian tumors and is highly expressed in approximately 30% of ovarian tumors and established cell lines (Anand N, 2002). EEF1A2 enhances focus formation, allows anchorage-independent growth, and reduces the doubling time of rodent Þbroblasts. It may also be associated with increased risk for recurrence of disease (Sharma S, 2005). These data suggest a potential new prognostic marker and oncogene for CaO and may lead to the study of EEF1A2 as a suitable target for CaO therapy. AKT. AKT and mTOR (mammalian target of rapamycin) phosphorylation are frequently detected in CaO, and activation of the P13K/AKT pathway may contribute to tumorigenesis. AKT mediates survival signals that protect cells from apoptosis. Experiments have shown that pretreatment of SKOV3 cells, which exhibit constitutive AKT activity under low serum conditions, with a P13K inhibitor augmented cisplatin-induced apoptosis (Altomare DA, 2004). Furthermore, inhibition of mTOR activity with rapamycin resulted in G1 arrest in SKOV3 cells. These Þndings indicate that active AKT

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and downstream mTOR represent potentially important therapeutic and/or chemopreventive targets in CaO. In addition, experimental data have shown that the molecular chaperone heat shock protein-90 (HSP-90) sensitizes taxol-resistant CaO cells to cancer through its impact on the P13K/AKT pathway (Sain N, 2004). HtrA1. Expression of HtrA1, a candidate tumor suppressor gene, is downregulated in Þve of seven CaO cell lines. In total, 59% of primary ovarian tumors have either a complete absence or markedly reduced levels of HtrA1 expression compared with the brushings of ovarian surface epithelium (Chien J, 2004). HE4. Human epididymis protein-4 (HE-4) is a secreted glycoprotein that is overexpressed in 93% of serous and 100% of endometrioid CaOs (Drapkin R, 2005). A recent study produced monoclonal antibodies to HE4 epitopes and quantiÞed the HE-4 levels with an enzyme-linked immunosorbent assay (ELISA) to determine the efÞcacy of the assay using the sera of postmenopausal women with CaO (Hellstrom I, 2003). The speciÞcity of HE-4 alone was signiÞcantly greater than CA-125 in its ability to distinguish between malignant and benign disease, providing incentive for future investigations of HE-4 as a diagnostic CaO biomarker to be used in combination with CA-125. ATF3, NM23-H2, caveolin-1, and DLC-1. Researchers recently identiÞed four progesterone-regulated, antitumor genes for CaO that may serve as therapeutic targets (Syed V, 2005). Semi-quantitative reverse transcriptionpolymerase chain reaction (RT-PCR ) analyses conÞrmed loss or reduced transcription of ATF3, NM23-H2, caveolin-1, and DLC-1 in OVCA cells when compared with HOSE cells and their up-regulation following progesterone treatment. ATF-3 functions primarily as an apoptosis inducer and NM23-H2 as a suppressor of cell motility, and caveolin-1 and DLC-1 exhibit features of classical tumor suppressors. Mucins. Human epithelial tumors express various mucin glycoproteins on their cell surfaces. At least 16 different human mucin genes, designated the MUC family, have been characterized. CA-125 was recently designated as MUC-16 after its cDNA sequence was identiÞed. CA-125 is overexpressed in more than 80% of nonmucinous CaOs and is recognized as a tumor marker associated with CaO. Other mucins strongly associated with CaO are MUC1 and MUC4. MUC1 modulates the activity of ErbB receptors and MUC4 acts as a receptor ligand and binds to ErbB-2 (Carraway KL, 1999). IGF-2. Expression of the insulin-like growth factor-2 (IGF-2 ) gene is signiÞcantly higher in CaO relative to normal ovarian surface epithelium. Furthermore, high IGF-2 gene expression is associated with high-grade, advanced-stage disease and is an independent predictor of poor survival in patients with CaO. As such, IGF-2 is a molecular marker and potential therapeutic target for the most aggressive CaOs. The Yale School of Medicine

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is developing a test relying on the detection of the proteins IGF-2, leptin, prolactin, and osteopontin to detect CaO at the early stage (Mor G, 2005). Looking for raised levels of these proteins led to CaO being identiÞed with 95% accuracy in a test group of more than 200 women. However, the researchers acknowledge that their test needs further work because even a 5% failure rate could potentially lead to the misdiagnosis of large numbers of women. Human Kallikreins. Human Kallikreins (hKs) are secreted serine proteases. Several members of the hK family are overexpressed in CaO, indicating that they may have potential as biomarkers and play a future role in the diagnosis, management, and prognosis of CaO. Several studies have investigated the prognostic values of these proteins. One study suggested that hK4 expression might be a predictive marker for paclitaxel resistance (Xi Z, 2004). Others reported that expression of hK8, hK11, and hK13 are increased during the development of CaO and down-regulated during CaO progression, indicating they are favorable prognostic markers (Diamandis EP, 2004; Scorilas A, 2004; Shigemasa K, 2004). On the other hand, hK5, hK6, hK7, hK10, and hK14 are considered unfavorable prognostic markers and are associated with more aggressive forms of CaO or late-stage disease (Diamandis EP, 2003; Hoffman BR, 2002; Kyriakopoulou LG, 2003; Luo LY, 2003; Yousef GM, 2002). YKL-40. YLK-40 is a glycoprotein in the chitinase protein family. A recent study showed that 20 of 31 early-stage CaO patients had elevated serum YLK-40 levels compared with 11 of 31 for CA-125 (Dupont J, 2004). Furthermore, YLK-40 levels increased with stage, regardless of grade, histology, or patient age, and high expression of YLK-40 in early-stage tumors was associated with poor prognosis. Therefore, YLK-40 may represent a novel marker for the detection of early-stage CaO and may predict disease recurrence and survival.

Reproductive and Menstrual Risk Factors. Numerous studies have focused on the contribution of a woman’s reproductive and menstrual history to the risk of developing CaO (Alberts DS, 1995; Greene MH, 1999; Riman T, 2002; Wittenberg J, 1999). Incessant ovulation is an important determinant of CaO risk; factors that suppress ovulation, such as pregnancy, the use of oral contraceptive pills, and lactation, reduce the risk of CaO (Purdie DM, 2003). Epidemiological evidence suggests that late menopause may be associated with an increased risk of CaO (Franceschi S, 1991). Other studies have found no relationship between age at menarche and the risk of CaO (Purdie D, 1995). Nulliparity (not having given birth to any children) is associated with an increased risk of CaO (Bristow RE, 1996). A study showed that multiparous (having had two or more offspring) women have a 30–70% lower risk of developing CaO compared with nulliparous women (Mosgaard BJ, 1997). Infertile women who received fertility drugs for extended periods of time are also at a higher risk of developing CaO (Rossing MA, 1994).

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One etiologic hypothesis suggests that the ovarian epithelium becomes damaged by each ovulation and that this damage must be repaired through the proliferation of replacement cells (Ozols RF, 2001). The greater the number of ovulations, the greater the number of repairs needed and the higher the chance of mutations—and thus malignancies. Authors of a large Swedish epidemiological study propose a different hypothesis—retrograde transportation of contaminants (Riman T, 2002). They argue that retrograde bleeding through the fallopian tubes may transport carcinogenic contaminants to the ovaries. Their hypothesis is based on the epidemiological Þndings that the cessation of retrograde bleeding after tubal ligation and hysterectomy is associated with a reduced risk of CaO. Hormone-Dependency Risk Factors. Some epidemiological studies indicated that CaO is endocrine-related and hormone-dependent. Exposure to estrogens during different periods in a woman’s life has been associated with the risk of CaO. The most common sources of exposure to hormones for women are the use of hormone replacement therapies (HRT) and oral contraceptives. Hormone Replacement Therapy. HRT usually involves treatment with either estrogen alone or in combination with progestin to compensate for reduced hormonal levels at menopause. Although the association between HRT and CaO remains unclear, several recent case-control studies have found an increased risk of epithelial CaO in relation to the use of HRT (Riman T, 2002; Lacey JV, 2002). Riman and colleagues found that women who took estrogen replacement therapy (ERT) were about 40% more likely than women who never took any form of HRT to develop CaO. Women who took HRT with sequential progestin were 50% more likely to develop CaO than were women who never underwent HRT. However, women who took low-potency estrogens or HRT with continuous progestin were at no higher risk of CaO than women who had never taken either. The women at highest risk were those who had taken HRT for more than ten years. Oral Contraceptive Pills. Any use of oral contraceptive pills (OCPs) has been shown to reduce CaO risk by 40–50% compared with never having taken them (Purdie D, 1995). The reduction in risk appears greatest following more than ten years of use. In addition, women with a family history of CaO appear to be at reduced risk of CaO with increasing duration of OCP use. One study demonstrated a 60% risk reduction in hereditary cancer after six years of OCP use in women with pathogenic mutations in the BRCA1 or BRCA2 gene (Narod S, 1998). One study suggested that four to eight years of OCP use can halve the risk of cancer in women with a family history of CaO by age 70 years, as compared with nonusers. Age. The risk for developing CaO increases with age. Most cases occur after menopause, which usually takes place around the age of 51. The median age at diagnosis is 63; the course of the disease is more rapid in women aged 65 or older.

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Lifestyle Risk Factors Obesity and Physical Activity A study from the American Cancer Society (ACS) found a higher rate of death from CaO in obese women (Rodriguez C, 2002). Obesity increased the risk of dying of CaO by 25% over women of normal weight. The ACS study also found that women who were 5 feet 9 inches or taller were 41% more likely to die of CaO than shorter women. Two explanations have been offered for this increase. The Þrst is that obesity causes the CaO deaths because obese women make higher amounts of the female hormone estrogen. Fat cells are an important source of estrogens in postmenopausal women. The second explanation involves another hormone, insulin-like growth factor (IGF) (as mentioned in the section “Acquired Genomic and Proteomic Factors,” expression of the IGF-2 gene is signiÞcantly higher in CaO relative to normal ovarian surface epithelium). This hormone has been involved in other hormone-dependent cancers such as breast and prostate cancer. It is elevated in obese people. Because these growth factors are also responsible for height, this explanation accounts for both risk factors-—height and obesity. Evidence suggests that physical activity may have a protective effect against CaO. In one study, the risk of CaO declined with increased frequency and duration of activity among premenopausal women (Zhang M, 2004). Diet. As with some other cancers, such as breast cancer, CaO is more common in developed industrialized countries, except for Japan. This phenomenon may reßect the consumption of diets low in vegetables or high in fat, particularly animal fat. No clear evidence links diet with CaO risk. One study compared women with CaO with similar women who had nonmalignant gynecological problems (Bosetti C, 2001). The cancer patients were more likely to eat lots of red meat, bread, soups, and sugar. Those who ate the most Þsh cut their risk in one-half. Lots of vegetables and pulses (peas, beans, and lentils) were also protective. Alcohol. In a study of 696 women with conÞrmed epithelial CaO and 786 cancerfree control women, consumption of any alcohol was associated with a reduced risk of CaO (Webb PM, 2004). Women who reported an average consumption of 25 g/day of alcohol (two standard drinks) had about one-half the risk of CaO compared with nondrinkers. Even an average consumption of less than one standard drink/week was associated with a 20% reduction in the risk of CaO. The study also reviewed all published data that comprehensively examined the association between alcohol consumption and the risk of CaO. When the researchers examined the intake of different types of alcohol, they determined that wine drinkers had a lower risk of CaO than both self-reported nondrinkers and women who reported drinking only beer or spirits. Red wine consumption in particular conferred the most protection: women who consumed more than one glass of red wine per day were almost seven times less likely to develop CaO than were women who never drank alcohol. It is therefore suggested that the apparent

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reduction in risk associated only with wine intake and not other types of alcohol may be because of something other than the alcohol in the wine. It is possible that the high levels of antioxidants and phytoestrogens found in wine (from grapes and grapeskins) reduce the risk of CaO. Environmental Risk Factors. Studies have indicated an increased risk of CaO associated with the use of talcum powder in genital hygiene (Cramer DW, 1999; Chang S, 1997). Talc (an ingredient in many body powders and feminine hygiene products) has been implicated in CaO because in the past, it was sometimes contaminated with asbestos, a known cancer-causing substance. Although such products are now required to be asbestos-free, proof of their safety or carcinogenicity (cancer-causing potential) has yet to be determined. Pathophysiology Characterization of Ovarian Epithelial Tumors. Approximately 50% of epithelial tumors are serous adenocarcinomas, which are both solid and cystic in appearance. Mucinous adenocarcinomas account for 36% of epithelial tumors and are made up of cysts containing mucinous ßuid. The remaining tumors are endometrioid adenocarcinomas—cysts containing brown ßuid—and clear-cell adenocarcinomas—cysts containing clear ßuid (Omura GA, 1991). In some of the major markets, tumor characteristics determine treatment. In addition to being distinguishable by tumor type, CaO cases can be classiÞed according to tumor histological grade, which refers to the degree of maturity of the cells forming the malignant ovarian tumor. The three grades of CaO are grade 1, “well differentiated”; grade 2, “moderately differentiated”; and grade 3, “poorly differentiated.” The lower the grade, the slower the tumor growth and the better the prognosis. Grading systems are based on the degree to which a tumor forms papillary structures or glands rather than solid tumors. Spread of Ovarian Cancer. The most common form of epithelial tumor dissemination throughout the peritoneal cavity is by exfoliation of malignant cells through the surface of the ovarian capsule (Chang TC, 1997). In the peritoneal cavity, the malignant cells follow the normal circulation of peritoneal ßuid to the undersurface of the hemidiaphragm, where they may implant and grow as surface nodules. Such exfoliation and implantation make up one of two primary modes of CaO spread within the peritoneal cavity. The other primary mode is via the retroperitoneal lymphatic nodes that drain the ovary, resulting in the involvement of lymph nodes lying along the aorta and vena cava up to the level of the renal vessels, inguinal lymphatics, and pelvic wall lymphatic nodes. Lymph node metastasis increases from 8% of stage I lesions to 29% of stage III cases and 53% of stage IV cases (Zang RY, 2000). The initial spread of CaO by both intraperitoneal and lymphatic routes is clinically occult. A signiÞcant proportion of women who appear to have

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tumors conÞned to one or both ovaries actually have widespread disease. These micrometastases can be detected only by histological examination of visually normal cells that are sampled during surgery (Ozols RF, 2001). For these reasons, inadequate assessment of the extent of disease spread has been a major problem in the management of patients with CaO. In the advanced stages of the disease, hematogenous spread is common and metastases can implant in the liver, the bones, and, less frequently, the brain. Signs and Symptoms. Epithelial cancers of the ovary have been described as silent killers because the overwhelming majority of patients present with disease that has spread outside the ovary and indeed outside the pelvis. Common symptoms include abdominal discomfort, abdominal distention (bloating), gastrointestinal (GI) symptoms, urinary frequency, and vaginal bleeding. Because of the nonspeciÞc nature of most of these symptoms, the diagnosis of CaO is seldom considered at Þrst presentation. Staging. When CaO is suspected, patients usually undergo an exploratory laparotomy (a surgical incision through the abdomen to the ovaries) to determine the stage of the disease (i.e., how far it has spread). A number of imaging techniques may also be used to help stage the cancer, including X-rays, ultrasound, and computed tomography (CT) scans. The International Federation of Gynecology and Obstetrics (FIGO) staging system categorizes CaO cases into stages I–IV—stage I referring to the least severe cases and stage IV to the most severe. Table 1 details the system. Prognostic Factors. Several clinical characteristics of CaO are considered valuable in assessing a patient’s prognosis. They can be useful in determining whether the patient will likely be cured with surgery and Þrst-line chemotherapy or will likely have a recurrence. Gene and protein expression proÞling of CaO samples using tissue microarrays is being used to investigate new prognostic tools. Tumor Stage. The treatment and prognosis of CaO are tied to the stage of disease: the higher the stage, the more aggressive the treatment and the poorer the prognosis. Stage I patients have a greater than 90% Þve-year survival rate; patients with stage II disease have a Þve-year survival rate of approximately 80%. Patients with stage III disease have a Þve-year survival rate of approximately 15–20% that depends in large part on the volume of disease present in the upper abdomen. Patients with stage IV disease have less than a 5% Þve-year survival rate. Peritoneal Fluid Cytology. Cytological samples of peritoneal ßuid are routinely obtained during staging procedures for CaO. These Þndings are important in substaging early (FIGO I and II) CaO; malignant cells in peritoneal washings or ascites warrant assignment of tumors to stage Ic or IIc.

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TABLE 1. International Federation of Gynecology and Obstetrics (FIGO) Staging System for Primary Carcinoma of the Ovary Stage Stage I Stage Ia

Stage Ib Stage Ic

Stage II Stage IIa Stage IIb Stage IIc

Stage III

Stage IIIa

Stage IIIb

Stage IIIc Stage IV

Description Growth limited to the ovaries. Growth limited to one ovary. Ascites (buildup of fluid inside the abdominal cavity) not present, nor tumors on the external surface of the ovary. Ovarian capsule is intact. Growth limited to both ovaries. Ascites not present, nor tumors on the external surface of the ovary. Ovarian capsule is intact. Tumor stage Ia or Ib but with tumor on the surface of one or both ovaries, with capsule rupture, with malignant ascites present, or with positive peritoneal washings. Growth involving one or both ovaries with pelvic extension. Extensions or metastases to the uterus or fallopian tubes associated with ovary and primary tumor. Growth involving one or both ovaries with extension into the pelvic cavity. Tumor either side stage IIa or IIb but with tumor on the surface of one or both ovaries, with capsule rupture, with malignant ascites present, or with positive peritoneal washings. Tumor involving one or both ovaries, with peritoneal metastases outside the pelvis, or positive retroperitoneal or inguinal nodes, or superficial liver metastases. Tumor is limited to the pelvis, but a histologically verified malignant extension to the small bowel or omentum is present. Tumor is grossly limited to the pelvis with negative nodes, but histologically confirmed macroscopic seeding of abdominal peritoneal surfaces is present. Tumor of one or both ovaries with histologically confirmed implants of abdominal peritoneal surfaces, none exceeding 2 cm in diameter; nodes negative. Abdominal implants greater than 2 cm in diameter or positive retroperitoneal or inguinal nodes. Growth involving one or both ovaries with distant metastasis; if pleural effusion is present, positive cytological test results are required to assign a case to stage IV. Parenchymal liver metastasis equals stage IV.

Note: The FIGO system is now used internationally in place of the tumor-node-metastasis (TNM) system.

Tumor Histology. The histological grade of a tumor correlates with relapse rates. For instance, a woman with grade 1 disease has a 22% chance of relapse after treatment; a woman with grade 3 disease has a 56% chance of relapse. The grading of CaO is particularly important in stage I patients because chemotherapy is withheld for low-grade tumors in view of their good prognosis when untreated. Performance Status. Physicians use the Karnofsky performance scale to grade the patient’s level of mobility and ability to manage daily life functions. The scale ranges from 10% (patient requires hospitalization for supportive care, moribund) to 100% (patient shows no evidence of the disease). The higher the rating, the better the prognosis. In addition, performance status often determines the choice of therapy: physicians are unlikely to administer aggressive therapy to

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TABLE 2. Karnofsky Performance Status Scale Description Able to carry on normal activity and to work; no special care needed.

% Rating 100 90 80

Unable to work; able to live at home and care for most personal needs; varying amount of assistance needed.

Normal; no complaints; no evidence of disease Able to carry on normal activity; minor signs or symptoms of disease Normal activity with effort; some signs or symptoms of disease

70

Cares for self; unable to carry on normal activity or do active work

60

Requires occasional assistance, but is able to care for most personal needs Requires considerable assistance and frequent medical care

50 Unable to care for self; requires equivalent of institutional or hospital care; disease may be progressing rapidly.

Definition

40

Disabled; requires special care and assistance

30

Severely disabled; hospital admission is indicated although death not imminent Very sick; hospital admission necessary; active supportive treatment necessary Moribund; fatal processes progressing rapidly Dead

20 10 0

patients whose performance status is poor. Table 2 details the Karnofsky performance scale. Age. Age is another important prognostic factor. In particular, cancer patients aged 60 or older have poorer survival rates than do younger patients. Median survival is at least two years longer in women younger than 65 than in women aged 65 or older. Older women are also at risk for developing more virulent subtypes of epithelial CaO. As they do for patients with poor performance status, physicians are likely to treat older women less aggressively. Volume of Residual Disease. For patients with advanced disease, the volume of residual disease following cytoreductive (tumor-debulking) surgery has a signiÞcant impact on survival. After administration of postoperative platinum-based chemotherapy, the Þve-year survival rate for patients with optimal stage III disease (deÞned as no residual nodule greater than 1 cm) is approximately 35%. This survival rate is signiÞcantly greater than the 15% survival rate of patients with suboptimal residual disease. CA-125 Levels. Approximately 80% of ovarian tumors express the circulatory protein cancer antigen-125 (CA-125), the most widely studied and used

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serum tumor marker for epithelial CaO. Physicians measure CA-125 to assess a patient’s prognosis and to gauge therapy success and disease progression. However, although elevated CA-125 can indicate the presence of CaO, CA-125 is not a suitable tool for screening purposes (Krishnan STM, 2002; Topalak O, 2002). The test has very poor speciÞcity in this setting; any cause of ascites, malignant or benign, leads to raised CA-125. The sensitivity of the test as a screening test is also poor; CA-125 is elevated in only 50% of stage I patients, although by the time disease has progressed to stage III and IV disease, it is elevated in more than 90% of patients. However, CA-125 levels measured after three cycles of chemotherapy are a relatively accurate prognostic tool. If CA-125 levels drop after therapy, remission is likely. A recent study concluded that CA-125 cannot be used for comparison of CaO tumor mass among patients, but serial estimation of CA-125 in individuals is fairly reliable as regards to the course of the tumor (Kumar P, 2005). The study also found that a CT scan is more informative as an evaluator and response indicator but is too expensive. DNA Ploidy. Ploidy represents the number of copies of DNA carried by a cell. Normal somatic cells are diploid (with two DNA copies); cancer cells can be aneuploid (with more than two DNA copies). DNA ploidy is assessed using ßow cytometry, which adds valuable data about tumor cell characteristics. The higher the DNA index (a direct measure of the number of copies), the poorer the prognosis (Trope C, 2000). However, not all researchers agree that aneuploidy indicates more serious disease (Resnik E, 1997). At this time, the Gynecologic Oncology Group, an international organization concerned with research as well as treatment guidelines, does not include aneuploidy as a criterion for predicting outcomes in women with early-stage disease. CURRENT THERAPIES Surgery plays a central role in the treatment of ovarian cancer (CaO); surgery with or without chemotherapy is performed in most patients regardless of stage of disease. Radiotherapy is very rarely used and only as a palliative treatment for patients with inoperable cancer to relieve symptoms such as bleeding, pain, and genitourinary obstruction. Most patients with CaO receive chemotherapy, the exception being those with stage Ia disease and those who are deemed too unÞt. Chemotherapeutic agents from a wide variety of classes have been shown to elicit responses in patients with CaO; consequently, many different regimens are in use, depending on physician opinions and preferred toxicity proÞles. Also, many other areas remain controversial in CaO treatment such as chemotherapy dose intensity, period of treatment, number of cycles, triplet versus doublet drug regimens, the use of maintenance therapy, and the method of administration. In any case, the Þrst-line chemotherapy regimen for CaO usually consists of a platinum-containing agent (platin)—cisplatin (Bristol-Myers Squibb’s ([BMS’s] Platinol-AQ, generics) or carboplatin (BMS’s Paraplatin, generics)—combined with another chemotherapy.

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This section focuses on regimens that are in regular clinical use or that may have a potentially greater role in the future. This coverage of current therapies is not intended to be inclusive of all drugs used to treat CaO because, as with many types of cancer, many drugs are used off-label or are used too rarely to warrant detailed discussion. Table 3 lists the leading therapies and combinations of therapies used to treat CaO. Agents that are not covered in detail but that are used, albeit rarely, include altretamine (MGI Pharma/Ipsen’s Hexalen, ProStrakan’s Hexastat), treosulfan (Medac’s Ovastat, generics), vinorelbine (GlaxoSmithKline/ Pierre Fabre’s Navelbine), irinotecan (SanoÞ-Aventis’s Campto, PÞzer’s Camptosar), tamoxifen (AstraZeneca’s Nolvadex, PÞzer’s Kessar, generics), doxorubicin (PÞzer’s Adriamycin, generics), and etoposide (generics). These agents are used as single agents or in combination, mostly in the second-line setting. With the exception of gemcitabine’s approval in Germany, altretamine’s approval in the United States and the United Kingdom, irinotecan’s approval in Japan, and treosulfan’s approval in Germany and the United Kingdom, these agents are used off-label; their use is likely to remain low because they do not offer signiÞcant advantages over standard treatments. Paclitaxel/Carboplatin Overview. In most of the major markets, the combination of paclitaxel (BMS’s Taxol, generics) (Figure 2) and carboplatin (Figure 3) is considered the standard of care. Compared with cisplatin, carboplatin is better tolerated and easier to administer (typically as a one-hour infusion rather than a six- to eight-hour infusion) and offers similar antitumor activity (Aabo K, 1998). Carboplatin does not require hydration and can be administered on an outpatient basis. However, its major and dose-limiting adverse effect—myelosuppression—is more pronounced than with cisplatin. Mechanism of Action. The individual components of the paclitaxel/ carboplatin regimen contribute the following mechanisms to achieve its overall activity: •

•

Paclitaxel is a taxane. Taxanes inhibit the dynamic reorganization of microtubule networks that is essential to vital interphase and mitotic cellular functions. Taxanes exhibit a unique mechanism of action. Paclitaxel, which achieves selective toxicity against rapidly proliferating cells, is mainly active in the G2/M phase of the cell cycle. Carboplatin is a platin. Platins inhibit tumor-cell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that carboplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle.

Clinical Performance. Phase III data comparing paclitaxel/carboplatin (TC) with paclitaxel/cisplatin (TP) have shown that both regimens have similar efÞcacies, but TC has a better toxicity proÞle.

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TABLE 3. Current Regimens/Classes Used for Ovarian Cancer Regimen or Class Paclitaxel/ carboplatin

Paclitaxel/ cisplatin

Cyclophosphamide/ doxorubicin/ cisplatin

Regimen Components Agent

Availability

Dose mg/m2

Paclitaxel (Bristol-Myers Squibb’s Taxol, generics)

on day US, F, G, 175 1 of a 21-day I, S, cycle of 4–10 UK, J cycles. IV.

Carboplatin (Bristol-Myers Squibb’s Paraplatin, generics) Paclitaxel (Bristol-Myers Squibb’s Taxol, generics)

US, F, G, AUC of 5.0–7.5 on I, S, day 1 of a 21-day UK, J cycle for a total of 4–10 cycles. IV.

Cisplatin (Bristol-Myers Squibb’s Platinol-AQ, generics) Cyclophosphamide (Bristol-Myers Squibb’s Cytoxan, Baxter’s Endoxan/ Endoxana, generics) Doxorubicin (Pfizer’s Adriamycin/ Adriblastine, Kyowa’s Adriacin, generics) Cisplatin (Bristol-Myers Squibb’s Platinol-AQ, generics)

US, F, G, 135 mg/m2 on day 1 of a 21-day I, S, cycle for a total of UK, J 3–6 cycles. IV.

US, F, G, 75 mg/m2 on day 2 of a 21-day cycle I, S, for a total of 3–6 UK, J cycles. IV. US, F, G, 350 mg/m2 on days 1 and 8 of a I, S, 28-day cycle for a UK, J total of 4–6 cycles. IV.

US, F, G, 20 mg/m2 on days 1 and 8 of a I, S, 28-day cycle for a UK, J total of 4–6 cycles. IV. US, F, G, 60 mg/m2 on days 1 and 8 of a I, S, 28-day cycle for a UK, J total of 4–6 cycles. IV.

Common Toxicities • Myelosuppression • Neurotoxicity • Thrombocytopenia • Gastrointestinal toxicity • Granulocytopenia • Anemia

• • • •

Leukopenia Neurotoxicity Hypersensitivity Gastrointestinal toxicity

• Alopecia • Renal toxicity

• Leukopenia • Alopecia • Gastrointestinal toxicity • Nausea • Thrombocytopenia

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TABLE 3. (continued) Regimen or Class

Regimen Components Agent

Gemcitabine/ Gemcitabine (Eli carboLilly’s Gemzar) platin

Docetaxel/ carboplatin

Availability

Dose

Common Toxicities

mg/m2

• Neutropenia and other hematological effects • Minor myelosuppression

on US, F, G, 1,000 days 1 and 8 of a I, S, 21-day cycle UK, J

Carboplatin US, F, G, AUC of 4 on day 1 (Bristol-Myers I, S, of a 21-day cycle Squibb’s UK, J Paraplatin, generics) Docetaxel US, F, G, 75 mg/m2 on day 1 of a 21-day cycle (Sanofi-Aventis’s I, S, for a total of 4–6 Taxotere) UK, J cycles. IV.

• Myelosuppression • Neutropenia • Thrombocytopenia • Gastrointestinal toxicity • Granulocytopenia • Anemia

Carboplatin (Bristol-Myers Squibb’s Paraplatin, generics) Bristol-Myers Squibb’s Paraplatin, generics Bristol-Myers Squibb’s Platinol-AQ, generics

US, F, G, AUC of 5 on day 1 I, S, of a 21-day cycle UK, J for a total of 4–10 cycles. IV.

Liposomal doxorubicin (single agent) Topotecan (single agent)

Ortho Biotech’s Doxil, ScheringPlough’s Caelyx/Myocet

US, F, G, 50 mg/m2 on day 2 of a 21-day cycle I, S, UK for a total of 6 cycles. IV.

GlaxoSmithKline/ Merck’s Hycamtin

US, F, G, 1.5 mg/m2 on days 1-5 of a 21-day I, S, UK cycle for a total of 4–6 cycles. IV.

• Neutropenia • Thrombocytopenia • Mild nonhematological toxicities

Docetaxel (single agent)

Sanofi-Aventis’ Taxotere

US, F, G, 100 mg/m2 on day 1 of a 21-day I, S, cycle for a total of UK, J 4–6 cycles. IV.

• Myelosuppression (leukopenia/neutropenia)/neutropenic infection • Neurotoxicity

Carboplatin (single agent) Cisplatin (single agent)

US, F, G, 300 mg/m2 on day 1 of a 28-day I, S, cycle for a total of UK, J 4–6 cycles. IV. US, F, G, 100 mg/m2 on day 1 of a 28-day I, S, cycle for a total of UK, J 6 cycles. IV.

• Neurotoxicity • Anemia • Gastrointestinal toxicity • Thrombocytopenia • Nephrotoxicity • Neutropenia • Anemia

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TABLE 3. (continued) Regimen or Class

Regimen Components Agent

Availability

Dose

Etoposide (single agent)

Bristol-Myers Squibb’s VePesid/ Etopophos, Pfizer’s Lastet, generics

US, F, G, 50 mg/m2 on day 1-21 of a 28-day I, S, cycle for a total of UK, J 6 cycles. PO.

Treosulfan (single agent) Altretamine (single agent)

Medac’s Ovastat, generics

G, UK

1–2 gm/day daily. PO.

US, UK

260 mg/m2 on day 1–14 of a 28-day cycle for a total of 12 cycles. PO.

(MGI Pharma/Ipsen’s Hexalen, ProStrakan’s Hexastat Cisplatin/ Cisplatin irinotecan (Bristol-Myers Squibb’s Platinol-AQ, generics) Irinotecan (Sanofi-Aventis’ Campto, Pfizer’s Camptosar) Irinotecan Sanofi-Aventis’ (single Campto, Pfizer’s agent) Camptosar

Tamoxifen (single agent)

AstraZeneca’s Nolvadex, Pfizer’s Kessar, generics

US, F, G, 60 mg/m2 on day 1 of a 28-day cycle. I, S, IV. UK, J

Common Toxicities • Myelosuppression • Leukopenia • Anemia • Secondary myelodysplasia • Mucositis • Nausea • Minor myelosuppression • Emesis • Myelosuppression • Anemia • Peripheral sensory neuropathy • Leukopenia • Neutropenia • Diarrhea

US, F, G, 60 mg/m2 on days 1, 8, and 15 of a I, S, 28-day cycle. IV UK, J US, F, G, 130 mg/m2 on day 1 of a 21-day I, S, cycle UK, J

US, F, G, 20 mg/day daily. I, S, PO. UK, J

• Diarrhea • Alopecia • Nausea • Vomiting • Cholinergic symptoms • Neutropenia • Thromboembolic events • Increased risk of endometrial cancer (2–7 times) • Hot flashes • Gastrointestinal toxicity

US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan. AUC = Area under curve. IV = Intravenous. PO = Orally.

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FIGURE 2. Structure of paclitaxel.

FIGURE 3. Structure of carboplatin.

A Phase III trial randomized 798 patients with International Federation of Gynecology and Obstetrics (FIGO) stage IIb-IV CaO to receive six cycles of either TC or TP (Du Bois A, 2003). The primary endpoint was the proportion of patients without progression at two years. Secondary endpoints included toxicity, response to treatment, quality of life, and overall and progression-free survival time. Quality of life was evaluated using the European Organization for Research and Treatment of Cancer quality-of-life questionnaire (EORTC QLQ C-30). The proportion of patients without progression at two years was not statistically signiÞcantly different between the two treatment arms (40.0% for TP versus 37.5% for TC). Median progression-free survival time in the TC arm (17.2 months) and the TP arm (19.1 months) was also not statistically much different; the same was true of median overall survival time (43.3 months versus 44.1 months, for the TC and TP arms, respectively). The TC regimen was associated with a higher frequency of hematologic toxicity but a lower frequency of gastrointestinal and neurological toxicity than the TP regimen. Mean global quality-of-life scores at the end of treatment were signiÞcantly better in the TC arm than in the TP arm (65.25 versus 51.97, respectively). Another trial of TP and TC, but in the adjuvant setting, similarly showed that no efÞcacy differences exist between TP and TC (Ozols FR, 2003); 792 FIGO stage III CaO patients with no residual mass greater than 1.0 cm after surgery received TC or TP. Gastrointestinal, renal, and metabolic toxicity, as well as National Cancer Institute Common Toxicity Criteria (NCI CTC) grade 4 (life-threatening) leukopenia, was signiÞcantly more frequent in patients receiving TP. Grade 2 (moderate) or greater thrombocytopenia was more common

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among patients receiving TC. Neurological toxicity was similar in both regimens. Median progression-free survival and overall survival were 19.4 and 48.7 months, respectively, for TP, compared with 20.7 and 57.4 months, respectively, for TC. However, in 2001, a third large trial found no survival beneÞt for the addition of paclitaxel to a platinum-based treatment. The International Collaborative Ovarian Neoplasm (ICON 3) study, a 2,075-patient, randomized Phase III trial, found that the three-year survival rate was 50% for both the taxane and nontaxane arms of the study. In this study, carboplatin rather than cisplatin was the platinum used (see discussion further on regarding the equivalence of cisplatin and carboplatin) (ICON Group, 2002). In light of these conßicting trial data, the inclusion of paclitaxel as part of Þrst-line treatment for CaO has become controversial in some countries. The use of carboplatin/paclitaxel for previously untreated patients with stage III/IV CaO who have undergone optimal or suboptimal cytoreductive surgery leaves room for improvement in terms of response rate and both progression-free and overall survival. Gynecologic Oncology Group (GOG) 182ICON 5 is a Þve-arm international collaborative study designed to improve on the efÞcacy of standard platinum/taxane therapy by incorporating newer cytotoxic agents in sequential doublet and triplet combinations. The Þve arms of this study are as follows: (1) four cycles of gemcitabine/carboplatin followed by four cycles of paclitaxel/carboplatin; (2) four cycles of liposomal doxorubicin/carboplatin followed by four cycles of paclitaxel/carboplatin; (3) four cycles of topotecan/carboplatin followed by four cycles of paclitaxel/carboplatin; (4) eight cycles of gemcitabine/paclitaxel/carboplatin; and (5) eight cycles of paclitaxel/carboplatin, acting as the control arm. Recruitment for the trial closed in August 2004 after 4,000 women were recruited worldwide. Paclitaxel/Cisplatin Overview. Cisplatin (Figure 4) is approved for Þrst- and second-line therapy in combination with other chemotherapeutic agents for the treatment of metastatic CaO. One of the Þrst pivotal studies establishing the role of platin-based chemotherapy in CaO treatment used TP. This regimen remains very popular because of its established efÞcacy and its lower price compared with that of TC. The main drawbacks are that cisplatin is considered slightly more toxic than carboplatin and it requires patient hydration. Mechanism of Action. The individual components of the paclitaxel/cisplatin regimen contribute the following mechanisms to achieve its overall activity: •

Paclitaxel is a taxane. Taxanes inhibit the dynamic reorganization of microtubule networks that is essential to vital interphase and mitotic cellular functions. Taxanes exhibit a unique mechanism of action. Paclitaxel, which

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FIGURE 4. Structure of cisplatin.

•

achieves selective toxicity against rapidly proliferating cells, is mainly active in the G2/M phase of the cell cycle. Cisplatin is a platin. Platins inhibit tumor-cell replication by creating intraand interstrand DNA cross-links. Early studies suggested that cisplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle.

Clinical Performance. The combination of a platin with paclitaxel became the gold-standard treatment for CaO in 1996, when the Gynecological Oncology Group (GOG 111) study found a signiÞcant survival beneÞt for the TP regimen. The GOG 111 study randomized more than 400 patients to receive either TP or cyclophosphamide/doxorubicin/cisplatin (CAP) (McGuire WP, 1996). Researchers found that TP was signiÞcantly more effective and better tolerated than CAP. In the TP arm, the median disease-free survival period was 18 months (versus 13 months for CAP), and the median overall survival time was 38 months (versus 24 months). Two years later, a large multicenter trial in Europe and Canada conÞrmed these Þndings (Stuart G, 1998). As noted in the previous section, Phase III data comparing TP with TC have shown that both regimens have similar efÞcacies, but TC has a better toxicity proÞle. A Phase III trial randomized 798 patients with FIGO stage IIb-IV CaO to receive six cycles of either TC or TP (Du Bois A, 2003). The primary endpoint was the proportion of patients without progression at two years. Secondary endpoints included toxicity, response to treatment, quality of life, and overall and progression-free survival time. Quality of life was evaluated using the EORTC QLQ C-30. The proportion of patients without progression at two years was not statistically signiÞcantly different between the two treatment arms (40.0% for TP versus 37.5% for TC). Median progression-free survival time in the TC arm (17.2 months) and the TP arm (19.1 months) was also not statistically signiÞcantly different; the same was true of median overall survival time (43.3 months versus 44.1 months for the TC and TP arms, respectively). The TC regimen was associated with a higher frequency of hematologic toxicity but a lower frequency of gastrointestinal and neurological toxicity than the TP regimen. Mean global quality-of-life scores at the end of treatment were signiÞcantly better in the TC arm than in the TP arm (65.25 versus 51.97, respectively). Another trial of TP and TC, but in the adjuvant setting, similarly showed that no efÞcacy differences exist between TP and TC (Ozols FR, 2003). In this trial, 792 CaO patients with no residual mass greater than 1.0 cm after surgery received TC or TP. Gastrointestinal, renal, and metabolic toxicity, as well as grade 4 leukopenia, was signiÞcantly more frequent in patients receiving TP. Grade 2 or higher

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thrombocytopenia was more common among patients receiving TC. Neurological toxicity was similar in both regimens. Median progression-free survival and overall survival were 19.4 and 48.7 months, respectively, for TP compared with 20.7 and 57.4 months, respectively, for TC. However, in 2001, a third large trial found no survival beneÞt for the addition of paclitaxel to a platinum-based treatment. The ICON 3 study, a 2,075-patient, randomized Phase III trial, found that the three-year survival rate was 50% for both the taxane and nontaxane arms of the study. In this study, carboplatin rather than cisplatin was the platinum (ICON Group, 2002). In light of these conßicting trial data, the inclusion of paclitaxel as part of Þrst-line treatment for CaO has become controversial in some countries. Cyclophosphamide/Doxorubicin/Cisplatin Overview. The agents cyclophosphamide (BMS’s Cytoxan, generics) (Figure 5), doxorubicin (PÞzer’s Adriamycin, generics) (Figure 6), and cisplatin (Figure 4) make up the CAP regimen. Until 1996, when the GOG published its data on the taxane-based regimens, platin-based CAP was the regimen of choice for CaO treatment.

Cl O

O P

N

NH Cl FIGURE 5. Structure of cyclophosphamide.

FIGURE 6. Structure of doxorubicin (R = OCH3 , R1 = OH, R2 = H, R3 = H, R4 = OH).

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Mechanism of Action. The individual components of the cyclophosphamide/ doxorubicin/cisplatin regimen contribute the following mechanisms to achieve its overall activity: •

•

•

Cyclophosphamide is an alkylating agent. Alkylating agents exert their biologic activity via covalent binding and cross-linking of a variety of macromolecules including DNA, RNA, and proteins. DNA cross-linking, probably the most important biologic action of these drugs, impairs DNA replication and transcription, ultimately leading to either cell death or altered cellular function. Doxorubicin is an anthracycline. Anthracyclines interact with several different cellular targets, most importantly topoisomerase II. By inhibiting this DNA re-ligation enzyme, anthracyclines exert their cytotoxic effect. Another mechanism leading to cell death, known as DNA intercalation, involves insertion of the anthracycline molecule between base pairs. This phenomenon causes single- and double-stranded breaks in DNA that inhibit cell proliferation. The free radicals generated by the reductive metabolism of anthracyclines may also damage cellular structures. Cisplatin is a platin. Platins inhibit tumor-cell replication by creating intraand interstrand DNA cross-links. Early studies suggested that cisplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle.

Clinical Performance. The CAP regimen’s declining use follows the publication of numerous clinical trials showing that it is more toxic than other regimens without offering any signiÞcant efÞcacy beneÞt. One of the most notable of these trials is the GOG 111 Phase III study, which randomized more than 400 FIGO stage III/IV patients to receive either TP or CAP (McGuire WP, 1996). Researchers found that the former regimen was signiÞcantly more effective and better tolerated than CAP. In the TP arm, the median disease-free survival period was 18 months (versus 13 months for CAP), and the median overall survival time was 38 months (versus 24 months). Two years later, a large multi-center trial in Europe and Canada conÞrmed this Þnding (Stuart G, 1998). The ICON 2 Phase III trial randomized 1,526 CaO patients (77% were FIGO stages III and IV) to receive either single-agent carboplatin or CAP (ICON Collaborative Group, 1998). After 728 patients died, survival curves showed no evidence of a difference between CAP and carboplatin. Median survival was 33 months and two-year survival was 60% for both groups. No efÞcacy difference between treatment groups in different subgroups deÞned by age, stage, residual disease, differentiation, histology, and coordinating center was evident. CAP was much more toxic than carboplatin, causing more alopecia, leukopenia, and nausea. Carboplatin caused more thrombocytopenia. Similar data came from the ICON 3 Phase III study, which randomized 2,075 FIGO stage I–IV patients to receive either carboplatin alone, TC, or CAP (Columbo N, 2000). Results showed that none of the treatment arms was more

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or less effective than the others. The three-year survival rate was 50% for both the carboplatin and TC arms of the study. Gemcitabine/Carboplatin Overview. Gemcitabine (Eli Lilly’s Gemzar) (Figure 7) is used both as a single agent and in combination. However, it is approved only for the treatment of CaO within the regimen gemcitabine/carboplatin, and the only major market in which it is approved is Germany. Gemcitabine is used off-label in other markets. Mechanism of Action. The individual components of the gemcitabine/ carboplatin regimen contribute the following mechanisms to achieve its overall activity: •

•

Gemcitabine is an antimetabolite. Antimetabolites block normal DNA synthesis, thus stopping cell replication. Gemcitabine exhibits cell-phase speciÞcity, primarily killing cells that are undergoing DNA synthesis (S phase) and blocking the progression of cells through the G1/S-phase boundary. Carboplatin is a platin. Platins inhibit tumor-cell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that carboplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle.

Clinical Performance. The Phase III trial that led to Germany’s approval of gemcitabine/carboplatin randomized 356 patients with recurrent platinum-sensitive CaO to receive six cycles of either gemcitabine/carboplatin or carboplatin every 21 days (PÞsterer J, 2005[b]). With a median follow-up of 17 months, median progression-free survival was 8.6 months for gemcitabine/carboplatin and 5.8 months for carboplatin. The response rate for the gemcitabine/carboplatin group was 47.2%; the response rate for the carboplatin group was 30.9%. Patients treated with gemcitabine/carboplatin reported signiÞcantly faster palliation of abdominal symptoms and a signiÞcantly improved global quality of life. Gemcitabine is well tolerated and produces only mild side effects, such as minor myelosuppression. In two small studies employing gemcitabine in refractory or recurrent advanced CaO, 13% of patients responded to treatment NH2 N HO

O O

OH

N F F

FIGURE 7. Structure of gemcitabine.

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(Friedlander M, 1998; Hansen SW, 1999[a]). In contrast, a gemcitabine/cisplatin combination and a gemcitabine/paclitaxel combination induced responses of 53% and 40%, respectively (Bauknecht T, 1998; Poole CJ, 1998). In previously untreated patients with a poor prognosis, use of gemcitabine alone produced a 24% response rate, but when the drug was given in combination with cisplatin, remissions occurred in 53–71% of patients. Dose-limiting toxicity is mainly hematological and includes neutropenia. Docetaxel/Carboplatin Overview. The taxane docetaxel (SanoÞ-Aventis’s Taxotere) (Figure 8) has not been approved for the treatment of CaO in any of the major markets except Japan, but it is used off-label for patients with advanced epithelial CaO who have developed neurotoxicity during treatment with paclitaxel. Usually, it is combined with carboplatin. Mechanism of Action. The individual components of the docetaxel/ carboplatin regimen contribute the following mechanisms to achieve its overall activity: •

•

Docetaxel is a taxane. Taxanes inhibit the dynamic reorganization of microtubule networks that is essential to vital interphase and mitotic cellular functions. Taxanes exhibit a unique mechanism of action. Docetaxel, which achieves selective toxicity against rapidly proliferating cells, is mainly active in the G2/M phase of the cell cycle. Carboplatin is a platin. Platins inhibit tumor-cell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that carboplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle.

Clinical Performance. A multi-center randomized trial comparing docetaxel/ carboplatin with paclitaxel/carboplatin as Þrst-line treatment in 1,077 patients found equivalence in terms of response rates and progression-free survival and the expected differences in toxicity. With a median follow-up of 23 months, median

FIGURE 8. Structure of docetaxel.

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progression-free survival was 15.4 and 15.1 months for the docetaxel/carboplatin and paclitaxel/carboplatin regimens, respectively. Two-year survival was 69.8% and 65.4%, respectively. Paclitaxel caused signiÞcantly more neurotoxicity (30% versus 11%), leading to early treatment discontinuation. Although docetaxel demonstrated a higher incidence of neutropenia, this side effect was not associated with an increase in toxic deaths or treatment discontinuation (Kaye SB, 2001). Docetaxel is slightly more convenient to deliver than paclitaxel because it can be administered as a one-hour infusion once every three weeks. Several clinical trials have examined docetaxel for the management of platinum-chemotherapy-resistant CaO. Researchers documented a response rate of approximately 20-35% in this setting (Piccart MJ, 1995; Kavanagh JJ, 1999). Carboplatin, Single Agent Overview Carboplatin monotherapy is a standard treatment option for CaO, most notably as a second-line treatment in patients with a response longer than 6-12 months to Þrst-line treatment but who then relapse (i.e., platinum-sensitive recurred patients). Compared with cisplatin, carboplatin is better tolerated and easier to administer (typically as a one-hour infusion rather than a six- to eighthour infusion) and offers similar antitumor activity (Aabo K, 1998). Carboplatin does not require hydration and can be administered on an outpatient basis. However, its major and dose-limiting adverse effect—myelosuppression—is more pronounced than with cisplatin. Mechanism of Action. Carboplatin is a platin. Platins inhibit tumor-cell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that carboplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown it to have complex and variable effects on the cell cycle. Clinical Performance. Randomized data on the use of single-agent carboplatin in CaO treatment are conßicting. A meta-analysis performed in 1998 found no difference in survival among patients treated with single-agent carboplatin or cisplatin (Aabo K, 1998). Initial data supporting single-agent carboplatin came from the ICON 2 Phase III trial, which randomized 1,526 CaO patients to receive either singleagent carboplatin or CAP (ICON Collaborative Group, 1998). After 728 patients died, survival curves showed no evidence of a difference between CAP and carboplatin. Median survival was 33 months and two-year survival was 60% for both groups. No efÞcacy difference between treatment groups in different subgroups deÞned by age, stage, residual disease, differentiation, histology, and coordinating center was evident. CAP was substantially more toxic than carboplatin, causing more alopecia, leukopenia, and nausea. Carboplatin caused more thrombocytopenia. Similar data came from the ICON 3 Phase III parallel study, which randomized 2,075 FIGO stage I–IV patients to receive either TC, carboplatin, or CAP (the

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control arm) (Columbo N, 2000). Results showed that none of the treatment arms was more or less effective than the others. The three-year survival rate was 50% for both the carboplatin and TC arms of the study. However, the ICON and Arbeitsgemeinschaft Gynaekologische Onkologie (AGO) groups’ ICON 4/AGO-OVAR-2.2 Phase III study did show an improvement in the second-line setting in the efÞcacy of a platinum-containing agent (platin) plus paclitaxel over carboplatin alone or any other regimen that does not contain paclitaxel (ICON and AGO Collaborators, 2003). In the paclitaxelcontaining multidrug regimen group, 80% of patients received TC, 10% received TP, and 10% received other regimens. In the “conventional” non-paclitaxelcontaining arm, 70% received carboplatin alone, 17% received the CAP regimen, and 13% received other regimens. With a median follow-up of 42 months, 530 patients died. Survival data showed a difference in favor of paclitaxel plus platin corresponding to an absolute difference in two-year survival of 7% between the paclitaxel and conventional treatment groups (57% versus 50%) and median survival of Þve months (29 versus 24 months). An absolute difference in one-year progression-free survival of 10% (50% versus 40%) and in median progressionfree survival of three months (13 versus 10 months) was also observed. Cisplatin, Single Agent Overview. Cisplatin is approved for Þrst- and second-line therapy in combination with other chemotherapeutic agents for the treatment of metastatic CaO. Although cisplatin is approved only for treating CaO in combination regimens, single-agent cisplatin is sometimes used because it is inexpensive and has demonstrated reasonable efÞcacy in randomized trials. Mechanism of Action. Cisplatin is a platin. Platins inhibit tumor-cell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that cisplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown it to have complex and variable effects on the cell cycle. Clinical Performance. A Phase III trial showed that single-agent cisplatin has efÞcacy comparable to that of paclitaxel/cisplatin but a worse toxicity proÞle (Muggia FM, 2000). In this trial, patients with suboptimally debulked FIGO stage III/IV CaO received either cisplatin (100 mg/m2 ), 24-hour infusion paclitaxel (200 mg/m2 ), or paclitaxel (135 mg/m2 ) followed by cisplatin (75 mg/m2 ). Patients (n = 614) were randomized to receive six cycles of one of the treatments every three weeks. Monotherapies were discontinued more frequently (cisplatin because of toxicity or patient refusal [17%] and paclitaxel because of progression [20%]) than was combination therapy (7% and 6%, respectively). Neutropenia, fever, and alopecia were more severe with paclitaxel-containing regimens, while anemia, thrombocytopenia, neurotoxicity, nephrotoxicity, and gastrointestinal toxicity were more severe with cisplatin-containing regimens. The overall response rate on paclitaxel monotherapy (42%) was signiÞcantly lower than that of the cisplatin regimens (67%). Median overall survival—30.2 months, 25.9 months, and 26.3 months for cisplatin, paclitaxel, and combination arms,

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respectively—was not statistically different. Relative to cisplatin, the death rate on paclitaxel was 15% higher and the death rate on the combination treatment was 1% less. These differences among treatment groups were not statistically signiÞcant. Liposomal Doxorubicin, Single Agent Overview. Liposomal doxorubicin (Ortho Biotech’s Doxil in the United States, Schering-Plough’s Caelyx/Myocet in Europe) was approved in the United States in 1999 for relapsed or treatment-refractory advanced-stage CaO. The drug was approved in Europe in 2000 (after having been approved for Kaposi’s sarcoma) for the same indication. The approval was based on results from Phase II trials and interim results from a Phase III clinical trial. Liposomal doxorubicin is not available in Japan. Mechanism of Action. Doxorubicin is an anthracycline. Anthracyclines interact with several cellular targets, most importantly topoisomerase II. By inhibiting this DNA religation enzyme, anthracyclines exert their cytotoxic effect. Another mechanism leading to cell death, known as DNA intercalation, involves insertion of the anthracycline molecule between base pairs. This phenomenon causes single- and double-stranded breaks in DNA that inhibit cell proliferation. The free radicals generated by the reductive metabolism of anthracyclines may also damage cellular structures. Formulation. Liposomal doxorubicin uses “stealth” liposome technology, in which polyethylene glycol (PEG) is incorporated into the liposomal membrane surrounding the drug. This modiÞcation prevents extensive uptake of the drug by cells of the reticuloendothelial system (e.g., macrophages and reticular and endothelial cells of the bone marrow). The half-life of the liposomal formulation is longer than that of conventional doxorubicin, so the agent is cleared more slowly from plasma. The liposome technology keeps the drug localized to the tumor by means of direct release into the tumor. These two features substantially reduce the systemic toxicities associated with the conventional formulation of doxorubicin (Martin FJ, 1997). Clinical Performance. A Phase III trial showed that liposomal doxorubicin is more efÞcacious than topotecan (GlaxoSmithKline/Merck’s Hycamtin) (Figure 9) in the treatment of patients with recurrent, platinum-sensitive disease (Gordon AN, 2004). CaO patients (n = 474) whose disease recurred after or failed to respond to Þrst-line platinum-based chemotherapy received either liposomal doxorubicin (50 mg/m2 every 28 days) or topotecan (1.5 mg/m2 per day for 5 days every 21 days). Most patients had been previously treated with platinum and taxanes (74% in the pegylated liposomal doxorubicin group and 72% in the topotecan group). Patients treated with liposomal doxorubicin experienced an 18% reduction in the risk of death (median survival was 62.7 weeks for liposomal doxorubicin and 59.7 weeks for topotecan-treated patients). Patients with platinum-sensitive

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CH3 N CH3 HO

O N N O HO H3C

O

FIGURE 9. Structure of topotecan.

disease who were treated with liposomal doxorubicin experienced a 30% reduction in the risk of death (median survival 107.9 weeks for pegylated liposomal doxorubicin versus 70.1 weeks for topotecan-treated patients). Among patients with platinum-refractory disease, survival was similar between treatment groups. A Phase II trial involved 35 CaO patients whose disease did not respond to initial paclitaxel/platinum therapy (Gordon AN, 1998). Nine of the 35 women (26%) responded to liposomal doxorubicin, and one of these responses was complete. The median duration of progression-free survival was 5.7 months; the median duration of overall survival was 11 months. Liposomal doxorubicin appeared to alleviate the cardiotoxicity, alopecia, and neurotoxicity associated with conventional doxorubicin, although it reduced neutropenia only slightly. Additionally, 13 of the 35 patients experienced nonhematologic skin and mucosal toxicities such as stomatitis. Topotecan, Single Agent Overview. Topotecan (GlaxoSmithKline/Merck’s Hycamtin) (Figure 9) is approved in the United States and Europe as second-line chemotherapy for advanced CaO. It is available in Japan for other malignancies but is not currently approved for treating CaO. Mechanism of Action. Topotecan is a topoisomerase I inhibitor. It binds to the topoisomerase I DNA complex and prevents religation of DNA singlestrand breaks that occur naturally during DNA synthesis. Researchers believe the cytotoxicity of topotecan results from double-strand DNA damage produced during DNA synthesis, when replication enzymes interact with the ternary complex formed by topotecan, topoisomerase I, and DNA. Mammalian cells cannot efÞciently repair these double-strand breaks. Clinical Performance. The FDA gave topotecan fast-track clearance for the treatment of CaO based on a multi-center European study that compared topotecan with paclitaxel as second-line therapy (ten Bokkel Huinink W, 1997). The trial involved 226 patients with recurrent disease who had been treated with

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a platinum-based regimen. Topotecan was administered to 112 patients in a 30-minute infusion of 1.5 mg/m2 /day for Þve days. This treatment elicited a 20.5% response rate; paclitaxel produced a statistically equivalent response rate of 13.2%. The median response duration for topotecan was 32 weeks, compared with 20 weeks for paclitaxel. The duration of patient survival was 61 weeks with topotecan versus 42 weeks with paclitaxel—Þndings that were also statistically equivalent. However, the incidence of neutropenia was signiÞcantly higher in patients treated with topotecan—81.4% versus 22.9% of patients receiving paclitaxel. With topotecan, hematological toxicity usually occurs during the Þrst cycle and can be managed with dose reduction and/or introduction of hematopoietic growth factors; nonhematologic toxicities are generally mild and not doselimiting. In the same randomized study of second-line therapy, researchers assessed the degree of non-cross-resistance between topotecan and paclitaxel because 110 patients crossed over to the alternative agent as third-line therapy (Gore M, 2001). Sixty-one patients crossed over from paclitaxel to topotecan; 49 patients switched from topotecan to paclitaxel. Their overall response rate to topotecan after paclitaxel was 13% and to paclitaxel after topotecan, 10%. Hence, patients who have disease that is resistant to topotecan or to paclitaxel have a small chance of responding to the other drug. Trial data for topotecan in the Þrst-line setting have not been promising. A Phase III trial randomized 1,308 FIGO stages IIb–IV patients with previously untreated CaO to receive six cycles of paclitaxel/carboplatin (PÞsterer J, 2005[a]), followed either by observation or four cycles of topotecan. The primary endpoint was overall survival. The following grade 3 (severe) and 4 toxicities were observed with paclitaxel/carboplatin followed by topotecan: anemia in 6.4% of patients, thrombocytopenia in 10.7%, and neutropenia in 57%. Complete responses were seen in 69% of the topotecan group versus 76.2% of the observation group. Median progression-free survival was 18.2 months in the topotecan group versus 18.5 months in the observation group. Median overall survival was 43.1 months versus 44.5 months for the topotecan and observation groups, respectively. A Phase III trial showed that liposomal doxorubicin is more efÞcacious than topotecan in the treatment of patients with recurrent, platinum-sensitive CaO (Gordon AN, 2004). Patients (n = 474) whose disease recurred after or failed to respond to Þrst-line platinum-based chemotherapy received either liposomal doxorubicin (50 mg/m2 every 28 days) or topotecan (1.5 mg/m2 per day for 5 days every 21 days). Most patients had been previously treated with platinum and taxanes (74% in the pegylated liposomal doxorubicin group and 72% in the topotecan group). Patients treated with liposomal doxorubicin experienced an 18% reduction in the risk of death (median survival 62.7 weeks for liposomal doxorubicin versus 59.7 weeks for topotecan-treated patients). Patients with platinum-sensitive disease who were treated with liposomal doxorubicin experienced a 30% reduction in the risk of death (median survival 107.9 weeks for pegylated liposomal

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doxorubicin and 70.1 weeks for topotecan-treated patients). Among patients with platinum-refractory disease, survival was similar between treatment groups. Nonpharmacological Approaches The function of surgery in CaO includes diagnosis, staging, potential cure (via tumor removal in the case of early-stage disease), cytoreduction (in bulky disease), and palliation. A key surgical principle is to remove as much of the tumor as is safely possible. Removal of the ovarian mass or masses (surgical debulking) is followed by biopsies. If biopsy samples reveal a malignancy, removal of the uterus (total abdominal hysterectomy [TAH]) and both fallopian tubes and ovaries (bilateral salpingo-oophorectomy [BSO]) is carried out unless contraindicated by the patient’s reproductive status. The omentum, which is located at the top of the peritoneal cavity, is removed by a procedure called an omentectomy. Any areas of the primary tumor that adhere to the peritoneum are biopsied; pelvic and para-aortic lymph nodes are dissected (lymphadenectomy). In advanced disease (stages III and IV), removing all of the malignancy is not usually possible. In such cases, the surgeon excises as much as possible to leave minimum residual disease. Although many studies have shown that aggressive removal of tumor tissue in advanced disease improves survival time, a metaanalysis of 58 studies found no statistically signiÞcant increase in survival time (Hunter RW, 1992). EMERGING THERAPIES The gold-standard chemotherapy for previously untreated patients with ovarian cancer (CaO) is a combination of paclitaxel (Bristol-Myers Squibb [BMS] Taxol, generics) and carboplatin (BMS’ Paraplatin, generics). However, most patients still relapse, and more than 70% of patients have stage III or IV disease at the time of diagnosis, a fact that translates into a poor Þve-year survival of 20%. Based on advances in proteomics and DNA microarray analysis, recent advances in the understanding of the biology and genetics of CaO have led to the identiÞcation of multiple molecular targets that may soon change the standard treatment of CaO. Several of these molecularly targeted (biologic) agents have entered clinical trials, including angiogenesis inhibitors, epidermal growth factor inhibitors, and immunotherapies. These products are discussed in the following sections. Ultimately, the success of CaO therapy lies not just in the availability of new agents but in the ability to identify patients with biomarkers that may predict their response to these agents. This section discusses the agents in late-stage development for CaO, including those compounds in more advanced Phase II studies. Table 4 lists late-stage emerging therapies for CaO, and Table 5 identiÞes promising agents in early-stage development. Agents in development for the treatment of CaO may be granted orphan drug status for this indication. The accompanying sidebar, “Orphan Drug Status in the Major Markets,” explains the commercial signiÞcance of orphan drug status.

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TABLE 4. Emerging Therapies in Development for Ovarian Cancer Compound Novel taxanes CT-2103 (PG-TXL, Xyotax) United States Europe Japan BMS-184476 United States Europe Japan Novel platinum compounds Oxaliplatin (Eloxatin) United States Europe Japan Novel alkylating agents TLK-286 ( Telcyta) United States Europe Japan Trabectedin ( ET-743, Yondelis) United States Europe Japan Immunotherapies IDM-1 (Osidem) United States Europe

Development Phase

III III PC

Cell Therapeutics Cell Therapeutics Chugai

II II —

Bristol-Myers Squibb Bristol-Myers Squibb —

II III —

Sanofi-Aventis Sanofi-Aventis —

III — —

Telik — —

III III —

Ortho Biotech PharmaMar/Ortho Biotech —

III

Immuno-Designed Molecules/Sanofi-Aventis Immune-Designed Molecules/Sanofi-Aventis —

III

Japan — Interferon gamma-1b (Actimmune, Imukin) United States III Europe

Japan Oregovomab (OvaRex) United States Europe Japan Angiogenesis inhibitors Bevacizumab (Avastin) United States Europe Japan Thalidomide (Thalomid) United States Europe Japan

Marketing Company

—

InterMune Pharmaceuticals InterMune Pharmaceuticals/Boehringer Ingelheim —

III II —

ViRexx/Unither Pharmaceuticals ViRexx/Dompe Farmaceutici —

III — —

Genentech — —

III — —

Celgene — —

III

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TABLE 4. (continued) Compound Carboxyamido-triazole (CAI) United States Europe Japan Squalamine United States Europe Japan Epidermal growth factor inhibitors Gefitinib (Iressa) United States Europe Japan Erlotinib (Tarceva) United States Europe Japan Cetuximab (Erbitux) United States Europe Japan Pertuzumab (Omnitarg) United States Europe Japan

Development Phase

Marketing Company

II — —

National Cancer Institute — —

II — —

Genaera — —

II — —

AstraZeneca — —

II III —

Osi Pharmaceuticals/Genentech Osi Pharmaceuticals/Genentech —

II — —

ImClone/Bristol-Myers Squibb/ Merck KGaA — —

II II PC

Genentech Roche Chugai

PC = Preclinical. Note: Development phase is based on databases such as R & D Focus, R & D Insight, Pharmaprojects, and the Investigational Drugs Database (IDdb3); periodicals such as Scrip, the FDC’s Pink Sheet, and Marketletter; company reports and press releases; and industry contacts.

The introduction of novel, highly targeted molecules may enable more “intelligent” treatment of CaO. The potential of these agents lies in two avenues: single-agent activity and combination with chemotherapy. Signaling pathways that play a key role in response to chemotherapy and that may be particularly relevant in CaO include the P13K/AKT pathway (see “Etiology and Pathophysiology”). New speciÞc P13K inhibitors are at an advanced preclinical stage and are set to enter clinical trials shortly. Experimental data have shown that the molecular chaperone heat shock protein-90 (HSP-90) sensitizes taxol-resistant CaO cells to cancer through its impact on the P13K/AKT pathway (Sain N, 2004). Therefore, combinations of the HSP-90 inhibitor 17 allyl amino geldanamycin (17AAG) and taxol are being planned. Another targeted therapy worth mentioning is imatinib mesylate (Novartis’ Gleevec/Glivec). Imatinib is a receptor kinase inhibitor with speciÞcity for abl, platelet-derived growth factor receptor (PDGFR), and c-kit. It is approved by the FDA for the treatment of chronic myelogenous leukemia and gastrointestinal

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TABLE 5. Select Investigational Therapies in Early-Stage Development for Ovarian Cancer Compound Cytotoxic agents 3-AP (Triapine)

Development Phasea

Developing Companies

Comments

II (US)

Vion Pharmaceuticals

• 3-AP is an IV formulation of 3-AMP and a ribonucleotide reductase inhibitor. • The compound is in development for the potential treatment of leukemia and a variety of solid tumors.

17-AAG

Preclinical (US)

Kosan Biosciences

• 17-allylaminogeldanamycin is an injectable semi-synthetic derivative of geldanamycin and a polyketide inhibitor of Hsp-90. • In human ovarian tumor cell lines, 17AAG and paclitaxel were synergistic.

A-6

II (US/UK)

Angstrom Pharmaceutical

• A-6 is a small peptide therapeutic that induces apoptosis. • A-6 has been shown to inhibit cancer metastasis in animal studies and showed ‘‘encouraging responses’’ in patients with advanced CaO in Phase I clinical trials.

ABI-007 (Abraxane)

Launched for breast cancer (US)

American Pharmaceutical Partners

• ABI-007 is a nanoparticle formulation of the cytotoxic paclitaxel. • ABI-007 is in development in collaboration with the M.D. Anderson Cancer Center. • Experts suggest considerable potential in ovarian cancer.

Atrasentan

II (US)

Abbott

• Atrasentan is a selective endothelin ET-A receptor antagonist. • Atrasentan acts by inhibiting growth and potentiates activity of paclitaxel in ovarian cancer cells. • The compound is in preregistration for the treatment of prostate cancer.

Bortezomib (Velcade)

II (UK,US)

Millennium Pharmaceuticals

• Bortezomib inhibits tumor cell ubiquitin proteasome activity and induces apoptosis. • It displayed synergy with docetaxel in an ovarian xenograft model (SKOV3). • Bortezomib’s development is across a range of cancers. The compound is launched for the treatment of multiple myeloma.

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TABLE 5. (continued) Compound

Development Phasea

Developing Companies

Comments

Capecitabine (Xeloda)

II (US)

Roche

• Capecitabine is a cytotoxic oral prodrug of 5-fluorouracil (5-FU). • The drug is launched for the treatment of breast and colorectal cancers.

CT-2106 (polyglutamate camptothecin)

II (US)

Cell Therapeutics

• CT-2106 is camptothecin linked to polyglutamate. • CT-2106 is welltolerated and does not exhibit the gastrointestinal toxicities commonly associated with camptothecins.

Etanercept (Enbrel)

II (UK)

Amgen

• Etanercept affects tumor pathogenesis by tumor necrosis factor (TNF) antagonism. • It was launched for rheumatoid arthritis and is under investigation across a broad range of diseases. • In 17 patients treated twice weekly with 25 mg subcutaneously for 3 months, SD was seen in 2; median overall survival was 9.6 months.

FK-228 (FR901228)

II (US)

Gloucester Pharmaceuticals/NCI

• FK-228 is a histone deacetylase inhibitor. • The compound is under license from Fujisawa Pharmaceuticals. • FK-228 is in Phase III trials for the treatment of cutaneous T-cell lymphoma.

Irofulven

II (US)

MGI Pharma

• Irofulven binds to cellular macromolecules and inhibits DNA synthesis. • Of 55 evaluable refractory patients, 17 had SD, 1 complete response, and 9 partial responses; partial responses remained progression-free for up to 20 months. • Dainippon is the irofulven development partner in Japan.

Ispinesib

II (US)

Cytokinetics/ GlaxoSmithKline

• Ispinesib is a cytotoxic, mitotic kinesin spindle protein (KSP) inhibitor. • The compound is also in Phase II trials for a variety of other solid tumors.

Ixabepilone

II (US)

Bristol-Myers Squibb

• Ixabepilone is one of a series of epothilone tubulin inhibitors licensed from the German Research Centre for Biotechnology. • The compound is in Phase III trials for the treatment of breast cancer.

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TABLE 5. (continued) Compound

Development Phasea

Developing Companies

Comments • Protaxel is a cytotoxic, paclitaxel prodrug. • A study showed significant effects in paclitaxel-resistant ovarian tumor cell lines. • Patupilone is an intravenous formulation of the microtubule depolymerization inhibitor (microtubule stabilizer) epothilone B. • In a Phase II trial, 32 patients with relapsed/refractory ovarian cancer received a 5- to 20-minute IV infusion of patupilone (6.5 to 11 mg/m2 ) once every 3 weeks (1 cycle). The most common adverse effects reported were diarrhea, nausea, vomiting, and fatigue. One patient had a CR, seven had a PR, 8 had stable disease, and 16 had progressive disease.

Paclitaxel ceribate (Protaxel)

I (US)

Biophysica Foundation

Patupilone

II (US)

Novartis

Pemetrexed (Alimta)

II (US)

Eli Lilly & Co.

• Pemetrexed is a thymidylate synthase and transferase inhibitor. • The compound is launched for the treatment of NSCLC and mesothelioma.

Rubitecan (Camptogen)

II (US)

Supergen

• Rubitecan is an orally active topoisomerase I inhibitor. • Rubitecan had a 12% response rate (2 PR and 10 SD) in 17 evaluable refractory ovarian cancer patients. • The agent is licensed from the Stehlin Foundation for Cancer Research. • The compound is in preregistration for the treatment of pancreatic cancer.

CellControl Biomedical

• The agent is an anti-idiotypic MAb that induces a specific immune reaction against CA-125. • In 42 relapsed patients, survival in Ab3-positive patients (28) was 19.9 months versus 5.3 months in Ab3-negative responders (14).

NCI

• The agent is an immunotoxin composed of a single-chain antimesothelin MAb linked to Pseudomonas exotoxin PE-38. • Orphan drug status was granted in February 2002 in the United States.

Immunotherapeutics ACA-125 II (Germany)

SS (dsFv) PE38

II (US)

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TABLE 5. (continued) Compound

Development Phasea

Angiogenesis inhibitors Fenretinide II (US)

Developing Companies

Comments

McNeil (Johnson & Johnson)/ NIH

• Fenretinide, an oral retinoid, inhibits growth of IGF-I and induces apoptosis in human ovarian cancer cell lines. • Out of 22 patients who were administered 900 mg/m2 of fenretinide, there were no objective responses and eight patients achieved SD. The median PFS was 1.5 months and the median survival was 9.2 months. • The compound is in Phase III trials for the treatment of breast and bladder cancers.

Bayer/Onyx

• Sorafenib is an oral cytostatic Raf kinase inhibitor, PDGFR inhibitor, and VEGFR-2 inhibitor. • The compound is in preregistration for the treatment of renal cancer.

Growth factor inhibitors Imatinib II (US) (Gleevec)

Novartis

• Imatinib inhibits tyrosine kinases, including Bcr-Abl and PDGFR. • Imatinib inhibits the growth of CaO cells through PDGFR inactivation. In addition, in vitro results suggest that constitutive Akt activation modulates sensitivity to imatinib in CaO cells. • The compound is launched for the treatment of chronic myelogenous leukemia in several major markets and for gastrointestinal tumors in the United States.

Lapatinib

GlaxoSmithKline

• Lapatinib is an ErbB2 and EGFR dual tyrosine kinase inhibitor. • Lapatinib is considered to be in the next generation of EGFR inhibitors. • The compound is in Phase III trials for the treatment of breast cancer.

Sorafenib

II (US)

II (US)

a Development phase reflects status for ovarian cancer unless otherwise indicated.

CR = Complete response. EGFR = Epidermal growth factor receptor. IGF = Insulin growth factor. IV = Intravenous. PR = Partial response. NCI = National Cancer Institute. NIH = National Institute of Health. NSCLC = Non-small-cell lung cancer. PDGFR = Platelet-derived growth factor receptor. SD = Stable disease. VEGFR-2 = Vascular endothelial growth factor receptor-2. Note: Development phase is based on databases such as R & D Focus, R & D Insight, Pharmaprojects, and the Investigational Drugs Database (IDdb3); periodicals such as Scrip, the Pink Sheet, and Marketletter; company reports and press releases; and industry contacts.

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stromal tumors. Although normal ovarian surface epithelium expresses c-kit and its ligand, which is stem-cell factor, they are overexpressed in more than 70% of CaO cases. In addition, approximately 73% of CaOs express PDGFR and at least 50% of these cases co-express PDGFR-alpha (Henriksen R, 1993). Several groups have been evaluating imatinib for CaO, including the Gynecologic Oncology Group, M.D. Anderson Cancer Center, and the National Cancer Institute (NCI). The results of these trials should be available soon. The development of novel cytotoxics is likely to focus on increased selectivity and/or novel mechanisms of action rather than on simple assessment of analogues of existing agents. Four examples of such agents that have exhibited signiÞcant activity in the most difÞcult scenario—platinum-refractory/resistant CaO—are patupilone (Novartis’s EPO-906), trabectedin (PharmaMar/Johnson & Johnson’s Yondelis), TLK-286 (Telik’s Telcyta), and capecitabine (Roche’s Xeloda). Trabectedin and TLK-286 are described in the section “Novel Alkylating Agents.” Patupilone is an epothilone, a novel natural (bacterial) product whose antitumor action is similar to that of taxanes (microtubule stabilization) but is much more potent. At least four epothilone analogues have reached clinical testing, including patupilone and BMS’s ixabepilone. Because of these compounds’ early development stage for the treatment of CaO, they are not discussed in detail here. The oral ßuorouracil pro-drug capecitabine reaches the tumor cells by a threestep enzyme-activation process following oral administration. The third of these steps relies on activation by thymidine phosphorylase; this enzyme is expressed in certain tumors, including CaO. Trials of intravenous (IV) capecitabine several years ago indicated that the drug does possess activity for CaO, but its use has declined as other drugs have been developed. More recent trials have focused on its use in combination with gemcitabine and either epirubicin or carboplatin. In view of the lack of current clinical activity, this product is not discussed in detail here. Other cytotoxic classes in early-stage development for the treatment of CaO include topoisomerase I inhibitors and microtubule inhibitors. They are not discussed here given the lack of data for them. In the future, clinical trials will assess new agents in combination treatment with liposomal doxorubicin (Ortho Biotech’s Doxil, Schering-Plough/Essex Pharma’s Caelyx) to treat patients whose disease is platinum-refractory. Likewise, a number of candidates may begin to emerge for combination with carboplatin in the treatment of platinum-sensitive CaO. A number of alternative formulations of existing agents—particularly topoisomerase I inhibitors and taxanes—have been actively pursued. Researchers theorize that changes in tissue distribution and pharmacokinetics can result in favorable shifts in efÞcacy and toxicity proÞles, as with liposomal doxorubicin (see “Current Therapies”). Examples include liposomal lurtotecan (OSI Pharmaceuticals’ OSI211), polyglutamated conjugates of camptothecin (Cell Therapeutics’ CT-2106), and paclitaxel (Cell Therapeutics’ CT-2103). CT-2103 has entered Phase III trials (see the section “CT-2103” further on), and CT-2106 entered a Phase I/II singleagent trial in CaO at the end of 2004. OSI-211 has been the subject of two recent

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trials: a Phase I study in combination with liposomal doxorubicin in refractory CaO and a randomized Phase II parallel evaluation of OSI-211 and topotecan (GlaxoSmithKline/Merck’s Hycamtin) in relapsed CaO. By May 2005, OSI Pharmaceuticals had cut back its oncology programs and development of OSI-211 had been discontinued. In clinical trials involving emerging therapies, physicians use X ray, computerized tomography (CT) scan, and magnetic resonance imaging (MRI)—known as Response Evaluation Criteria in Solid Tumors (RECIST)—to evaluate the ovarian tumor response. The technique is recommended but not mandatory for NCI-sponsored trials and involves formalized rules for measuring tumor target lesions. The criteria are based on a simpliÞcation of former methods and on measurable disease (i.e., the presence of at least one measurable lesion). The criteria encompass four response categories: • • • •

Complete response (CR) = disappearance of all target lesions. Partial response (PR) = 30% reduction in the sum of the longest diameter of target lesions. Progressive disease (PD) = 20% increase in the sum of the longest diameter of target lesions. Stable disease (SD) = small changes that do not meet above criteria.

Novel Taxanes Overview. Paclitaxel has been a highly successful agent for the treatment of a variety of solid tumors. As a result, the development of new formulations and analogues to address problems with paclitaxel’s administration has generated much interest. The goal is to develop more-soluble, cremophor (a solvent)-free formulations and analogues that enable the administration of higher doses of paclitaxel for greater clinical efÞcacy while reducing the associated toxicity. These agents may also have an improved pharmacokinetic proÞle and different mechanism of intracellular uptake. These agents’ commercial potential was recently demonstrated with the launch of American Pharmaceutical Partners’ nanoparticle formulation ABI-007 (Abraxane) for the treatment of metastatic breast cancer. This agent is being evaluated in CaO patients. As yet, no data on this indication have been published, but off-label use in the CaO setting is likely. Mechanism of Action. Taxanes suppress microtubule dynamics and disassembly, thereby stopping mitosis at the metaphase/anaphase transition and inducing cell death. Formulation. Companies are investigating a variety of approaches to modifying taxanes. They include oral formulations of paclitaxel (Ivax’s Paxoral), liposomal formulations (PÞzer/NeoPharm’s LEP), pegylated formulations (Enzon’s

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Macromed), and, until recently, microsphere formulations (Guilford Pharmaceuticals’ Paclimer—however development on this particular product was just recently ceased). This discussion focuses on the most promising agents, a paclitaxel polyglutamated formulation (Cell Therapeutics’ CT-2103) and a paclitaxel analogue (BMS’ BMS-184476). CT-2103. Cell Therapeutics is developing CT-2103 (paclitaxel poliglumex, PGTXL, Xyotax), a biodegradable poly-L-glutamate/paclitaxel conjugate, for the treatment of various tumors. It is in Phase III studies for both non-small-cell lung cancer (NSCLC) and CaO. The agent is licensed from the M.D. Anderson Cancer Center. Chugai has development and marketing rights in select Asian markets, including Japan. Conjugation of paclitaxel to poly-L-glutamate enhances aqueous solubility and eliminates the need for the toxic solubilizing agent cremophor, resulting in a tenminute infusion. This approach offers the prospect of less formulation-related toxicity and the delivery of higher and potentially more effective cytotoxic doses to tumors. In May 2005, clinical data on CT-2103 in combination with carboplatin were presented at the 41st American Society of Clinical Oncology (ASCO) meeting in Orlando, Florida (Herzog T, 2005). In the Phase II study, 82 patients with stage III/IV ovarian or primary peritoneal carcinoma who had standard debulking surgery received induction therapy of a ten-minute IV infusion of CT-2103 (175 mg/m2 [62 patients] or 210 mg/m2 [20 patients]) plus carboplatin AUC (area under curve [carboplatin dosage (mg) = AUC(mg/mL × min) × carboplatin clearance (mL/min)]) 6. Patients with stable disease after four to six cycles received single-agent maintenance therapy (175 mg/m2 CT-2103). Based on CA-125 levels, 54% of patients experienced a CR and 10% a PR. Grade 3 (severe) neuropathy occurred in 7 of 23 patients at 210 mg/m2 and resulted in the withdrawal of six patients. As a result, the dose of CT-2103 was reduced to 175 mg/m2 for subsequent patients. The international Phase III trial of CT-2103, initiated in July 2002, will involve 950 chemotherapy-naive CaO patients. It will evaluate the combination of a Þxed dose of carboplatin (AUC 6) and either CT-2103 at 210 mg/m2 or paclitaxel at 175 mg/m2 , given for six cycles. Patients who achieve complete remission will go on to a consolidation arm consisting of single-agent CT-2103 or paclitaxel, both at 175 mg/m2 once monthly for 12 months. The primary endpoint is progressionfree survival (PFS). As well as being investigated in combination with existing treatments, CT2103 is being evaluated as monotherapy for the treatment of CaO. Study CTI1071 was an open-label multi-center study to determine the response rate and time to disease progression (TTP) in heavily pretreated patients with recurrent CaO or fallopian tube or peritoneal cancer (Sabbatini P, 2004). CT-2103 was given to 99 patients with measurable disease at a dose of 175 mg/m2 over ten minutes every three weeks without routine premedications. Platinum-sensitive (n = 42) and platinum-refractory or platinum-resistant patients (n = 57) were

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enrolled. Thirty-nine patients (39%) had received one or two prior regimens, and 60 patients (61%) had received 3–12 regimens. The median number of cycles was three (range, 1–14 cycles). The response rate for all patients was 10%; the median TTP was two months. The PR in platinum-sensitive and platinumresistant patients was 14% and 7%, respectively. In patients with only one or two prior regimens, the PR in platinum-sensitive and platinum-resistant patients was 28% and 10%, with a median TTP of four and two months, respectively. Grade 2 (moderate) or 3 neuropathy was reported in 30% of patients. Grade 2 hypersensitivity occurred in 8% of patients who were subsequently treated with premedications; one patient had grade 3 hypersensitivity and was removed. Grade 2 alopecia was absent. The Gynecologic Oncology Group has initiated a Phase III trial of CT2103. The multi-center, U.S. and European study will enroll 1,550 patients who achieved a CR following standard Þrst-line chemotherapy. Subjects will receive a ten-minute infusion of CT-2103 once monthly for 12 months, with no maintenance therapy or monthly paclitaxel. The endpoints will include progression-free and overall survival. Researchers hope that the trial will demonstrate that monthly maintenance of CT-2103 therapy improves both overall and progression-free survival of patients with advanced CaO, without signiÞcant side effects that diminish patients’ quality of life. BMS-184476. BMS-184476 is an analogue of paclitaxel under development for the treatment of solid tumors, including breast and NSCLC tumors. The agent is the most-advanced compound in a series of taxanes under development by BMS. It is in Phase II studies in the United States and Europe for CaO. At ASCO 2002, researchers presented the results of a U.K. Phase II study involving 30 CaO patients who had previously received platinum- and paclitaxelbased chemotherapy. The patients had progressed six months or more after receiving paclitaxel. BMS-184476 was administered at a dose of 50 mg/m2 by infusion for one hour at days 1 and 8 of a 28-day cycle. Of the 15 evaluable patients, 6 (40%) experienced a PR and 7 (47%) experienced disease stabilization. The authors considered the agent to be well tolerated; major toxicities encountered in 17 evaluable patients included grades 3 and 4 (life-threatening) neutropenia (62%), grade 1 (mild) sensory neuropathy (47%), and alopecia (35%) (Poole CJ, 2002). Novel Platinum Compounds Overview. Platinum compounds have been used for more than 20 years as anticancer therapeutics; their efÞcacy in treating CaO is well documented—making this market difÞcult to penetrate. However, the use of these compounds is sometimes limited by acquired drug resistance, such as that seen with cisplatin and carboplatin. Therefore, the goal of new platinum agents is to retain the class effect of platinum compounds while combating this drug resistance. Mechanism of Action. Platinum-containing compounds (e.g., alkylating agents) react with many biologic compounds; however, their cytotoxic effect

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is believed to be mediated by their ability to create intra- and interstrand DNA cross-links. In this way, they interfere with DNA replication and cell division. Oxaliplatin. Oxaliplatin (SanoÞ-Aventis’s Eloxatin) (Figure 10), a thirdgeneration platinum cytotoxic agent, is marketed in Europe and the United States for the treatment of colorectal cancer. The agent is in Phase III trials in France for CaO, in a Phase II study in the United States in combination with topotecan, and in a Phase I/II study in France in combination with docetaxel (SanoÞAventis’ Taxotere). SanoÞ-Aventis obtained worldwide rights to oxaliplatin (with the exception of Japan and several smaller markets) from Debiopharm in 1994. In 2000, SanoÞ-Aventis bought out Eli Lilly’s stake in a U.S. joint venture, SanoÞ Lilly Oncology, which had been formed to market oxaliplatin. Oxaliplatin is the Þrst available diaminocyclohexane platinum, and its mechanism of action is thought to be similar to that of cisplatin. Oxaliplatin exerts its antitumor effects by cross-linking two adjacent guanine DNA bases, two adjacent guanine and adenosine DNA bases, or two guanine DNA bases on complementary DNA strands. This cross-linking prevents DNA replication and so induces tumor cell apoptosis. Studies have shown that oxaliplatin is active in cell lines that are resistant to carboplatin and cisplatin (Raymond E, 1998). DeÞciencies in mismatch repair (MMR) (DNA MMR defects occur in a wide variety of sporadic human cancers) and increases in replicative bypass are thought to contribute to cisplatin resistance. Diaminocyclohexane platinum adducts do not appear to be recognized by the MMR complex, and, therefore, have not been shown to induce similar resistance to oxaliplatin. A Phase II study conducted to determine the efÞcacy of oxaliplatin monotherapy (130 mg/m2 IV over two hours every 21 days) in 25 patients with platinumresistant or platinum-refractory CaO demonstrated minimal activity (Fracasso PM, 2003). Therefore, studies investigating this agent have concentrated on oxaliplatin in combination with other active agents and, for the most part, in women with platinum-n¨aive or platinum-sensitive CaO. Researchers evaluated oxaliplatin as a Þrst-line therapy in combination with cyclophosphamide in patients with previously untreated, advanced-stage CaO (Misset JL, 2001). In the Phase II/III comparative study, 177 patients received a maximum of six cycles with IV regimens of oxaliplatin/cyclophosphamide or cisplatin/cyclophosphamide (CPC). Eighty-six patients were given 130 mg/m2 oxaliplatin and 1,000 mg/m2 cyclophosphamide, both infused for two hours, and 91 patients received 100 mg/m2 cisplatin for one hour and 1,000 mg/m2

NH2

O

O

O

O

Pt NH2

FIGURE 10. Structure of oxaliplatin.

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cyclophosphamide infused for two hours. The oxaliplatin combination displayed efÞcacy similar to that of the CPC regimen, with no signiÞcant difference in objective response rates (64% of the oxaliplatin patients versus 67% of the CPC patients); median PFS (13.0 months versus 13.3 months); or overall survival (36.0 months versus 25.1 months). However, toxicities were signiÞcantly less severe in the oxaliplatin arm: researchers observed grades 3 and 4 leukopenia in 37% of the oxaliplatin patients versus 56% of the CPC patients; anemia in 7% of the oxaliplatin patients versus 32% of the CPC patients; and blood transfusions had to be given to 8% of the oxaliplatin patients versus 21% of the CPC patients. At ASCO 2002, investigators presented preliminary Þndings of a Phase II study evaluating an oxaliplatin/paclitaxel combination in 45 patients pretreated with one platinum- or platinum-taxane-based regimen (Viens P, 2002). The platinum-free interval was longer than 12 months in 31 patients and was 6–12 months in 14 patients. Thirty-two patients had received prior treatment with a taxane. Patients received paclitaxel 175 mg/m2 , followed by oxaliplatin 130 mg/m2 every 21 days, for six cycles. The overall response rate was 87%, with CRs seen in 9 patients and PRs in 30. Median projected PFS was 12 months. The most common adverse effects observed were grade 2 reversible neurotoxicity in 53% of patients, grade 3 and 4 neutropenia (40%), and grade 2 alopecia (33%). A multi-center feasibility study determined the toxicity proÞle and antitumor activity of the gemcitabine plus oxaliplatin combination as a second-line treatment in paclitaxel-resistant or paclitaxel-refractory advanced CaO (Raspagliesi F, 2004). Twenty patients received a 30- to 60-minute infusion of gemcitabine once a week for two weeks, followed by a 120- to 180-minute infusion of oxaliplatin every three weeks. The doses used were 1,000 and 130 mg/m2 , respectively. Seventeen cases were platinum-resistant and three were platinum-refractory. Grade 3 and 4 thrombocytopenia occurred in 14 cases. Two patients required platelet transfusion, and eight patients received hydrocortisone. The dose-limiting toxicity was thrombocytopenia. Combined grade 3 and 4 neutropenia was observed in eight cases. Five patients were treated with recombinant erythropoietin because of grade 3 anemia and four patients received granulocyte colony-stimulating factor (G-CSF) prophylactically from the Þrst cycle. The overall response rate of the combination in terms of antitumor activity was 26%. The researchers concluded that a combination of gemcitabine and oxaliplatin using this schedule gave rise to a moderate/severe toxicity proÞle and would be feasible only if growth factors were used and/or gemcitabine were administered at lower doses. Furthermore, the antitumor activity of the combination was seen as insufÞcient reward for the resultant toxicity proÞle; however, it was equivalent to that of other drugs used to treat platinum-refractory and platinum-resistant patients. The preliminary results of a Phase II trial of oxaliplatin and topotecan as a Þrst-line treatment for 17 patients with advanced-stage CaO (stage III–IV) were presented at ASCO 2005 (Gorbounova V, 2005). The primary aim of the study is to evaluate the response rate and toxicity proÞle and the secondary aim will be time to disease progression and overall survival. Twelve patients

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received oxaliplatin 85 mg/m2 and Þve patients received 100 mg/m2 on day 1 (over 120 minutes) plus topotecan 1.5 mg/m2 on days 2–4 (over 30 minutes) every 21 days. Five patients had CRs, seven PRs, four SD, and one PD. The toxicity proÞle was acceptable. Novel Alkylating Agents Overview. A nitrogen mustard alkylating agent was the Þrst nonhormonal chemical to demonstrate signiÞcant antitumor activity. Subsequently, less toxic and more clinically effective alkylating agents have been developed—for example, cyclophosphamide, ifosfamide, melphalan, mitomycin C, and nitrosourea derivatives. Despite the large number of agents available, an improvement in the toxicity of alkylating agents is still required. Two novel alkylating agents are in Phase III studies for CaO. Mechanism of Action. Alkylating agents exert their cytotoxic action by alkylation of DNA bases. Alkylating agents may be mono- or bifunctional (the latter induces DNA cross-links). Unless repaired, the alkylated lesions will prevent the cell from replicating effectively. One mechanism of resistance to alkylating agents is DNA repair enzymes such as O6-alkylguanine-alkyltransferase, which removes the alkyl group from the O6 position in guanine. TLK-286. TLK-286 (Telcyta) is the lead compound in a series of alkylating agents in development by Telik (formerly Terrapin) as a potential treatment for cancer. Telik has retained full worldwide rights to TLK-286. The agent is under investigation across a range of tumors; Phase III trials are being conducted in NSCLC and CaO and Phase II trials in breast, colon, and lung cancer. Fast-track designation for third-line therapy for platinum-refractory or platinum-resistant CaO was granted in September 2003. In December 2003, the FDA granted TLK-286 fast-track designation for third-line therapy of locally advanced or metastatic NSCLC. TLK-286 is an apoptosis-inducing small-molecule drug that targets glutathione S-transferase (GST) P1–1. This protein plays a role in the development of resistance to common chemotherapeutic drugs, and it is elevated in a variety of tumors, possibly worsening the prognosis in CaO. The drug may also be useful for treating drug-resistant disease. Researchers at the ASCO 2002 meeting presented data on a Phase II trial involving 36 patients (31 evaluable) with advanced CaO, of whom 64% were deemed platinum-refractory and 36% platinum-resistant (Kavanagh JJ, 2002). Patients received TLK-286 1,000 mg/m2 given intravenously once every three weeks. Disease control was achieved in 52% of patients, including one with a CR, three with PRs, and 12 with SD. The most common toxicities were mild (grade 2) fatigue, nausea, vomiting, microscopic hematuria, and anemia. A few higher-grade toxicities were also reported. In November 2002, at the European Organization for Research and Treatment of Cancer-National Cancer InstituteAmerican Association for Cancer Research (EORTC-NCI-AACR) symposium,

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Telik reported further results: 63% of the patients were alive, and the median survival, which exceeded 16 months, had not yet been reached. Another Phase II trial presented at the ASCO 2003 meeting examined TLK286 administered weekly in patients with platinum-refractory or -resistant thirdline advanced-stage CaO (Kavanagh JJ, 2003). Twenty-three patients with a median of three prior chemotherapy regimens received TLK-286 at 960 mg/m2 IV weekly until tumor progression or unacceptable toxicities. The most common possibly drug-related toxicities (grade 2) were fatigue, nausea, and thrombocytopenia. No grade 4 toxicities occurred. Grade 3 toxicities were infrequent and no cumulative toxicities were seen. No dose reductions were required. At interim analysis, 11 of 23 patients were evaluable and objective tumor responses were seen in 2 of the 11 patients. The longest duration of therapy is in the PR patients, who were free of tumor progression more than Þve months later. In December 2002, two Phase II trials were initiated for TLK-286 in CaO in the United States. One was in combination with liposomal doxorubicin (TLK286 960 mg/m2 plus liposomal doxorubicin 50 mg/m2 ) to treat patients who had failed platinum-based chemotherapy; the second evaluated TLK-286 in combination with carboplatin (TLK-286 500 mg/m2 plus carboplatin AUC 5 or 6) in the treatment of recurrent, platinum-sensitive CaO. In October 2004, data from both trials were reported at the Tenth Biennial International Gynecologic Cancer Society. An October 4, 2004, press release for Telik announced that for the trial of TLK-286 in combination with liposomal doxorubicin, 19 of 51 patients enrolled were evaluable for efÞcacy at the time of analysis. The objective response rate was 42%, with eight PRs. The overall disease control rate was 74% and median PFS was 34.1 weeks. The combination was well tolerated. In the study of TLK-286 in combination with carboplatin, 26 patients were evaluable out of 53 enrolled (Telik, press release, October 4, 2004). The objective response rate was 54%, including four CRs and 10 PRs. Overall disease control rate was 73%, and median PFS was 25.6 weeks. The combination was well tolerated. A Phase III study began in March 2003. Assessment of Survival in Solid Tumors-1 (ASSIST-1) involves 440 patients whose disease has progressed following platinum-based chemotherapy and one second-line treatment. Patients are randomized to receive treatment with TLK-286, liposomal doxorubicin, or topotecan. In December 2004, Telik initiated ASSIST-3, a Phase III trial to evaluate a combination of TLK-286 plus carboplatin versus liposomal doxorubicin in the second-line treatment of platinum-refractory or platinum-resistant CaO. The study will enroll 244 women; end points will be objective response rate, PFS, and overall survival. The preliminary response rates for TLK-286 do not seem to represent a signiÞcant improvement in efÞcacy over currently available agents. Combination with another agent is a likely way forward given the mild toxicities associated with the compound. It may be the case that as a result of the currently running ASSIST-1 and ASSIST-3 trials, TLK-286 will be adopted in a second- or

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third-line role in combination with liposomal doxorubicin, topotecan, or carboplatin. Trabectedin. Trabectedin (ecteinascidin-743, ET-743, Yondelis), a marine compound isolated from the Caribbean tunicate Ecteinascidia turbinata, is being developed by the Spanish company PharmaMar and Ortho Biotech (a subsidiary of Johnson & Johnson) as a potential treatment for a number of tumors, including sarcoma, breast cancer, and NSCLC. The agent is in Phase III studies for advanced-stage CaO in the United States and Europe. In October 2003, trabectedin was awarded European Union orphan drug status for CaO. In April 2005, the FDA granted it orphan drug status for the same indication. Trabectedin has a unique mechanism of action that makes it a novel antitumor agent. It binds to the minor groove of DNA, resulting in alkylation of the guanine N2. Defects in DNA repair pathways have paradoxical effects on the antitumor activity of trabectedin: loss of mismatch repair does not affect trabectedin’s toxicity, while loss of DNA-dependent protein kinase activity enhances its toxicity. However, defects in transcription-coupled nucleotide excision repair confer patient resistance to trabectedin. Given that a DNA repair capability appears to be necessary for at least one mechanism of trabectedin-mediated cytotoxicity, the drug may interact with the DNA repair machinery to induce lethal strand breaks. One of the most novel aspects of trabectedin is its effect on RNA polymerase II-mediated gene transcription. Trabectedin selectively inhibits activation of the multidrug-resistant gene while leaving constitutive gene expression relatively unaffected (Scotto KW, 2002). The Þnal results of a Phase II study using a weekly schedule of trabectedin (0.58 mg/m2 IV) for three weeks followed by a week of rest in second- or thirdline CaO were presented at the ASCO 2005 meeting (McMeekin DS, 2005). In the platinum-sensitive group, 55 patients were treated; the response rate was 28.8% and the median PFS was 5.1 months. In the platinum-resistant group, 64 patients were treated; the response rate was 4.8% and the median PFS was two months. Grade 3 and 4 toxicities in pooled cycles were reversible liver ALT (alanine aminotransferase) elevation (12%), neutropenia (8%), fatigue (5%), and nausea/vomiting (4%). Researchers conclude that trabectedin is active and well tolerated when administered as a three-hour infusion for three weeks out of a four-week cycle in platinum-sensitive patients. In April 2005, PharmaMar initiated a Phase III pivotal, international, multicenter study to compare liposomal doxorubicin monotherapy with the combination of trabectedin and liposomal doxorubicin in CaO patients who have relapsed after receiving standard Þrst-line chemotherapy. This randomized study will enroll 650 patients in 110 hospitals in 16 countries. Immunotherapies Overview. Several potential immunotherapies are under investigation for the treatment of CaO in a variety of settings, including both Þrst-line and consolidation therapy. These immunotherapies include humanized bispeciÞc antibodies,

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radiolabeled murine antibodies, interferon gamma (an immunomodulator that also exerts an antiproliferative and antiangiogenic effect), and vaccines. Their uptake and positioning in the therapeutic armamentarium will likely be governed by developments in the basic understanding of CaO’s pathophysiology and advances in molecular biology. These developments will facilitate characterization of tumor markers speciÞc to CaO and should, in turn, lead to a stratiÞcation of the market by patient type in the same way that breast cancer patients are stratiÞed by HER2/neu overexpression. Mechanism of Action. All cancer immunotherapeutic approaches use elements of the patient’s immune system to kill cancer cells, but their means of doing so varies. IDM-1. Immuno-Designed Molecules (IDM) is developing IDM-1 (Osidem), a combination of Medarex’s MDX-210 with IDM’s macrophage-activated killer (MAK) cells, for the treatment of CaO. This “cell drug” therapy is in Phase III trials for stage III CaO in Europe, Canada, and Australia. In May 2002, IDM received FDA clearance to begin Phase III trials in the United States. In January 2002, IDM signed a codevelopment agreement with SanoÞ-Synth´elabo (now part of SanoÞ-Aventis) giving the latter the right of Þrst refusal for up to 20 of IDM’s cellular immunotherapies and therapeutic vaccines for cancer. MDX-210 is a bispeciÞc antibody consisting of a humanized anti-HER2/neu fragment and a humanized anti-CD64 fragment (the CD64 receptor of Fc gamma R1 is present on cytotoxic cells involved in attacking tumor cells). The antibody is immunologically active, recruits circulating monocytes, and stimulates certain cytokines. IDM-1 couples tumor-cell-killing cells derived from the patient’s own MAK cells with the antibodies in MDX-210 that link the killer cells to speciÞc antigens on the surfaces of the targeted cancer cells—in this case, HER2 -expressing CaO cells. The goal of the cellular immunotherapy treatment is to eliminate residual tumor cells following surgery and chemotherapy. At ASCO 2000, researchers presented the results of a Phase II trial of IDM-1 (de Gramont A, 2000). The study involved 17 late-stage CaO patients with minimal residual disease following surgery and chemotherapy. Patients were treated with six doses of IDM-1, injected intraperitoneally (IP) over a one-month period. EfÞcacy was evaluated at the time of third-look surgery performed three months after the initiation of cell therapy. Of the 14 evaluable patients, 8 demonstrated HER2/neu overexpression at the surface of ovarian tumor cells. Disease progressed in Þve patients and stabilized in three; six patients experienced a complete clinical response. No serious (grade 3 and 4) side effects were reported during the treatment. Although the CR rate observed in the aforementioned study is encouraging, only a small proportion of CaO patients overexpress HER2/neu. A further barrier to widespread use is the IP administration. This form of administration is a complex procedure that requires specially trained physicians available at a comparatively small number of clinics. Despite trials showing improved survival

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using IP delivery of current treatments, few centers have adopted this mode of therapy. Interferon Gamma-1b. Interferon gamma-1b (InterMune Pharmaceuticals’ Actimmune; Boehringer Ingelheim’s Imukin) is in Phase III trials for the treatment of CaO. The agent has been launched extensively for chronic granulomatous disease, and it is licensed in the United States for the treatment of severe malignant osteopetrosis. It is also under investigation for idiopathic pulmonary Þbrosis (IPF), cryptococcal meningitis, cystic Þbrosis, atypical mycobacterial infections, asthma, hepatitis C virus infection, liver disease, and non-Hodgkin’s lymphoma. In 1999, InterMune acquired the development and commercialization rights for all oncology indications from the developer, Genentech. In 2001, InterMune established an international strategic partnership with Boehringer Ingelheim. Under this partnership, InterMune has the responsibility for the clinical and regulatory development of interferon gamma-1b, and Boehringer Ingelheim retains an option to exclusively promote interferon gamma-1b under Boehringer Ingelheim’s trade name in all countries outside of North America and Japan. InterMune has also formed an alliance with Maxygen to develop a next-generation version of interferon gamma-1b that can be taken once weekly rather than three times a week as required by the current regimen. Interferon gamma-1b is a naturally occurring human protein that exerts both an antiproliferative and an antiangiogenic effect. It stimulates the immune system by activating macrophages, an effect that triggers antitumor activity. The induction of macrophages and other cytotoxic effector T cells may also have a nonspeciÞc immunomodulatory inßuence that enhances the patient’s responsiveness to chemotherapy. Researchers in Europe conducted a randomized Phase III trial involving 148 women in stages Ic-Illc CaO who had undergone primary surgery (Windbichler GH, 2000). The control group received cisplatin 100 mg/m2 and cyclophosphamide 600 mg/m2 ; the experimental group also received interferon gamma-1b 0.1 mg subcutaneously on days 1, 3, 5, 15, 17, and 19 of each 28-day cycle. Results showed that interferon gamma-1b was well tolerated, although the proportion of patients developing fever and ßu-like symptoms was higher than in the control group. Researchers did not observe any appreciable increase in gastrointestinal, neurological, or hematological side effects. The CR rate was higher (68%) in the study group than in the control group (56%), as were the median TTP (48 months versus 17 months) and the three-year overall survival rate (74% versus 58%). After reviewing the results of the European trial, InterMune decided to pursue a U.S. trial. This multicenter randomized Phase III Gamma Interferon and Chemotherapy EfÞcacy Study (GRACES) will evaluate the efÞcacy and safety of interferon gamma-1b in combination with standard chemotherapy (carboplatin plus paclitaxel) in 800 patients recently diagnosed with advanced-stage CaO. The study’s endpoints are PFS and overall survival. Enrollment for this trial was completed in April 2004.

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Although the Þndings of the European trial are important, chemotherapy used for CaO has since been supplanted by more effective and less toxic combinations of carboplatin and paclitaxel. Thus, the success of interferon gamma-1b as a Þrst-line therapy will depend on the results of the GRACES trial. In addition, interferon is associated with side effects (lethargy, fatigue, and inßuenza-like symptoms) that can greatly affect quality of life during the course of treatment. Current Phase III trials, then, will need to show that interferon gamma-1b’s impact on overall and progression-free survival is signiÞcant enough to make enduring these side effects worthwhile. Oregovomab. ViRexx’s oregovomab is one of several therapeutic vaccines in development to boost the immune response to CaO, but the only one in late-stage development. Vaccines in early-stage development include Cancer Vac’s immunotherapeutic vaccine, which involves priming of dendritic cells with a mannan-MUC1 conjugate; CellControl Biomedical’s ACA-125, an antiidiotype antibody directed against CA-125; and Endocyte’s FolateImmune, a novel immunotherapy composed of a vaccine EC90 with GP-0100 adjuvant and a folate-conjugated vaccine (EC-17) that targets folate-receptor-positive tumor cells. These approaches are all in Phase I/II studies. ViRexx (formerly AltaRex) is developing oregovomab (B43-13, OvaRex), a murine monoclonal antibody vaccine, for the treatment of CaO as monotherapy and in combination with chemotherapy. The company acquired the agent in 1995 under an exclusive license from Biomira and, in 2002, entered into an exclusive license agreement with Unither Pharmaceuticals (United Therapeutics) covering North America and Japan. Unither is responsible for conducting clinical programs in the United States. Oregovomab is also licensed to Dompe Farmaceutici for several European Union countries, including Italy and Spain. In the United States, oregovomab received orphan drug status in 1996 and fasttrack designation in 1998; in the European Union, it received orphan drug status in 2002. The agent is in two pivotal Phase III clinical studies with Unither in the United States for the treatment of stage III or IV CaO as postchemotherapeutic consolidation therapy. ViRexx has submitted a Phase II protocol to the European Organization for Research and Treatment of Cancer to examine the potential of oregovomab in combination with chemotherapy in patients with platinumsensitive relapsed disease. Although it is a murine monoclonal antibody to tumor-speciÞc antigen CA-125, oregovomab also acts like a cancer vaccine, binding circulating CA-125 antigens and forming complexes that the body recognizes as foreign. This behavior induces a host immune reaction that results in the production of antigen-antibody complexes, potentiating the immune reaction of the host to the attempt by CA-125 to produce CaO tumors. Oregovomab was originally developed as a tumor-imaging agent for CaO, but researchers noticed that patients who were exposed to it unexpectedly lived longer (Schultes BC, 1998). In September 2004, data were published from a trial of oregovomab in 145 women with advanced-stage CaO (Berek JS, 2004). The purpose of the trial was

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to assess oregovomab as consolidation treatment and reÞne the immunotherapeutic strategy for subsequent study. Patients who had a CR to primary treatment were randomly assigned to oregovomab (73 patients) or placebo (72 patients) administered at weeks 0, 4, and 8, and every 12 weeks up to two years or until recurrence. Patients assigned to active treatment received 2 mg doses of oregovomab diluted in 50 mL of sodium chloride injection administered via IV over 20 minutes. The primary endpoint was time to relapse (TTR). For the population overall, median TTR was not different between treatments—13.3 months for oregovomab and 10.3 months for placebo. However, a subset of 67 patients was identiÞed who responded more favorably to surgery and chemotherapy. These patients had remaining tumors of 2 cm or less following surgery and no conclusive evidence of disease during and after chemotherapy. Within this “successful front-line treatment” (SFLT) group, patients who received oregovomab experienced double the remission time when compared with those patients who did not receive the investigational treatment; TTR was 24.0 months in the oregovomab group compared with 10.8 months for placebo. A set of conÞrmatory Phase III studies has been initiated to determine whether the SFLT population derives beneÞt from oregovomab treatment. In January 2003, two identical pivotal 177-patient randomized Phase III trials of oregovomab were initiated in the United States. Called Immunotherapy Pivotal Ovarian Cancer Trials (IMPACT) I and II, these trials involve stage III or IV CaO patients who have had successful initial surgery, are responsive to chemotherapy, and are in remission (based on a clinical and CT exam) at the end of six months of chemotherapy treatment. The primary endpoint for the trials is TTR between active and placebo groups; secondary objectives include quality of life, immune response, and safety. Patients will also be followed for survival. Another ongoing U.S. trial began enrollment in February 2005. This multicenter Phase III study will assess survival, quality of life, immune response, and safety in 350 women with stage III or IV CaO. Participants will receive best standard supportive care plus oregovomab or placebo. Two-thirds of enrolled patients will be randomized to receive oregovomab. The study medication will be administered by 20-minute infusions once a month for three consecutive months, followed by quarterly doses for up to Þve years. All participants will be followed for relapse and survival status for up to seven years. Angiogenesis Inhibitors Overview. Angiogenesis, the formation of new blood vessels, plays a major role in many normal physiological processes. It also plays a central role in a number of pathological conditions, including solid tumor growth and metastasis, and therefore represents an area of considerable commercial interest. Approximately 15 angiogenesis inhibitors are in clinical development for the treatment of CaO. Because of this high number, only those agents in late-stage development are discussed here.

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Mechanism of Action. Most anticancer angiogenesis inhibitors under development inhibit a single antiangiogenic pathway; the most promising pathway appears to be inhibition of vascular endothelial growth factor (VEGF). VEGF, a multifunctional cytokine and potent permeability factor secreted in response to hypoxia, has a major angiogenesis-promoting effect, is overexpressed in CaO, and is associated with ascites formation, malignant progression, and poor prognosis. However, inhibition of a single stimulatory signal is likely to have an effect only in a subset of patients and cannot be universally applied to all cancers, especially end-stage cancers, when multiple proangiogenic factors are expressed. Inhibition of VEGF receptors is another approach under investigation. The VEGF receptor is expressed on normal endothelial cells, and these cells therefore have a much lower mutation rate than that of tumor cells and are less likely to become resistant to a VEGF-receptor-targeted drug. Bevacizumab. Genentech’s bevacizumab (Avastin) is a recombinant humanized VEGF monoclonal antibody. In July 2003, Roche acquired worldwide rights to bevacizumab except in the United States, where Genentech retains exclusive rights. In February 2004, Genentech launched bevacizumab in the United States for use in combination with IV 5-ßuorouracil (5-FU)-based chemotherapy for treating Þrst-line metastatic cancer of the colon or rectum. In January 2005, the drug gained European Union approval for treating patients with previously untreated metastatic colorectal cancer. Bevacizumab is being developed as a potential treatment for other solid tumors. Phase III trials include NSCLC, metastatic breast cancer, renal cell carcinoma, and pancreatic cancer, and Phase II trials include prostate cancer and melanoma. A Phase III trial, GOG-0218, is under development for CaO and will investigate concurrent treatment with bevacizumab and chemotherapy as well as the effect of 15 months’ treatment with bevacizumab postchemotherapy. Also recruiting is an open-label, single-arm, two-stage, multi-center Phase II study evaluating the efÞcacy and safety of bevacizumab in women with platinum-resistant, advancedstage CaO who subsequently progressed either during treatment with liposomal doxorubicin or topotecan therapy or within three months of discontinuing treatment with liposomal doxorubicin or topotecan. Bevacizumab binds to all human forms of VEGF, thus preventing VEGF from binding to VEGF receptors on the endothelial cells. This action theoretically prevents one step in the process of angiogenesis. In a Phase II Gynecologic Oncology Group (GOG) study presented at ASCO 2005, 64 patients with persistent or recurrent ovarian or primary peritoneal cancer after receiving one to two prior cytotoxic regimens were treated with bevacizumab 15 mg/kg every 21 days until disease progression or prohibitive toxicity (Burger RA, 2005). Primary endpoints were PFS at six months, clinical response rate, and toxicity. The treatment was well tolerated; no grade 3 or 4 hematologic toxicities occurred. Four patients had grade 4 gastrointestinal adverse events, and four had grade 3 hypertension. Two patients experienced grade 3 allergic reactions. A clinical response rate of 17.7% was achieved with a six-month

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PFS of 38.7%. Median PFS was 4.8 months, and median overall survival was 17 months. Reduction of ascites was an early marker of response, and even patients who had only tumor stabilization typically had complete resolution. Another abstract presented at ASCO 2005 described early results from a Phase II trial of combination therapy with bevacizumab and low-dose metronomic cyclophosphamide in recurrent ovarian and primary peritoneal cancer in 29 patients (Garcia A, 2005). Based on an encouraging partial response rate of 28% with six-month PFS of 57% and median TTP of 7.5 months, the study has begun its second stage of patient accrual. In September 2005, Genentech announced that enrollment in a multi-center, single-arm Phase II study of bevacizumab for platinum-refractory CaO had been discontinued as a result of a higher rate of gastrointestinal (GI) perforations reported than in previous studies with bevacizumab (GI perforations are a known possible adverse event with bevacizumab) (Genentech, press release, September 23, 2005). Five GI perforations were observed in the Þrst 44 patients enrolled in the proposed 53-patient study. The patients currently enrolled in the trial will be informed of the new safety information and, in consultation with their physician, may continue to receive protocol treatment with bevacizumab or elect to discontinue treatment. Despite these negative results, Genentech states that it is continuing with plans to study bevacizumab in CaO based on the activity seen to date and the signiÞcant unmet need within this patient population. In addition, it must be noted, the patients enrolled in the discontinued study had late-stage disease, which typically involves the bowel, and had received more prior chemotherapy regimens than had patients in previous clinical trials of bevacizumab for CaO. Thalidomide. Thalidomide (Celgene’s Thalomid) (Figure 11) is an immunomodulator marketed for the treatment of leprosy. The compound is under investigation for several cancer indications, including multiple myeloma, colorectal cancer, and lung cancer. The Gynecologic Oncology Group has initiated a Phase III trial to compare thalidomide with tamoxifen in patients with CaO who have an elevated CA-125 level during follow-up front-line chemotherapy. Thalidomide suppresses levels of several growth factors, including tumor necrosis factor-alpha (TNF-alpha), basic Þbroblast growth factor (bFGF), VEGF, and interleukin-6 (IL-6), suppressing tumor growth and angiogenesis. Recently, several small studies were presented investigating the use of thalidomide to treat recurrent and advanced-stage CaO. The purpose of a study presented O N O

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FIGURE 11. Structure of thalidomide.

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at ASCO 2005 was to assess the rate of response and TTP of recurrent CaO and primary peritoneal cancer in women treated with carboplatin and thalidomide (Wong C, 2005). Patients were treated with carboplatin every 21 days and daily with thalidomide; both adjusted for toxicities. After six cycles and a CR, patients began six months of thalidomide treatment. Thirteen patients were initially enrolled in the study between October 2001 and September 2002. One was asked to leave the study and another dropped out because of carboplatin intolerance. The mean TTP on treatment was 98.0 days and the mean number of cycles was 4.7. Six of the 11 patients were diagnosed with progression by CT scan and three of 11 were diagnosed by rising CA-125. As of the close of the study, one patient was maintained on thalidomide and was free of disease. The researchers concluded that carboplatin and thalidomide demonstrate a brief progression-free interval with a minimal side-effect proÞle. In a prospective study presented at the 2005 Society of Gynecologic Oncologists’ Annual Meeting on Women’s Cancer, thalidomide was found to be similar in effectiveness and toxicity to IV chemotherapy in advanced-stage, recurrent CaO (Gordinier K, 2005). Patients eligible for the study had recurrent CaO after two prior regimens for recurrence. Eighteen patients received chemotherapy (regimen not speciÞed) and 18 received oral thalidomide (200 mg daily). Of the 15 evaluable patients on chemotherapy, one experienced a PR, nine had SD, and Þve had disease progression. For the 13 patients who received thalidomide, one experienced a PR, seven had SD, and Þve had disease progression. Statistically, the study found no difference in the progression-free interval, which was three to four months regardless of treatment. Carboxyamido-Triazole. The NCI is developing a micronized, oral formulation of carboxyamido-triazole (CAI), an angiogenesis inhibitor, as a potential treatment for a range of cancers. CAI is in Phase III studies for NSCLC and Phase II studies for metastatic renal-cell carcinoma and CaO in the United States. The agent was originally developed by Merck as L-651582 as a coccidiostat (a group of chemical agents mixed in feed or drinking water to control coccidiosis in animals) for use in poultry. The compound inhibits calcium inßux and calcium-mediated signaling events in endothelial cells, thereby suppressing their proliferation. Antitumor activity may also be exerted via inhibition of interleukin-8 (IL-8) and VEGF expression. A Phase II study investigated whether CAI would promote disease stabilization lasting at least six months in patients with relapsed CaO (Hussain MM, 2003). Oral CAI was given daily using a pharmacokinetic-dosing approach to maintain plasma concentrations between 2 and 4 µg/mL. Radiographic imaging to assess response was performed every eight weeks. Thirty-six patients were assessable for primary endpoint analysis, and 38 were assessable for toxicity. Forty-four percent of the patients had three prior regimens, more than 50% had four or more disease sites, and 48% had liver metastases. Thirty-three patients reached the targeted concentration range during the Þrst cycle. Eleven patients (31%) attained the >6-month outcome endpoint, with one PR (8 months) and three

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minor responses (8, 12+, and 13 months). Median TTP was 3.6 months (with a range of 1.6–13.3 months). CAI was well tolerated; toxicity was mostly grade 1 and 2. Grade 3 events included fatigue (5%), vomiting (2%), neutropenic fever (2%), and neutropenia (2%). No grade 4 adverse events occurred. Researchers concluded that CAI may have a role in maintenance therapy. Squalamine. Genaera (formerly Magainin) is developing squalamine (MSI1256F), a potent antiangiogenic agent and calmodulin-sequestering agent for the treatment of several cancers, including NSCLC and CaO, for which it is in Phase II studies in the United States. In 2001, the FDA granted squalamine orphan drug status for the treatment of CaO. An extract of dogÞsh shark liver, squalamine blocks endothelial-cell activation, proliferation, and migration. Preclinical data suggest that squalamine may suppress mitogen-activated protein (MAP) kinase signaling in VEGF-stimulated endothelial cells and that low doses of radiation could potentiate squalamine’s antiangiogenic effects. At ASCO in 2002, researchers presented data from a Phase IIa trial (Davidson SA, 2002), in which squalamine was administered as a Þve-day continuous infusion in conjunction with carboplatin (AUC 6) every three weeks at 200 mg/m2 /day to 33 patients who displayed either progression on primary paclitaxel and carboplatin therapy (refractory disease) or recurrence within six months of initial response to the same regimen (resistant disease). Response data were available for 26 of the enrolled patients. Nine objective responses were observed: Þve were CRs and four were PRs. Grade 4 thrombocytopenia, anemia, leukopenia, myalgia, and asthenia occurred in three patients; other grade 4 adverse events, including headache, nausea, and pain, occurred in two patients. Although the 19% CR rate appears promising, squalamine requires continuous infusion, and therefore any usage is likely to be restricted to hospitalized patients. This cumbersome administration protocol required for squalamine, together with the grade 4 toxicities, is likely to relegate this agent to end-stage use unless a signiÞcant survival beneÞt is achieved in a larger trial. Epidermal Growth Factor Inhibitors Overview. The epidermal growth factor receptor (EGFR, also known as HER or erbB) family includes four related transmembrane receptors: HER-1 (erbB1), HER-2 (erbB2), HER-3 (erbB3), and HER-4 (erbB4). Each receptor contains an extracellular-binding region and an intracellular kinase. The HER-3 kinase is defective; thus, signaling through HER-3 requires heterodimerization with another EGFR family member. HER-2 has no known natural ligand and so typically requires heterodimerization as well. However, at high concentrations, HER-2 homodimers may signal even in the absence of ligand. Ligand binding results in homo- or heterodimer formation with activation of the associated receptor tyrosine kinase. Receptor activation triggers multiple intracellular pathways, including the mitogen-activated protein kinase (MAPK) and AKT/PI3K that result in increased proliferation, resistance to apoptosis, and increased angiogenesis.

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A strong association is observed between expression of the EGFR tyrosine kinases and the aggressiveness and prognosis of several types of solid tumors, including CaO. Mechanism of Action. When growth factors (such as epidermal growth factor and transforming growth factor-alpha) bind to the EGFR, the receptor’s intracellular enzymatic tyrosine kinase portion is activated and, in turn, triggers a cascade of intracellular events that stimulate growth and survival of the tumor cell. The activation of EGFR can be inhibited, thereby preventing proliferation and subsequent tumor growth, by blocking the binding of the ligand by the receptor. This blocking can be done with monoclonal antibodies (cetuximab [ImClone/Merck KGaA/BMS’s Erbitux] and Merck KGaA’s matuzumab, which are not discussed here because of insufÞcient clinical data) or inhibition of the enzyme activity of the receptor tyrosine kinase with small molecules such as geÞtinib (AstraZeneca’s Iressa) or erlotinib (Osi Pharmaceuticals/Genentech’s Tarceva). The next generation of EGFR inhibitors, such as pertuzumab (Genentech/Roche/Chugai’s Omnitarg; discussed in a later section) and GlaxoSmithKline’s lapatinib, will affect the interactions of erbB family members. The latter compound is a small-molecule dual inhibitor of EGFR (ErbB1) and HER2 (erbB2) that blocks the tyrosine kinases of both these receptors. Clinical trials of lapatinib are in progress for patients with breast or lung cancer, and a Phase II trial for persistent or recurrent CaO or peritoneal cancer has begun recruiting. Lacking clinical data, this section does not discuss lapatinib further. Gefitinib. GeÞtinib (AstraZeneca’s Iressa) was the Þrst EGFR tyrosine kinase inhibitor launched. Following Japanese approval in 2002, geÞtinib was launched in October 2003 in the United States for third-line treatment of advanced NSCLC after gaining accelerated FDA approval. However, in a follow-up Phase III NSCLC trial reported in December 2004, geÞtinib failed to meet its endpoint regarding survival. The FDA, whose accelerated approval program authorizes the agency to remove a drug from the market if a postmarketing clinical study fails to verify clinical beneÞt, subsequently issued a statement informing physicians and patients that “alternative therapies are available.” European approvals for geÞtinib are on hold. The compound is also undergoing several other clinical trials in various other cancer indications, including CaO in Phase II. GeÞtinib is an orally active, quinazoline-derived selective EGFR tyrosine kinase inhibitor that blocks signal transduction. The drug’s precise mechanism of action is unknown. The GOG-170C trial involving 30 patients with recurrent CaO found geÞtinib to be well tolerated (Schilder RJ, 2003). Fifteen patients had one prior chemotherapy regimen and 13 had two prior chemotherapy regimens. Patients received a median of two cycles. The primary endpoint was a progressionfree interval of at least six months on therapy to be achieved in at least 30% of patients. Four patients achieved this endpoint, including one patient who achieved a PR. Grade 3 or 4 hematologic toxicity was observed in one patient

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(grade 4 neutropenia). The following grade 3 or 4 nonhematologic toxicities were observed: skin (four grade 3), infection (one grade 3), pain (three grade 3), diarrhea (seven grade 3), emesis (two grade 3), other gastrointestinal (one grade 4), and hypokalemia (two grade 3). The researchers concluded that geÞtinib has minimal activity as a single agent in this unselected population of patients with recurrent disease. However, molecular characterization of tumor subsets may identify patients who are likely to beneÞt from EGFR tyrosine kinase inhibition resulting in tumor growth inhibition. To test this theory, the researchers examined archived tumor tissue from the Phase II trial, and they sequenced genes in an additional 32 CaO tumor samples not treated with geÞtinib (Schilder R, 2005). Overall, the researchers detected mutations in the tyrosine kinase domain region of EGFR in two of 56 (3.6%) ovarian adenocarcinomas and observed that a patient in the clinical trial with a mutation in the catalytic domain of EGFR responded to geÞtinib. This Þnding suggests a method to preselect a subset of patients whose tumors may be more responsive to this EGFR-targeted therapy. Erlotinib. Erlotinib (Tarceva) is an orally active selective inhibitor of the EGFR tyrosine kinase that has been developed and launched by Osi Pharmaceuticals and Genentech for the treatment of NSCLC. Osi, Genentech, Roche, and Roche’s Japanese subsidiary, Chugai (formerly Nippon Roche), are also developing erlotinib for the potential treatment of various other solid tumors; the compound is in preregistration for pancreatic cancer. The Scottish Randomized Trial in Ovarian Cancer (SCOTROC) will soon begin evaluating erlotinib as part of front-line therapy for advanced CaO. This Phase III trial will use erlotinib as postinduction treatment following carboplatin and docetaxel. Like geÞtinib, erlotinib is an oral, once-a-day, small-molecule agent that inhibits the intracellular tyrosine kinase domain of the EGFR, thus blocking receptor activity. Data from a Phase II trial were reported at ASCO 2001 (Finkler N, 2001). Thirty-four patients with platinum-refractory CaO received daily oral doses of 150 mg erlotinib. Researchers observed a 10% PR rate. At eight weeks, they noted that a further 50% of the 30 evaluable patients showed evidence of disease stabilization. Adverse events included an acneiform rash affecting the face, upper torso, and arms; diarrhea, nausea, and vomiting; headache; and fatigue. A trial presented at ASCO 2005 investigated the tolerability of carboplatin, paclitaxel, and erlotinib as a Þrst-line treatment for CaO, fallopian tube cancer, or primary peritoneal cancer (Blank SV, 2005). Since June 2003, patients with stage III or IV CaO have received paclitaxel (175 mg/m2 ) and carboplatin (AUC 6) every 21 days, along with erlotinib (150 mg) orally daily. Data are based on 28 patients enrolled thus far in the trial. Sixteen patients completed the full six cycles of chemotherapy with erlotinib, and 13 have undergone reassessment surgery. Researchers conclude that the addition of erlotinib to paclitaxel and carboplatin as a front-line regimen for CaO and related epithelial cancers is feasible and well tolerated similar to standard chemotherapy, with the exception

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of skin rash; all patients who completed therapy experienced at least a grade 1 rash. Cetuximab. Cetuximab (Erbitux, IMC-C225) was developed by ImClone, in collaboration with Merck KGaA and BMS. The product was Þrst launched in 2004 for the IV treatment of irinotecan-refractory colorectal cancer. The companies are also evaluating the compound both as a single agent and in combination with current therapies for numerous solid tumors; the compound is in Phase II for the treatment of CaO. Cetuximab is a recombinant, human-murine chimeric monoclonal antibody. The antibody binds to HER1 on both normal and tumor cells and competitively inhibits the binding of EGF and other ligands, such as transforming growth factoralpha. It is composed of the Fv regions of a murine anti-EGFR antibody with human IgG1 heavy and kappa light chain constant regions. Preliminary data presented at ASCO 2005 from an ongoing study showed that cetuximab in combination with standard paclitaxel/carboplatin chemotherapy was well tolerated in 31 previously untreated patients having advanced CaO with EGFR-positive tumors (Aghajanian C, 2005). Patients received an initial dose of cetuximab 400 mg/m2 on day 1 of cycle 1, followed by 250 mg/m2 weekly. Paclitaxel (175 mg/m2 ) and carboplatin (AUC 6) were given on day 1 of each three-week cycle. The presentation featured preliminary data from the Þrst 27 patients completing six cycles of initial combination therapy. Fourteen patients showed CRs and were eligible to continue cetuximab monotherapy. A total of 18.5% of patients had grade 3 and 4 febrile neutropenia during initial combination therapy, with no febrile neutropenia during maintenance monotherapy with cetuximab. Grade 3 and 4 nonhematologic toxicities were uncommon in this patient population. The researchers concluded that long-term maintenance treatment with cetuximab monotherapy is feasible and well tolerated. Pertuzumab. Genentech/Roche/Chugai’s next-generation HER-2-directed monoclonal antibody, pertuzumab (Omnitarg), inhibits dimerization (the pairing of receptors) of HER-2 with EGFR. It is in Phase II clinical trials for a range of solid cancers, including CaO. Genentech claims that pertuzumab represents the Þrst in a new class of targeted potential therapeutic agents known as HER-dimerization inhibitors (HDIs). Binding of the antibody to the extracellular dimerization domain of HER2 directly inhibits the ability of HER2—the most common pairing partner—to dimerize with other HER-receptor proteins. Inhibiting receptor dimerization prevents the activation of HER-signaling pathways. The mechanism of action of pertuzumab is distinct from tyrosine kinase inhibitors, such as erlotinib and geÞtinib, which bind competitively to the intracellular adenosine-triphosphate-binding site of the HER receptors. In May 2005 at the 41st ASCO meeting, Phase II data were presented from a study in which 65 patients with relapsed CaO were administered a loading dose of 840 mg, followed by 420 mg pertuzumab IV every three weeks (Gordon MS,

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Pancreatic Cancer

ETIOLOGY AND PATHOPHYSIOLOGY Introduction The human pancreas is a secretory organ with two main functions: to produce hormones (endocrine pancreas) and to produce a number of digestive enzymes (exocrine pancreas). In the endocrine pancreas, the pancreatic production of insulin, somatostatin, and glucagon plays an important role in regulating sugar and salt balance in the body. The digestive enzymes from the exocrine pancreas are secreted into the intestines to help digest food. In the normal pancreatic architecture, acinar cells, which secrete the digestive enzymes chymotrypsin, trypsin, and carboxypeptidase, make up more than 80% of pancreatic cells; the islet cells, which secret insulin, make up only 1–2% of the pancreas. Ten to Þfteen percent of the pancreas is structured of single-layered, cuboidal ductal cells; a sparse interlacing network of blood vessels, lymphatic vessels, nerves, and collagenous stroma is also present. Figure 1 shows the structure of the normal human pancreas and the altered structure in a carcinoma. This architecture is clearly altered in carcinoma, in which the predominant histological feature in pancreatic cancer (PC) is a dense collagenous stroma and a moderate increase in the number of ducts of both normal and cancerous appearance (Evans DB, 2001[a]). Islet cell clusters are generally preserved. The vast majority (95%) of malignant neoplasms arise from the exocrine portion of the gland; malignant neoplasms originate nine times more often from duct cells than from acinar cells (Evans DB, 2001[a]; Ringel J, 2003). As many as 70–80% occur in the head of Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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Stomach Spleen Liver Gallbladder

Pancreas

Small intestine Large intestine

Anatomical location in the abdomen Hepatic ducts of liver Gallbladder

Common hepatic duct Bile duct Pancreas Small intestine Pancreatic duct

Neighboring ducts and tissues Celiac plexus Pancreatic duct

Portal vein

Pancreas tail

Small intestine Tumor invasion of small intestine

Pancreas body

Bile duct

Mesenteric plexus Pancreas head

Tumor invasion from the head of the pancreas into the small intestines

FIGURE 1. Diagram of the pancreas.

the pancreas (Cancer Help UK, 2004). Endocrine tumors, arising from islet cells, are seen much less frequently, while nonepithelial lymphomas or sarcomas are extremely rare. Because of the low incidence of nonexocrine tumors, this section focuses on exocrine PC. Oncogenes and Tumor Suppressor Genes A number of oncogenes and tumor suppressor genes (TSGs) have been linked with PC (Table 1). Oncogenes stimulate cell growth, whereas TSGs coordinate

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TABLE 1. Oncogenes and Tumor Suppressor Genes in Pancreatic Cancer

Gene

Function in Normal Cells

Effect of Mutation

Frequency in Pancreatic Cancer

Possible Applications

K-ras

Promotes cell division

Constitutive activation

75–90%

p53

Promotes cell death, inhibits cell division Inhibitor of cell-cycle progression Prevents cell division

Inactivation

50–75%

Inactivation

>80%

Prognostic value. Inherited mutation

Inactivation

>60%

Unique to pancreatic cancer

p16

DPC4

Prognostic value in combination with other markers. Early cancer detection. Therapeutic target Prognostic value. Therapeutic target

and limit cell growth. Mutations in at least three TSGs—deleted in pancreatic cancer 1/2 (DPC1/2), DPC3, and DPC4—are common in pancreatic tumors (Heinm¨oller E,2000). Oncogenes K-ras. The ras oncogene family (K-ras, H-ras, and N-ras) encodes cellmembrane guanosine-triphosphate (GTP) proteins that are involved in cell growth. Mutations that activate the ras oncogene result in inappropriate, prolonged signaling for continued cell growth and division. Mutations in ras genes contribute to 20–30% of all types of cancer, but are involved in 75–90% of PC tumors (Evans DB, 2001[a]; Cowgill SM, 2003). K-ras mutation appears to occur early in the carcinogenesis process (Inoue S, 2001), suggesting that it is a potential target for gene-based screening or diagnostic tests. Bcl-2. The proto-oncogene bcl-2 (B-cell lymphoma/leukemia-2) encodes a protein—pBcl-2—that blocks apoptosis (Nio Y, 2001). Overproduction of pBcl2 allows cells to escape apoptosis. In some studies, pBcl-2 expression was found to be a beneÞcial prognostic factor in PC. A trend toward improved survival was seen in well-differentiated tumors and in those with increased pBcl-2 expression (Sinicrope FA, 1996; Nio Y, 2001). However, other studies have found that although pBcl-2 regulates pancreatic morphogenesis and can be considered a phenotypic marker of normal exocrine pancreas, pBcl-2 does not play a central role in pancreatic tumorigenesis and cancer progression (Campani D, 2001). Tumor Suppressor Genes. p53. The p53 protein normally prevents the propagation of DNA damage by causing cell arrest at the G1/S checkpoint and inducing apoptosis, or programmed

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cell death. Mutations in p53 remove this regulatory control, allowing mutationcarrying cells to progress through the cell cycle. p53 is the most commonly mutated gene product in human cancer, and more than 70% of pancreatic carcinomas display lost function (Hruban RH, 1998). Clinical outcome has been found to bear no relation to p53 expression (Sinicrope FA, 1996), although it has been suggested to be of beneÞt in predicting the effects of adjuvant chemotherapy (Nio Y, 2001). p16. The p16 gene regulates the function of a protein called pRb by inhibiting CDK4 enzyme activity. Inactivation of the p16 gene disrupts pRb-mediated cellcycle control and unregulated cell growth. Some 85% of pancreatic tumors lack normal p16 function (Caldas C, 1994). In a study investigating the incidence of malignant melanoma in melanoma-prone families, an increased risk of developing PC was, coincidentally, observed in family members with p16 mutation (Goldstein AM, 1995). Several studies have addressed the effects of p16 mutation on prognosis. One found that the survival period was shorter and metastasis more likely to occur in those cases that did not show p16 expression (Hu Y, 1997). While p53 mutation may be a relatively early event in pancreatic carcinogenesis, alteration of the p16 gene is more likely to be correlated with tumor progression (Naka T, 1998). Breast Cancer 2 (BRCA2). The BRCA2 gene appears to be a TSG that participates in DNA damage repair. Mutations in this gene have been linked to breast and ovarian cancers, and recent studies have shown that patients with BRCA2 mutation have a signiÞcantly increased risk of developing PC (Lin Y, 2001). DPC1/2, DPC3, and DPC4. Unlike K-ras, p53, and p16, which are associated with many different cancers, three TSGs have been identiÞed that appear to be speciÞc to PC: DPC1/2, DPC3, and DPC4. DPC1/2 is located on chromosome 13q (the region of the BRCA2 gene), DPC3 (p16/MTS-1) on chromosome 9q, and DPC4 on chromosome 18q. DPC4 (also called Smad4 ), a recently discovered TSG on chromosome 18q21, is an important component of the transforming growth factor-beta (TGF-β) signaling pathway that normally down-regulates the growth of epithelial cells, stimulates differentiation, and promotes apoptosis. DPC4 has been found to be deleted or inactivated in 50% of PC cases (Hruban RH, 1998). Consequent loss of this important growth regulatory pathway contributes to unregulated cell growth (Hahn SA, 1996). Although it is assumed that the growth inhibitory function of TGF-β is important for DPC4 tumor suppressor activity, recent evidence suggests that restoration of DPC4 function in human pancreatic carcinoma cells suppresses tumor formation in vivo but does not restore TGF-β sensitivity (Schneider G, 2003), implying that TGF-β is independent of DPC4 function. The ability to restore (and possibly overexpress) DPC4 function in human PC cells remains an active area of investigation and may have therapeutic potential in PC.

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Other Factors in PC CDC25. CDC25 phosphatases consist of CDC25A, -B, and -C. This family of cyclin-dependent kinase (CDK) activators act at different points of the cell cycle, including G1-S and G2-M transition. The cell cycle consists of events that result in cell growth and division through stages G1-S to G2-M. CDC25A is expressed in the late G1 phase and controls G1/S transition while CDC25B and -C are essential for the G2 and G2/M transition. CDC25A and -B, but not -C, exhibit oncogenic potential, and overexpression of CDC25A or -B has been found in different human tumors including non-Hodgkin’s lymphoma and lung, colorectal, and prostate cancers (Guo J, 2004). Researchers investigating the expression and functional signiÞcance of CDC25s in pancreatic ductal adenocarcinoma found that CDC25B mRNA was overexpressed in PC (7.5-fold) when compared with the normal pancreas (Guo J, 2004). A strong nuclear CDC25B immunoreactivity was observed in both pancreatic and metastatic cancer samples. The study found two novel CDC25B inhibitors (NSC663284 and NSC668394) that dose-dependently reduced the growth of PC cells by blocking G2/M phase transition. This result suggests an important role of CDC25B in cell-cycle progression and implies that inhibition of CDC25B may have therapeutic potential in PC (Guo J, 2004). Epidermal Growth Factors and Epidermal Growth Factor Receptors. Various growth factors are expressed at increased levels in PC, including epidermal growth factors (EGFs) and their receptors (HER-1 [also known as EGF-R], HER-2, HER-3, and HER-4), which promote cell division. These growth factors and receptors are expressed at high levels in more than 50% of PC tumors and thus represent potential targets for treatment. The concomitant presence of the EGF-receptor HER-1 and its ligands EGF and TGF-alpha (-α) is associated with enhanced tumor aggressiveness and shorter survival periods following resection. A number of other growth factors and their respective receptors, such as Þbroblast growth factor, nerve growth factor, platelet-derived growth factor, and insulinlike growth factor, are expressed at increased levels in PC and are thought to contribute to aggressive growth of the tumor (Ozawa F, 2001). Vascular Endothelial Growth Factor. Vascular endothelial growth factor (VEGF) is an endothelial cell-speciÞc mitogen that promotes angiogenesis in solid tumors. Angiogenesis, the process of developing new blood vessels, is essential for tumor growth and must occur for metastasis formation. VEGF upregulation occurs as a result of the activation of mutations of the ras oncogene (Rak J, 1995). The TSGs p53 and p16 have also been shown to regulate the expression of this factor (Bouver M, 1998; Harada H, 1999). Flk-1/KDR and ßt1, two VEGF receptors, are also overexpressed in PC (Itakura J, 2000). Although VEGF and its receptors are upregulated in this disease, pancreatic tumors are not highly vascularized and therefore this factor plays only a limited role in the development of PC.

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Sonic Hedgehog. Sonic Hedgehog (Shh), a secreted glycoprotein, is an essential signaling molecule in numerous embryonic processes, including pancreatic development. Shh signaling is activated by binding of Shh to a membrane receptor called patched (Ptc). Misregulation of Shh signaling has been implicated in several forms of cancer, including lung, basal cell carcinomas, and PC (Wicking C, 1999). Recent data suggest that overexpression of Shh may contribute to pancreatic tumorigenesis. The Shh pathway is required for both cell division and the suppression of apoptosis (Thayer SP, 2003). Cyclopamine, a steroidal alkaloid, inhibits Shh signaling through a direct interaction with Smo (serpentine receptor, smoothened), a proto-oncogene acting downstream of Shh. Cyclopamine represses Smo function, ultimately inducing apoptosis and blocking proliferation in a subset of PC cell lines both in vivo and in vitro (Thayer SP, 2003). Targeting Shh signaling may therefore be an effective therapeutic approach for the treatment of PC, and agents are already being investigated in preclinical studies. Mucins. Mucins are high-molecular-weight glycoproteins that are produced by various epithelial cells including those found in the pancreas. Under normal circumstances, mucins provide a protective layer on epithelial surfaces. In addition, their involvement in the renewal and differentiation of the epithelium and modulation of cell adhesion as well as cell signaling has been proposed (Ringel J, 2003). Mucins are synthesized either as membrane-bound or secreted glycoproteins. Generally, the membrane-bound mucin MUC1 is expressed in most epithelial tissues and is highly expressed in the pancreas and breast. A secreted mucin, MUC2, is expressed by the goblet cells of the colon, small intestine, and respiratory airways. It has been reported that MUC1 and MUC2 are expressed differentially between nonneoplastic and neoplastic pancreatic lesions in surgically resected tissue (Chhieng DC, 2003). Results of a clinical trial examining MUC1 and MUC2 expression in pancreatic lesions obtained by Þne-needle aspiration biopsy found MUC1 to be overexpressed in 96% of invasive pancreatic ductal carcinomas, 9% of nonneoplastic lesions, and none of the nonductal neoplasms. MUC2 immunoreactivity was noted in 13% of pancreatic ductal carcinomas and 9% of non-neoplastic lesions (Chieng DC, 2003). This Þnding suggests that the expression of MUC1 can be used as an ancillary marker for diagnosing pancreatic ductal carcinoma. However, MUC2 did not appear to be a useful marker for recognizing pancreatic ductal carcinoma (Chieng DC, 2003). Further studies are necessary to conÞrm the Þndings of this study. Risk Factors PC incidence is highest in industrialized countries, although considerable variation exists among geographically and ethnically dissimilar populations (e.g., relatively low rates have been reported in Spain compared with Northern Europe). As yet, no plausible explanation for these differences has been proposed (Evans DB, 2001[a]). The incidence of PC has historically been higher in males than in females. Over time, however, male incidence rates have fallen, while those in females

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have increased. The latter effect has been attributed to a rise in the percentage of women who smoke. Some ethnic groups appear to be particularly susceptible to the disease; in the United States, blacks suffer the highest incidence and mortality rates (Solomon L, 2002). Age. Age is the strongest risk factor for PC; the risk of developing PC increases with age. Incidence increases from less than 1 per 100,000 at age 35 to 96 per 100,000 after age 85. The median age at diagnosis is 69 years. Smoking. Cigarette smoking is the most consistently cited environmental risk factor for PC. Estimates suggest that 25–30% of cases are due to smoking and that smokers have a signiÞcant (70%) increased risk compared with nonsmokers (Fuchs C, 1996). The risk of PC increases in parallel with the daily amount smoked and the duration of smoking, while long-term cessation (>10 years) reduces risk by approximately 30% relative to the risk of current smokers (Silverman DT, 1994). Pancreatitis. Various studies have established an association between PC and chronic pancreatitis (Lowenfels AB, 1993), and estimates of risk suggest that chronic pancreatitis may explain as many as 5% of PC cases (Fernandez E, 1995). Increased cell division as a result of pancreatitis could heighten the risk of mutation leading to cancer, and some studies found a speciÞc K-ras mutation (mutation at codon 12) to be frequent in patients with chronic pancreatitis (25–42%), providing a possible molecular link between chronic inßammation and the initiation of pancreatic carcinogenesis (Yanagisawa A, 1993; Zalatnai A, 2003). However, such studies are compromised by difÞculties in distinguishing the symptoms of pancreatitis from those of early-stage PC and by the fact that early-stage PC can cause pancreatitis. The issue of cause and effect remains debatable and unsettled. Diabetes. An association between diabetes and PC has been recognized for many years, but the precise nature of the relationship has yet to be determined. Diabetes has been implicated both as a predisposing factor for, and as an early manifestation of, PC. A higher frequency of PC in patients with long-standing diabetes was reported in a meta-analysis of studies published from 1975 to 1995 (Everhart J, 1995). Against this background, a large-scale Italian study found that in 56% of cases of co-disease, diabetes was diagnosed either concomitantly with cancer or within two years of the cancer diagnosis, suggesting that diabetes was caused by the tumor and was not a risk factor (Gullo L, 1994). In a more recent study in northern Italy, researchers found that the overall risk for developing PC was higher for diabetes patients treated with insulin (relative risk = 6.49) than for patients treated with oral hypoglycemic drugs (relative risk = 2.12). Additionally, patients treated with insulin had a seven-fold increased risk for developing PC as compared with patients treated with oral antidiabetics (Bonelli L, 2003). Whatever

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the relationship, it is largely accepted that patients with new-onset diabetes, like patients with chronic pancreatitis, have a low but progressively increasing risk of having or developing PC (Simon B, 2001). Diet. Various studies have addressed the effect of diet on the risk of developing PC, but none has demonstrated conclusive evidence of an association. Meat and fat consumption appears to increase risk, whereas a diet high in fruits and vegetables has been suggested as conferring protection (Mills PK, 1988). Studies also suggest that increased risk of developing PC is associated with high consumption of salt, smoked meat, dehydrated food, fried food, and reÞned sugar. An inverse association was found with the consumption of food containing no preservatives and addictives, raw food, food prepared by high-pressure cooking, and food prepared using electricity or exposure to microwaves from microwave ovens (Li D, 2003). A number of carcinogenic materials such as heterocyclic amines and polycyclic aromatic hydrocarbons can be formed when grilling or barbecuing red meat. It has been suggested that this type of cooking should be regarded as a risk factor (Anderson KE, 2002). Alcohol consumption and coffee consumption have also been considered risk factors, but two large, prospective studies in the United States concluded that intake of neither was associated with a greater risk of PC (Michaud DS, 2001[a]). However, another analysis of the same study found that individuals with a body mass index (BMI) of at least 30 kg/m2 had an elevated risk (1.72:1) of PC compared with those with a BMI of less than 23 kg/m2 . Physical activity appeared to reduce risk, especially among the obese (Michaud DS, 2001[b]). Hereditary Disorders. A number of rare hereditary disorders predispose patients to PC. They include ataxia-telangiectasia, familial atypical multiple mole melanoma syndrome, Peutz-Jeghers syndrome, hereditary nonpolyposis colon cancer, BRCAI/BRCA2 in breast/ovarian cancers, and hereditary pancreatitis (Cowgill SM, 2003; Evans DB, 2001[a]). Currently, it is estimated that 5–10% of PC patients have a familial predisposition to the disease—they have one or more Þrst-degree relatives with PC (Li D, 2003; Lynch HT, 2001). Pathophysiology Prior to diagnosis, patients may present for several months complaining of vague, nonspeciÞc symptoms. These symptoms can range from upper abdominal and back pain and weight loss to symptoms of recent-onset diabetes such as glucose intolerance. These vague symptoms often complicate and delay deÞnitive diagnosis. The majority of patients suffer from late-stage disease when diagnosed. Jaundice due to biliary tract obstruction occurs in approximately 50% of patients at diagnosis and may be associated with a less advanced stage of disease than other symptoms (Bakkevold KE, 1992). Obstruction can be caused by a small tumor in the pancreatic head, with jaundice prompting the patient to seek medical attention while the mass is still localized and potentially resectable.

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Few patients present with potentially resectable disease in the absence of biliary obstruction. In locally advanced cancers, patients present with constant pain affecting the middle and upper back, caused by tumor invasion of the celiac and mesenteric plexus. Fatigue, rapid weight loss, and anorexia are common. Malabsorption and mild changes in stool frequency are also common; diarrhea occurs infrequently. Biliary or pancreatic duct obstruction can also cause abdominal pain. Invasion of the tumor into adjacent organs and regional metastasis is common in PC. Tumors in the pancreatic head extend into the stomach, duodenum, and colon; those of the body and tail frequently invade the spleen and adrenal glands (Figure 1). Tumor growth also results in compaction or invasion of the nerves surrounding the pancreas, a process that triggers abdominal pain. Metastasis into regional lymph nodes occurs in the majority of patients. Invasion into the splenic vein facilitates metastasis to the liver, and subclinical tumor nodules are often present in the liver on investigation. Peritoneal seeding and metastasis to the lung, although not as common as liver metastases, can also occur. Staging Several systems are used to stage PC. These systems can provide prognostic information and may aid clinical treatment decisions. The most widely used classiÞcation system was developed by the American Joint Committee on Cancer (AJCC) together with the Union Internationale Contre le Cancer (UICC). It uses TNM (primary tumor, lymph node spread, distant metastasis) criteria to classify PC based on the extent of primary tumor growth, regional lymph node involvement, and distant metastasis (Tables 2 and 3). This staging system has recently been updated to allow stage III to signify unresectable, locally advanced disease, while stage IV is reserved for patients with metastatic disease (AJCC, 2004). Table 3 shows the differences between the older and recently updated AJCC/TNM staging systems. Because most patients die within a few months of diagnosis, most patients with PC are not staged using the TNM system. Instead, clinicians often employ a clinical staging system that classiÞes a tumor based on whether it is resectable (localized), locally advanced, or metastatic (Table 4). This type of summary staging is frequently being used by tumor registries but is not fully characterized in the clinic (SEER, 2004). The summary staging system is the most basic staging system and is applicable to all anatomic sites (solid tumors, but not leukemias). Summary staging uses all medical, clinical, and pathological information available and is also referred to as general staging, California staging, and SEER staging (SEER, 2004). In Japan, physicians use staging criteria developed by the Japan Pancreas Society (JPS), although the UICC system is employed for international communications. The revised JPS classiÞcation published in 2002 (Table 5) is more reliable for predicting outcome compared with the UICC classiÞcation (Isaji S, 2004).

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TABLE 2. AJCC/UICC TNM Classification System for Pancreatic Cancer Primary Tumor (T) TX T0 Tis T1 T2 T3 T4

Criteria Primary tumor cannot be assessed No evidence of primary tumor In situ carcinoma Tumor 2 cm or less in greatest dimension Tumor more than 2 cm in greatest dimension Tumor extends directly to any of the following: duodenum, bile duct, or peripancreatic tissues Tumor extends to any of the following: stomach, spleen, colon, or adjacent large vessels

Regional node status (N) NX N0 N1 pN1a pN1b

Regional nodes cannot be assessed No regional node metastasis Regional node metastasis Regional node metastasis in a single regional node Regional node metastasis in multiple regional nodes

Distant metastasis (M) MX M0 M1

Distant metastasis cannot be assessed No distant metastasis Distant metastasis

AJCC = American Joint Committee on Cancer. TNM = Primary tumor, lymph node spread, distant metastasis. UICC = Union Internationale Contre le Cancer (International Union Against Cancer).

TABLE 3. AJCC/UICC Staging System for Pancreatic Cancer Stage 0 I IA IB II IIA IIB III IV IVA IVB

TNM Criteriaa

New TNM Criteriab (updated 2004)

Tis, N0, M0 T1-2, N0, M0 N/A N/A T3, N0, M0 N/A N/A T1-3, N1, M0 N/A T4, any N, M0 Any T, any N, M1

Tis, N0, M0 N/A T1, N0, M0 T2, N0, M0 N/A T3, N0, M0 T1-3, N1, M0 T4, any N, M0 Any T, any N, M1 N/A N/A

a See Table 2 for definitions of TNM criteria. b Definitions of the new TNM criteria are given below.

AJCC = American Joint Committee on Cancer; N/A = Not applicable; TNM = Primary tumor, lymph node spread, distant metastasis; UICC = Union Internationale Contre le Cancer (International Union Against Cancer). Notes: The T classification reflects the distinction between potentially resectable (T3) and unresectable (T4) primary pancreatic tumors. Tis = Carcinoma in situ. (This also includes the ‘‘PanInIII’’ classification) T3 = Tumor extends beyond the pancreas, but without involvement of the celiac axis or the superior mesenteric artery. T4 = Tumor involves the celiac axis or the superior mesenteric artery (unresectable primary tumor).

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TABLE 4. Clinical Staging of Pancreatic Cancer Clinical Stage

Description

TNM Criteria

I

Resectable

T1–3, selected T4, NX, M0

II

Locally advanced

T4, NX-1, M0

III

Metastatic

T1-4, NX-1, M1

Additional Characteristics No encasement of the celiac trunk or SMA Patent superior mesenteric vein-portal vein confluence No extra pancreatic disease Tumor extension involving celiac axis or SMA, or venous occlusion No extra pancreatic disease Metastatic (typically to the liver and peritoneum)

SMA = Superior mesenteric artery; TNM = Primary tumor, lymph node spread, distant metastasis.

TABLE 5. Japan Pancreas Society Staging System for Pancreatic Cancer

Stage I II III IV

Tumor Size (cm)

Lymph Node Metastasis

Anterior Capsular Invasion

Retroperitoneal Invasion

Portal Vein Involvement

T1, <2 T2, 2 to 4 T3, 4 to 6 T4, >6

N0 N1 N2 N3

S0 S1 S2 S3

Rp0 Rp1 Rp2 Rp3

PV0 PV1 PV2 PV3

N 0 = No involvement. 1 = Primary group of lymph nodes. 2 = Secondary group of lymph nodes. 3 = Tertiary group of lymph nodes.

S, Rp, PV 0 = Absence of tumor invasion. 1 = Suspected invasion. 2 = Definite invasion. 3 = Severe invasion.

Prognosis The prognosis for the vast majority of patients with exocrine PC is poor because the survival rate is very low. Fewer than 20% of newly diagnosed cases survive two years and only 3% of cases live for Þve years (Solomon L, 2002). Patients presenting with tumors conÞned to the pancreatic head—those with the best prognosis—have a Þve-year survival rate of approximately 20% and a median survival of 13–20 months because local or regional recurrence is common even among these patients. Median survival in patients with locally advanced, nonmetastatic disease is six to ten months, while that for metastatic disease is three to six months (Evans DB, 2001[a]). Additionally, some reports suggest that patients with unresectable PC live about 6 months; the Þve-year survival for resected cases is about 10–19% with a median survival of 12–18 months (Zalatnai A, 2003). A number of authors

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claim a Þve-year survival rate after resections of 30–58% of diagnosed incident cases, although it is likely that these Þgures refer only to limited cases. PC’s poor prognosis is largely due to the lack of effective therapy and the difÞculties in diagnosis (Solomon L, 2002; Zalatnai A, 2003). CURRENT THERAPIES Effective therapy for pancreatic cancer (PC) is very limited but can include surgery, radiotherapy, and chemotherapy. Surgery is the only potentially curative treatment, but owing to the tendency of pancreatic tumors to present late in the disease course, less than 20% of patients have resectable disease at presentation. Furthermore, most of these resectable patients will progress to advanced PC (APC, deÞned as nonresectable locally advanced disease or metastatic disease) for which the standard treatment is chemotherapy. Radiotherapy is sometimes used in conjunction with surgery and chemotherapy for resectable disease, but its role in nonresectable disease is as yet uncertain. Most patients with PC are therefore potential candidates for systemic management. However, for many reasons, the medical management of PC presents a considerable therapeutic challenge to oncologists. First, almost 70% of patients are age 65 or older at diagnosis, so many have premorbid conditions that limit their treatment options. Additionally, more than 80% of patients have diseaserelated symptoms, such as pain, weakness, weight loss, and poor performance status, that limit the option to deliver potentially toxic chemotherapy. In addition to these patient-related factors, PC is less chemosensitive than other commonly occurring solid malignancies; best-response rates to conventional agents are less than 10%. The reasons for this chemoresistance are poorly understood. Comparing chemotherapy clinical trial results in PC is difÞcult. Most trials recruit both locally advanced, nonresectable patients and metastatic patients. Because these two groups of patients have different survival times, the average length of survival reported in trials can vary widely depending on the case mix of the trial patients. Physicians have attempted to treat PC with many drugs, administered both singly and in combination. However, only two agents, 5-ßuorouracil (5-FU, generics) and gemcitabine (Eli Lilly’s Gemzar), have so far demonstrated sufÞcient clinical beneÞt to be accepted as standard monochemotherapies. Unfortunately, this clinical beneÞt is meager (median survival of APC patients is only six months) regardless of which current therapy patients receive. Because of the poor outcome of APC patients treated with 5-FU or gemcitabine monotherapy, physicians have investigated the efÞcacy of combination regimens. However, given that most PC patients present with poor performance status and comorbidities, the increased toxicities associated with combination chemotherapy signiÞcantly limit the use of these combination regimens in PC. Clinical trial results of 5-FU in combination with other agents have not been convincing; even discounting the increased toxicities of 5-FU combinations compared with single agents, combinations offer no substantial efÞcacy beneÞts over single agents (Berlin J, 2001; Di Costanzo F, 2001).

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NH2 N HO

O O

OH

N F F

FIGURE 2. Structure of gemcitabine.

The outlook for gemcitabine combinations is more optimistic; some trials have shown promising activity. Trials are proceeding with established chemotherapies such as cisplatin (Bristol-Myers Squibb’s Platinol-AQ, Nippon Kayaku’s Randa, generics), docetaxel (Aventis’ Taxotere), irinotecan (PÞzer’s Camptosar, Yakult/Aventis’ Campto, Daiichi’s Topotecin), oxaliplatin (SanoÞ-Synth´elabo’s Eloxatin), and paclitaxel (Bristol-Myers Squibb’s Taxol, generics), as well as with novel agents not yet approved for any indication (see “Emerging Therapies”). Table 6 summarizes chemotherapy regimens commonly used to treat PC. Gemcitabine, Single Agent Overview. Gemcitabine (dißuorodeoxycytidine) (Figure 2) is a deoxycytidine analogue with structural and metabolic similarities to those of the nucleoside cytarabine. Gemcitabine has superseded 5-FU as the Þrst-line chemotherapy for APC. Administered intravenously, it is indicated as a Þrst-line treatment for APC patients and for patients previously treated with 5-FU. Although gemcitabine is indicated for single-agent use in treating PC, it is sometimes used in combination with cytotoxic agents that are approved for other cancer indications. However, these gemcitabine-containing combinations are considered investigational, so they are discussed in the “Emerging Therapies” section. Gemcitabine has demonstrated activity in a variety of solid tumors and certain hematological malignancies. The growing importance of gemcitabine as a cytotoxic agent stems from its broad activity against common cancers and from an understanding of its mechanisms of action, enabling pharmacological intervention to potentiate its therapeutic indices. Mechanism of Action. Gemcitabine is an antimetabolite that shares characteristics common to all nucleoside analogues, including mediated transport by membrane transporters, activation by intracellular metabolic steps that retain the nucleotide residues in the cell, and the formation of active phosphate derivatives. Gemcitabine exhibits cell-phase speciÞcity, primarily killing cells undergoing DNA synthesis (S-phase) and blocking the progression of cells through the G1/S-phase boundary. It is metabolized intracellularly by nucleoside kinases to the active diphosphate and triphosphate nucleosides. The cytotoxic effect of gemcitabine is attributed to a combination of two actions of the diphosphate and the triphosphate nucleosides, which leads to inhibition of DNA synthesis. First,

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TABLE 6. Current Regimens Used for Pancreatic Cancer Regimen Components Regimen

Agent

Availability

Dose

Gemcitabine, Gemcitabine US, F, G, single (Eli Lilly’s I, S, agent, Gemzar) UK, J doseintense infusion Gemcitabine, Gemcitabine US, F, G, single (Eli Lilly’s I, S, agent, fixedGemzar) UK, J dose-rate infusion 5-FU, single 5-fluorouracil US, F, G, agent, (generics) I, S, continuous UK, J infusion

1,000 mg/m2 /week for 7 weeks followed by 1 week’s rest. Then weekly for 3 weeks followed by 1 week’s rest. Repeat 28-day cycle. 1,500 mg/m2 at a rate of 10 mg/m2 /min. Dosed weekly for 3 weeks followed by 1 week’s rest.

5-FU, single 5-fluorouracil US, F, G, agent, bolus (generics) I, S, UK, J

400–500 mg/m2 /day for 5 days

5-FU/LV

5-FU: 425 mg/m2 IV bolus.

5-fluorouracil US, F, G, (generics) I, S, UK, J

Leucovorin (generics) IV = Intravenous.

US, F, G, I, S, UK, J

1,000 mg/m2 /day continuous infusion for 5 days per week for 4 weeks

Leucovorin: 20 mg/m2 /d for 5 days. Repeat every 4 weeks for 6 months. Or 400 mg/m2 IV bolus followed by 5-FU: 600 mg/m2 IV infusion over 22 hours. Repeat on second day; repeat cycle after 2 weeks for 6–12 months. Leucovorin: 200 mg/m2 IV over 2 hours prior to 5-FU.

Common Toxicities Anemia Nausea/vomiting Thrombocytopenia Neutropenia Leukopenia Anemia Nausea/vomiting Thrombocytopenia Neutropenia Leukopenia Myelosuppression Neurotoxicity Mucositis Stomatitis Diarrhea Renal failure Nausea/vomiting Alopecia Infection Myelosuppression Neurotoxicity Mucositis Stomatitis Diarrhea Renal failure Nausea/vomiting Alopecia Infection Myelosuppression Neurotoxicity Mucositis Stomatitis Diarrhea Renal failure Nausea/vomiting Alopecia Infection

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gemcitabine diphosphate inhibits ribonucleotide reductase, which is responsible for catalyzing the reactions that generate the deoxynucleoside triphosphates for DNA synthesis. Inhibition of this enzyme by the diphosphate nucleoside causes a reduction in the concentrations of deoxynucleotides, including 2’-deoxycytidine 5’-triphosphate (dCTP). Second, gemcitabine triphosphate competes with dCTP for incorporation into DNA. The reduction in the intracellular concentration of dCTP (by the action of the diphosphate) enhances the incorporation of gemcitabine triphosphate into DNA (self-potentiation). After the gemcitabine nucleotide is incorporated into DNA, only one more nucleotide is added to the growing DNA strands. After this addition, further DNA synthesis is inhibited. DNA polymerase epsilon is unable to remove the gemcitabine nucleotide and repair the growing DNA strands (masked chain termination). Clinical Performance. In the Þrst multi-center trials reported on gemcitabine, objective responses were documented in only 11% of patients with APC. However, even in the absence of responses, the authors noted improvements in subjective symptomatic beneÞts (Casper ES, 1994). As a result of these observations, two pivotal trials were completed, and they established gemcitabine as the current standard chemotherapy for the treatment of APC. The primary efÞcacy parameter in these studies was “clinical beneÞt response,” assessed by measuring analgesic consumption, pain intensity, performance status, and weight change. One study compared gemcitabine with 5-FU in 126 previously untreated patients with unresectable disease, randomized to either gemcitabine or 5-FU (Burris HA, 1997). Median survival durations were 5.65 and 4.41 months for gemcitabine-treated patients and 5-FU-treated patients, respectively, while the survival rate at 12 months was 18% for the gemcitabine group and 2% for the 5-FU group. In addition, greater clinical beneÞt responses were seen with gemcitabine (24%) than with 5-FU (5%). The second study, which enrolled patients refractory to previous 5-FU regimens, conÞrmed these results. Median survival for the 63 patients treated with gemcitabine was 3.85 months, and a favorable clinical beneÞt response was achieved in 27% of patients (Rothenberg ML, 1996). Gemcitabine was considered to have a manageable toxicity proÞle in both studies, with comparatively low incidences of grade 3 or 4 toxicities. Common side effects of gemcitabine included ßu-like symptoms, leukopenia, and nausea. The most active dose and schedule of gemcitabine are being investigated in a Phase III Eastern Cooperative Oncology Group (ECOG) trial. The concept of a “Þxed-dose-rate” (FDR) infusion was prompted by pharmacokinetic data suggesting that the active metabolite, gemcitabine triphosphate, could be accumulated more effectively using a longer infusion. A Phase II trial randomized 92 patients to receive a dose-intense gemcitabine treatment of 2,200 mg/m2 as a 30-minute infusion (standard arm), or an FDR infusion with 1,500 mg/m2 at a rate of 10 mg/m2 /min on days 1, 8, and 15 of every four-week cycle (Tempero M, 2003). Results showed superior clinical activity for the FDR arm over the dose-intense arm. Median survival was Þve months in the standard arm and eight months in the FDR arm. The one- and two-year survival rates for all patients

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FIGURE 3. Structure of 5-fluorouracil (5-FU).

were 9% (standard arm) versus 28.8% (FDR) and 2.2% (standard arm) versus 18.3% (FDR), respectively. Patients in the FDR infusion arm experienced consistently more hematologic toxicity. Pharmacokinetic analyses demonstrated a two-fold increase in intracellular gemcitabine triphosphate concentration in the FDR arm, indicating more-effective inhibition of DNA synthesis. Although some investigational gemcitabine-containing cytotoxic combinations have shown positive early-phase trial data, a randomized Phase III trial of 322 APC patients has shown that gemcitabine in combination with 5-FU is not signiÞcantly more active than single-agent gemcitabine (Berlin JD, 2003). Median survival was 5.4 months for gemcitabine alone and 6.7 months for gemcitabine plus 5-FU. Progression-free survival for gemcitabine alone was 2.2 months, compared with 3.4 months for gemcitabine plus 5-FU. Objective responses were uncommon and were observed in only 5.6% of patients treated with gemcitabine and 6.9% of patients treated with gemcitabine plus 5-FU. Greater occurrences of hematologic and gastrointestinal toxicities were observed with gemcitabine/5-FU than with gemcitabine alone. Although the gemcitabine/5-FU arm displayed a trend toward improved activity compared with the gemcitabine single-agent arm, the difference was not signiÞcant. No other signiÞcant differences between the two treatment arms were noted. 5-FU, Single Agent Overview. First synthesized in 1957 (Heidelberger C, 1957) (Figure 3), 5-FU has been used to treat PC since the 1970s. Although the agent is the most extensively evaluated chemotherapeutic treatment in PC, few studies have rigorously addressed the response rate achieved with 5-FU as monotherapy. The agent’s use for treating colorectal cancer prompted its use in the treatment of PC; 5-FU’s use in PC has also diminished since the introduction of gemcitabine, a more active agent, in the late 1990s. The rationale for developing ßuoropyrimidines such as 5-FU came from the observation that rat cancer (hepatoma) cells use uracil, a pyrimidine, more efÞciently than do normal rat intestinal cells. This Þnding suggested that uracil metabolism might represent a potential therapeutic anticancer target. Many ßuoropyrimidines exist, but 5-FU is the only one that has found widespread acceptance as a standard treatment for gastrointestinal cancers. In fact, 5-FU is the only intravenously administered ßuoropyrimidine used in the treatment of PC. However, more recently, with the approval of the oral ßuoropyrimidine

CURRENT THERAPIES

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capecitabine (Roche’s Xeloda) for the treatment of colorectal cancer, physicians have been using capecitabine off-label for treating PC. This switching from 5-FU to capecitabine has occurred because the latter’s tablet formulation is much easier to administer. Capecitabine is considered an “investigational” PC therapy, so it is discussed in the “Emerging Therapies” section. 5-FU is traditionally given as a bolus (400–500 mg/m2 /day for Þve days), but the cytotoxic mechanism suggests a beneÞt in prolonged administration. For this reason, 24-hour continuous infusion is also employed. Although 5-FU has been evaluated as a treatment for PC in combination with other cytotoxic drugs, results have not been as positive as with gemcitabine combinations. Therefore, unlike gemcitabine-containing combinations, 5-FU-containing combinations are very rarely used and are no longer considered investigational in the treatment of PC. Mechanism of Action. 5-FU shares a biochemical pathway common to all ßuoropyrimidines and exerts its cytotoxic effect via several mechanisms. It requires intracellular activation to exert its effects, and it enters the cell in various ways. The agent is enzymatically converted to the active metabolite 5-ßuoro2’-deoxyuridine monophosphate (FdUMP). One of the most important targets for 5-FU is thymidylate synthase (TS), a key enzyme in de novo thymidine synthesis. This pyrimidine is an essential requirement for normal DNA synthesis. In the presence of a folate cofactor, FdUMP forms a stable covalent complex with TS, thereby inhibiting thymidine synthesis. Because 5-FU is a structural analogue of uracil, it is also misincorporated into RNA in place of uracil, interfering with normal RNA function and leading to cell death. Clinical Performance. Analysis of data collected from trials conducted before computed tomography (CT) scanning was common suggested response rates of 28% (Carter SK, 1975). However, CT scanning has more accurately shown that response rates of less than 10% can be expected. Four randomized trials conducted in the past decade have uniformly showed that 5-FU is only weakly active in the treatment of PC. Altogether, 313 patients treated with 5-FU reached a remission rate of 3.8%; survival times of between 3.5 and 5.1 months are not signiÞcantly greater than what would be expected for best supportive care alone (Burris HA, 1997; Cullinan S, 1990; Maisey N, 2001; Rougier P, 1999). 5-FU has also been studied in trials in combination with other chemotherapies, but data have not provided unequivocal evidence for using 5-FU combinations. One study did show a signiÞcant beneÞt of 5-FU combination over 5-FU alone (Rougier P, 1999). Locally advanced or metastatic patients (n = 207) were randomized to receive 5-FU plus cisplatin or 5-FU alone. After a median of two cycles, the incidence of grade 3/4 World Health Organization toxicity was lower in the 5-FU arm than in the 5-FU plus cisplatin arm (20% versus 48%; neutropenia: 6% versus 23%; vomiting: 4% versus 17%; mucositis: 5% versus 14%). Six-month and one-year survival for the 5-FU arm and the 5-FU plus cisplatin arm were 28% versus 38% and 9% versus 17%, respectively. Response rates

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(intent-to-treat analysis) were 0% for the 5-FU arm and 12% for the 5-FU plus cisplatin arm. Another randomized trial comparing 5-FU alone with 5-FU plus mitomycin C in 208 advanced PC patients did not show a signiÞcant beneÞt of using the 5-FU combination over 5-FU alone (Maisey N, 2002). After a median follow-up of 13.6 months, the overall response rate was 8.3% in the 5-FU arm compared with 20% in the 5-FU plus mitomycin C arm. No signiÞcant difference in median survival between the two arms was observed (5.1 months for 5-FU and 6.5 months for 5-FU plus mitomycin C). One-year survival was 23.3% for 5-FU and 28.2% for 5-FU plus mitomycin C. Global quality of life (QoL) improved signiÞcantly after 24 weeks of treatment compared with baseline only for the 5-FU plus mitomycin C, but no statistical difference in QoL was observed between treatment arms. Prior to commencement of treatment, 126 patients had pain, 88 had anorexia, 113 had weight loss, and 120 had lethargy. Considerable symptom improvement was reported in both arms, but no signiÞcant difference was observed between arms. Toxicities in both arms were mild and generally occurred in less than 10% of patients in both arms, with the exception of lethargy, which occurred in 28% of patients taking 5-FU monotherapy versus 25% for those on 5-FU plus mitomycin C. The study results showed an increased incidence of neutropenia in the combination arm, but no difference in the incidence of infection. 5-FU/Leucovorin Overview. A major milestone in the development history of 5-FU was the ability to biomodulate its activity. Biomodulation involves the concomitant administration of 5-FU with another drug to improve its therapeutic index. Many 5-FU modulators have been used in the past 20 years, but leucovorin (LV; generics) has emerged as the most active agent. Unfortunately, available data demonstrate that LV has this synergistic activity with 5-FU only in treating colorectal cancer, not PC (Crown J, 1991; DeCaprio JA, 1991; Evans DB, 2001). Mechanism of Action. •

•

5-FU shares a biochemical pathway common to all ßuoropyrimidines and exerts its cytotoxic effect via several mechanisms. It requires intracellular activation to exert its effects, and it enters the cell in various ways. The agent is enzymatically converted to the active metabolite 5-ßuoro-2’-deoxyuridine monophosphate (FdUMP). One of the most important targets for 5-FU is thymidylate synthase (TS), a key enzyme in de novo thymidine synthesis. This pyrimidine is an essential requirement for normal DNA synthesis. In the presence of a folate cofactor, FdUMP forms a stable covalent complex with TS, thereby inhibiting thymidine synthesis. Because 5-FU is a structural analogue of uracil, it is also misincorporated into RNA in place of uracil, interfering with normal RNA function and leading to cell death. LV is a biochemical modulator of 5-FU. LV enhances the activity of 5-FU by facilitating the binding of 5-FU to the enzyme thymidylate synthase, resulting in a greater degree of activity against cancer.

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Clinical Performance. Randomized data comparing 5-FU with or without LV in PC patients are lacking. However, one of the most inßuential studies in PC is the European Study Group for Pancreatic Cancer-1 (ESPAC-1) trial; although this trial did not randomize 5-FU/LV versus 5-FU alone, it did show that 5-FU/LV use led to a signiÞcant survival beneÞt when compared with no chemotherapy in surgically resected patients (Neoptolemos JP, 2004). In one Phase II trial, 42 previously untreated patients with advanced, measurable adenocarcinoma of the pancreas were treated with weekly infusional 5-FU and leucovorin IV (DeCaprio JA, 1991). Three partial responses (7%) and no complete responses were observed. Median survival was 6.2 months. The most common toxicity was diarrhea; there was one treatment-related death. In another study (Crown J, 1991), 22 eligible patients (18 previously untreated) with advanced pancreatic adenocarcinoma were treated in a Phase II trial of 5-FU/LV. Among the 20 assessable patients, no complete or partial responses (>50% reduction in tumor mass) were observed, although there was one minor response lasting four months. Three patients had stable disease for 5, 20, and 21 months, respectively. Median survival was ten weeks. Toxicity was predominantly mucosal; grade 2 or worse stomatitis was seen in Þve patients, and grade 2 or worse diarrhea was seen in four patients. Researchers concluded that this combination of 5-FU/LV did not demonstrate meaningful therapeutic activity in patients with adenocarcinoma of the pancreas and was associated with moderate to severe toxicity. Nonpharmacological Approaches In most cases of resectable PC and some cases of locally advanced disease, surgery and radiotherapy are combined with chemotherapy. Nonpharmacological approaches play a very small role in the treatment of metastatic disease, although occasionally they may be used to reduce painful symptoms. As previously mentioned, surgery is the only form of curative therapy for PC. However, less than 20% of patients are considered eligible for resection in Western centers; resection rates of 40% are normal in Japan, where surgeons routinely operate on more-advanced cases. The vast majority of patients present with cancers of the head of the pancreas. Many undergo the classic Whipple procedure, also known as pancreatoduodenectomy (PD), which involves a single-stage removal of the gallbladder, common bile duct, head of the pancreas, pylorus, and antrum of the stomach. Increasingly, a modiÞed form of the procedure, the pylorus-preserving PD, is employed. Proponents argue that it improves long-term upper gastrointestinal tract function and reduces diarrhea. Other surgical procedures performed include total pancreatectomy. Usually reserved for larger tumors of the pancreatic head and tail, it is more extensive than the classic Whipple procedure and involves the removal of the body and tail of the pancreas as well as the spleen and the surrounding lymph nodes. Distal (or left) pancreatectomy is used to excise a localized tumor in the body or tail of the pancreas—such tumors occur in approximately 20% of cases of PC.

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Palliative surgery may be undertaken when the patient is not a resection candidate. The three most common indications for palliative surgery are jaundice (resulting from biliary obstruction), duodenal obstruction, and pain control. Less invasive than palliative surgery, endoscopically placed biliary stenting is becoming the preferred mode of therapy for obstructive jaundice. The success rate for stenting is nearly 90%, although signiÞcant morbidity occurs in as many as 35% of cases. Metal stents are more likely to remain unobstructed than plastic stents and cause less morbidity. Two main types of radiotherapy procedures are used to treat PC. Multiple-Þeld external-beam radiation therapy (EBRT) is commonly used following curative resection and in patients with locally advanced PC. Intraoperative electron beam radiation therapy (IOERT) is also employed in some centers. During laparotomy, surgeons expose the pancreas or pancreatic bed to 10–30 Gy of radiation. This radiation can slow tumor growth and sterilize the tissue surrounding the pancreas. Although IOERT can produce marginal gains in survival by improving local tumor control in the locally advanced setting, it is of limited value because the majority of patients present with disease that is too advanced for radiotherapy to be effective. EMERGING THERAPIES Although pancreatic cancer (PC) is the Þfth-leading cause of cancer-related death for men and women (after lung, colon, breast, and prostate cancers), its 99% mortality rate is the highest of any cancer and signiÞcant unmet medical need remains in this indication. Few of the agents in development for the treatment of PC target this disease as their primary indication. Furthermore, few emerging therapies have demonstrated beneÞts in the management of PC; encouraging results in preliminary clinical trials are frequently not replicated in large-scale studies. Diagnosis commonly occurs late in the pathogenesis of PC, often beyond the point at which the cancer is localized and surgically resectable or responsive to systemic therapy. Consequently, PC is notoriously difÞcult to manage with pharmacotherapy, and agents with proven efÞcacy in other indications such as breast cancer or colorectal cancer (CRC) may not demonstrate sufÞcient efÞcacy in PC because of the relatively limited therapeutic window. Most of the emerging therapies reviewed in this section are being investigated in combination with standard agents—primarily gemcitabine (Eli Lilly’s Gemzar). Topoisomerase I inhibitors, taxanes, and platinum agents are increasingly used off-label in PC and may have some impact in the medium term. Immunotherapies and vascular endothelial growth factor (VEGF) inhibitors are among the most promising approaches now under investigation. Table 7 lists promising emerging therapies for PC. Immunotherapy Overview. Immunotherapeutic approaches to the treatment of cancer include vaccine and immunostimulatory therapies. The use of vaccines to stimulate

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TABLE 7. Emerging Therapies in Development for Pancreatic Cancer Compound Immunotherapies G17DT United States Europe Japan

Development Phase

Marketing Company

III III —

Aphton/Aventis Aphton/Aventis —

Virulizin United States Europe Japan

III — —

Lorus Therapeutics — —

CTP-37 United States Europe Japan

II — —

Avi BioPharma/SuperGen — —

Telomerase vaccine United States Europe Japan

— II —

— Gemvax —

Vascular endothelial growth factor inhibitors Bevacizumab United States II Europe — Japan — Platinum agents Oxaliplatin United States Europe Japan

Roche/Genentech — —

III III —

´ Sanofi-Synthelabo ´ Sanofi-Synthelabo —

III II —

Pfizer/Aventis Pfizer/Aventis

Rubitecan United States Europe Japan

PR III —

SuperGen SuperGen —

Exatecan United States Europe Japan

III III I

Daiichi Daiichi Daiichi

Topoisomerase I inhibitors Irinotecan United States Europe Japan

Epidermal growth factor receptor inhibitors Trastuzumab United States II

Roche/Genentech

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TABLE 7. (continued) Compound

Development Phase

Europe Japan Cetuximab United States Europe Japan Erlotinib United States Europe Japan Farnesyl transferase inhibitors Tipifarnib United States Europe Japan

Marketing Company

— — III II —

ImClone/Bristol-Myers Squibb Merck KGaA

III

OSI Pharmaceuticals/Genentech/ Roche

— —

III III —

Janssen Pharmaceutica Janssen Pharmaceutica

II II —

Schering-Plough Schering-Plough

III III —

Eli Lilly Eli Lilly —

Capecitabine United States Europe Japan

I/II III —

Roche Roche —

Taxanes Docetaxel United States Europe Japan

II II —

Aventis Aventis —

Gene therapy CYP2B1 United States Europe Japan

— II (Austria) —

SCH-66336 United States Europe Japan Thymidylate synthase inhibitors Pemetrexed United States Europe Japan

TNFerade United States Europe Japan

II — —

Austrianova — GenVec — —

EMERGING THERAPIES

195

TABLE 7. (continued) Compound Rexin-G United States Europe

Development Phase

Marketing Company

II —

Epeius Biotechnologies —

Note: Development phase is based on databases such as R & D Focus, R & D Insight, Pharmaprojects, and the Investigational Drugs Database (IDdb3); periodicals such as Scrip, the FDC’s Pink Sheet, and Marketletter; company reports and press releases; and industry contacts. PR = Preregistered.

therapeutic tumor-antigen-speciÞc immune responses holds promise as a complementary approach to current treatment options in PC. Important components of vaccines for active immunotherapy include immunogens in the form of tumorassociated antigens and an adjuvant or carrier molecule to promote presentation of the antigen to the immune effector cells. Possible antigens include carbohydrate structures and tumor-expressed proteins. Immunotherapy will be assured a place in the PC armamentarium if results from early studies with immunotherapeutic agents can be duplicated in randomized, Phase III studies. Because of its low toxicity, immunotherapy would likely be used in almost all PC patients in addition to standard therapy. Patients who are ineligible for gemcitabine therapy because of poor performance status will also be offered immunotherapy. Mechanism of Action. Cancer vaccines are designed to stimulate the immune system to launch a response against the speciÞc epitope contained in the vaccine. In general, research has shown that the most effective antitumor immune responses are achieved by stimulating T lymphocytes, which can recognize and kill tumor cells directly. Immunostimulants also activate other cells of the immune system, including macrophages, natural killer (NK) cells, and dendritic cells. These cells are capable of recognizing and destroying tumor cells. G17DT. G17DT (Gastrimmune), an antigastrin 17 immunogen vaccine, is under development by Aphton and Aventis. Under normal circumstances, gastrin is a potent stimulator of gastric acid secretion but, in the disease state, gastrin 17 (an intermediate) has been shown to be a growth factor for pancreatic, stomach, and colorectal cancers (Ohlsson B, 1999; Smith AM, 2000). The antigastrin 17 immunogen consists of four components: a large carrier protein molecule (diphtheria toxin, DT), a smaller peptide similar to a portion of gastrin 17 (which is targeted to be neutralized) attached to the carrier protein, an adjuvant to stimulate an immune response, and a liquid immersion vehicle. The peptide carried by DT is internalized and presented by B cells and macrophages. T cells then bind to the foreign epitopes, which in turn proliferate and signal the B cells, which bind to the peptides to proliferate and to “mass produce” the desired antibodies that neutralize gastrin-17.

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Aphton was conducting two Phase III clinical trials in patients with advanced PC in Europe and the United States comparing single-agent vaccine, vaccine plus gemcitabine therapy, or gemcitabine alone. However, trials of the combination therapy were halted in Europe in 2003 in favor of the single-agent vaccine, although the company has not disclosed the reasons. In December 2003, Aphton Þled for Australian regulatory approval of G17DT as monotherapy for the treatment of advanced pancreatic cancer (APC) in patients not receiving chemotherapy. The trial protocols had not been made available for a long time, and, at the time of composing this reference, the Phase III trials (of single agent) are still reportedly ongoing. However, recent Phase III results (of combination with gemcitabine) have been negative. Also, some discussion of the trial had been presented in a Þnancial analyst report (Simmer CM, 2003). A total of 154 treatment-naive stage II, III, and IV patients with unresectable PC were randomized to receive G17DT or placebo. The majority of the patients randomized had stage IV disease (85%). Two hundred and Þfty micrograms of G17DT were administered on day 1 of week 1 and then repeated in week 3. Intent-to-treat patient analysis revealed a median survival of 151 days for patients treated with G17DT compared with a median of 83 days for the placebo arm. The survival beneÞt at nine months was 22% in the G17DTtreated arm versus 11% in the placebo arm. No information on tolerability was presented. Additional Phase III trials investigating the efÞcacy of G17DT in combination with gemcitabine are ongoing in the United States and Europe. Two Phase II studies show that median survival in APC patients treated with G17DT compares favorably with historical data. In the Þrst trial—a dose-ranging study—30 patients were treated with either 100 µg or 250 µg by intramuscular injection in weeks 0, 2, and 6. Six of 13 patients (46%) treated with 100 µg produced antigastrin antibodies, compared with 14 of 17 patients (42%) treated with 250 µg. Median survival was 187 days (6.2 months) in the whole group, 217 days (7.1 months) in the antibody producers, and 121 days (4.0 months) in the antibody-negative group (Brett B, 2002). In the second study of G17DT monotherapy (250 µg intramuscularly, in weeks 0, 1, and 3), the outcomes of 37 APC patients were compared with those of historical controls treated with best supportive care (Gilliam AD, 2001). Median survival was 9.9 months in the vaccine group and 3.6 months for the control patients. G17DT is well tolerated. Adverse events were generally limited to injectionsite reactions, including discomfort, swelling, abscess development, and fever. Virulizin. The details of the mechanism of action of Lorus Therapeutics’ Virulizin remain uncertain. It is known that Virulizin, a mixture of proteins and peptides extracted from bovine bile, stimulates the immune system to modulate production of inßammatory cytokines, including tumor necrosis factor (TNF), interleukin 1-beta, and granulocyte-macrophage colony-stimulating factor (GMCSF). Virulizin also stimulates macrophages, and this stimulation has recently

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been shown to activate natural killer (NK) cells, increasing tumor penetration of these cells and resulting in apoptosis of the tumor cells. The agent is in Phase III trials for PC in the United States, Canada, and Mexico and was granted orphan drug status for this indication by the FDA in February 2001. The Phase III, double-blind, randomized clinical trial is designed to evaluate Virulizin as a treatment for advanced (unresectable, recurrent, or metastatic) PC. The trial was initiated in November 2001 and is being conducted at approximately 40 North American medical centers with the goal of enrolling 350 patients who will be randomized to receive gemcitabine either alone or in combination with Virulizin. Those patients who do not respond to treatment or become refractory to gemcitabine will then be treated with 5-ßuorouracil (5-FU) or with 5-FU in combination with Virulizin. At the time of composing this reference, the data have become available, and they are negative. Lorus reported a meta-analysis of three Phase I/II studies of Virulizin. In 49 evaluable patients with APC, 87% of whom had received prior treatment, median survival was 5.7 months. Six- and nine-month survival rates were 48% and 31%, respectively. In one contributing study with 19 evaluable patients, one patient experienced a complete response and seven patients had stable disease. Overall median survival duration was 6.7 months (an improvement of more than 3 months compared with average survival) and the 6-month survival rate was 58%. In addition, there was a signiÞcant improvement in total quality-of-lifechange score (Liu C, 1999). Virulizin was well tolerated in the studies included in the meta-analysis. CTP-37. Avi BioPharma (formerly AntiVirals) is developing CTP-37 (Avicine), a vaccine containing two peptides from human chorionic gonadotropin (hCG) linked to DT. CTP-37 is a peptide fragment of hCG, an oncofetal protein expressed by many cancer cells to counteract the immune system. SuperGen has acquired exclusive U.S. marketing rights to the agent. CTP-37 has completed several Phase II studies in PC and Avi BioPharma had been planning to initiate Phase III studies. However, in 2004, Avi announced that it was commissioning further Phase II studies and is seeking a development partner prior to moving into Phase III studies in PC. In a small Phase II trial, 55 patients with PC were randomized to receive either CTP-37 monotherapy or CTP-37 with gemcitabine. There was no signiÞcant difference in the overall survival times of the two groups, which were similar to those seen in patients treated with gemcitabine alone (Iversen P, 2001). Telomerase Vaccine. Gemvax is developing a telomerase vaccine, which is in Phase II clinical trials for PC in Europe. Gemvax also has a p21 Ras vaccine in Phase II trials for PC in Europe. Telomerase normally maintains the integrity of telomeres (structures that cap the ends of chromosomes) but is frequently overexpressed in cancers, promoting stabilization of telomere length and permitting continued cell proliferation. Telomerase vaccines disrupt this function, promoting telomere shortening and ultimately cell death.

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Data presented at the American Society of Clinical Oncology (ASCO) in 2003 demonstrate telomerase vaccine’s efÞcacy in treatment-naive PC. Forty-seven patients received the vaccine in combination with GM-CSF eight times over a ten-week period; select patients also received a monthly booster vaccination. Median survival was 3.5 months in the low-dose vaccine group and 9.8 months in the intermediate-dose group (Gaudernack G, 2003). Vascular Endothelial Growth Factor Inhibitors Overview. Angiogenesis—the formation of new blood vessels—is critical for both normal physiology and pathological conditions such as solid tumor growth and metastasis. A variety of endogenous angiogenic factors have been identiÞed, including vascular endothelial growth factor (VEGF), which is frequently overexpressed in PC. Both VEGF and its receptor tyrosine kinase (RTK)—the vascular endothelial growth factor receptor (VEGFR)—represent attractive therapeutic targets because inhibition of angiogenic signaling by this pathway should slow tumor growth. Angiogenesis inhibitors reduce the ability of growing solid tumors to vascularize, in turn reducing the availability of nutrients required to support fast-growing tumor tissue. Although non-VEGF-based angiogenesis inhibitors are reportedly in development for PC—including PÞzer’s histone deacetylase inhibitor CI-994 (acetyldinaline) and Merck KGaA’s αVβ3 and αVβ5 integrin inhibitor cilengitide (EMD-121974)—these agents appear unlikely to fulÞll their potential. VEGF inhibition, by contrast, is an area of considerable commercial interest. Semaxanib—a small-molecule inhibitor of RTK activity formerly in development by Sugen (a wholly owned subsidiary of PÞzer)—was among the Þrst agents in this class to be investigated for its anticancer properties. However, clinical development of semaxanib was halted in 2001, and most hope is now focused on monoclonal antibodies that inhibit VEGF binding to VEGFR. Mechanism of Action. VEGF, a multifunctional cytokine and potent permeability factor, is secreted in response to hypoxia (reduced oxygen) in tissues. VEGFR is an RTK consisting of three distinct sections: an extracellular ligandbinding domain, a transmembrane domain, and an intracellular tyrosine kinase domain. Ligand binding to the receptor generates tyrosine kinase activity and receptor autophosphorylation, which in turn activates tyrosine phosphorylation of membrane-anchored Ras. Ras signaling activates MAP kinase, which stimulates both angiogenesis and mitogenesis through activation of the transcription factors Fos and Jun supporting tumor growth. Drug intervention antagonizes receptor-mediated activation of this pathway and, in theory, inhibits angiogenesis, slowing tumor vascularization and growth. Antibodies to VEGF and VEGFR antagonize ligand binding to the receptor, preventing receptor activation. Small-molecule inhibitors of VEGFR inhibit tyrosine kinase activity, preventing autophosphorylation and subsequent activation of Ras following ligand binding.

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Bevacizumab. Bevacizumab (Genentech/Roche’s Avastin; rhuMAb-VEGF), a humanized monoclonal antibody that binds VEGF, was approved by the FDA for the treatment of metastatic CRC in combination with 5-ßuorouracil (5-FU) in 2004. Bevacizumab is in Phase II trials in combination with gemcitabine and Phase I trials in combination with capecitabine and radiation for APC. Published data supporting the efÞcacy of bevacizumab for the treatment of PC remain scarce. Interim data from an ongoing Phase II trial involving 40 patients with APC (presented at the ASCO’s First Annual Symposium on Gastrointestinal Cancers in 2004) indicate that the addition of bevacizumab to gemcitabine therapy improves both median time to progression and six-month survival rates relative to gemcitabine monotherapy. In this trial, intravenous gemcitabine (1,000 mg/m2 ) was administered on days 1, 8, and 15 and subsequently every 28 days. Intravenous bevacizumab (10 mg/kg) was given on days 1 and 15 and subsequently every 28 days. At the time of presentation, only 32 patients were available for evaluation (8 patients were still too early in the protocol); a median of three cycles of treatment had been completed. There were eight conÞrmed partial responses, lasting a median of 9.2 months, and 13 patients with stable disease, lasting a median of 5.8 months. Six-month survival was 74% and median survival was 12.4 months. Side effects were mostly consistent with gemcitabine use and included leukopenia (28%), neutropenia (25%), anemia (3%), and thrombocytopenia (5%). A single (fatal) incidence of gastrointestinal bleeding was reported, but this adverse reaction occurred in a patient with metastatic invasion of the duodenum (Kindler HL, 2004). A Phase III trial investigating bevacizumab in combination with gemcitabine and several Phase II trials investigating bevacizumab combination therapy are now in progress. Platinum Agents Overview. Cisplatin—the Þrst platinum compound to be used for the treatment of cancer—entered clinical trials in the 1970s and was found to have activity against numerous cancers, including testicular, bladder, and ovarian cancer. Second-generation agents (such as carboplatin) lack some of the renal and neurotoxicities associated with cisplatin but retain much of that agent’s efÞcacy against solid tumors. Oxaliplatin (SanoÞ-Synth´elabo’s Eloxatin), a third-generation agent with an improved toxicity proÞle, is displaying activity in a number of cancer indications. Like many other cytotoxic agents, platinum compounds are associated with cellular resistance by mechanisms, including decreased uptake of platinum-containing compounds by resistant cells, inactivation of the drug by cellular thiol compounds, and enhanced repair of platinum-related DNA damage. A number of platinum-containing compounds are being studied in combination with gemcitabine for the management of PC (Philip A, 2001). The most clinically advanced for PC treatment is oxaliplatin. Mechanism of Action. Platinum-containing compounds (e.g., alkylating agents) react with many biologic compounds. However, their cytotoxic effect

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FIGURE 4. Structure of oxaliplatin.

is believed to be mediated by their interference with DNA replication and cell division. Oxaliplatin. SanoÞ-Synth´elabo’s oxaliplatin (Figure 4), a third-generation platinum complex, is marketed in Europe and the United States for the treatment of CRC. This agent is in Phase III trials for PC—alone and in combination with gemcitabine or 5-FU. At the time of composing this reference, these international trials are still open to patient recruitment. The Þnal results of a European randomized Phase III trial of gemcitabine plus oxaliplatin versus gemcitabine alone were presented at ASCO 2004 (Louvet C, 2004). A total of 326 patients with APC were randomized to receive either arm “A” or arm “B.” Patients randomized to arm A received gemcitabine 1,000 mg/m2 weekly for seven weeks with one week of rest followed by a further four courses of gemcitabine every three weeks. Patients randomized to arm B received gemcitabine 1,000 mg/m2 on day 1 followed by oxaliplatin 100 mg/m2 (GEMOX) on day 2 every two weeks. The primary end point of the study was an increase in median survival from six months to eight months; this end point was not met. However, 28.7% of the patients treated with GEMOX had a partial response with a median duration of 42 weeks. This result was signiÞcantly different from the 16.7% partial responses achieved in the gemcitabine arm. Progression-free survival was signiÞcantly better in the GEMOX-treated group at 5.5 months versus 3.7 months for the gemcitabine-treated group. Toxicity data were presented at ASCO 2003 (Louvet C, 2003). GEMOX and gemcitabine treatment regimens had a comparable toxicity proÞle with similar grade 3/4 events occurring at similar frequencies. However, thrombocytopenia (12.8% versus 3.8%), vomiting (8.9% versus 3.2%), and neuropathy (15.3% versus 0.0%) occurred more frequently in the GEMOX arm. In a randomized, open-label, Phase II study, 63 patients with APC were randomized to receive oxaliplatin alone (130 mg/m2 , two-hour, intravenous infusion), 5-FU alone (1,000 mg/m2 /day, continuous intravenous infusion on days 1 to 4), or oxaliplatin plus 5-FU every three weeks (Ducreux M, 2004). Three partial responses occurred, all in the oxaliplatin/5-FU arm (10% response rate). Median time to progression and overall survival time were greater in the combination arm (4.2 and 9.0 months, respectively, versus 2.0 and 3.4 months for the oxaliplatin arm and 1.5 and 2.4 months for the 5-FU arm). The authors concluded that the response rate and encouraging safety proÞle warrant further studies of this combination in PC.

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In another Phase II study, 47 patients with previously untreated APC received oxaliplatin (100 mg/m2 intravenous on day 1) and gemcitabine (1,000 mg/m2 intravenous on days 1 and 8) for a three-week cycle (Alberts SR, 2003). More than 50% of patients lived for longer than six months (the trial’s primary end point); the median time to progression was 4.53 months and Þve conÞrmed responses were seen, with a median duration of response of 2.7 months. The treatment was found to be well tolerated. Topoisomerase I Inhibitors Overview. Cytotoxic agents targeting topoisomerase I (an enzyme that cleaves and reattaches DNA during cell replication) are commonly used to treat cancer. Camptothecin (CPT, generics; an alkyloid agent derived from the Campotheca acuminate tree) was the Þrst topoisomerase I inhibitor to reach clinical trials for the management of cancer. However, unacceptable toxicity, including severe myelosuppression, enteritis, and hemorrhagic cystitis, led to the discontinuation of initial trials. Subsequently, two safer CPT analogues—topotecan (GlaxoSmithKline’s Hycamtin) and irinotecan (PÞzer’s Camptosar, Yakult/Aventis’s Campto, Daiichi’s Topotecin)—received approval for broad clinical use. Although these agents are not currently approved for the management of PC, off-label use in this indication is relatively common. Mechanism of Action. Topoisomerase I catalyzes the conversion of DNA strands into single-strand DNA breaks—a process that is necessary to relieve chain tension during the DNA replication required for cell division—by nicking and then re-annealing the DNA strand. CPT analogues exert their cytotoxic effect by binding to topoisomerase I and stabilizing the intermediate enzyme-DNA cleavable complex, generating DNA strand breaks that are lethal to the cell. Irinotecan. Irinotecan (PÞzer’s Camptosar, Yakult/Aventis’s Campto, Daiichi’s Topotecin) (Figure 5) is a parenteral topoisomerase I inhibitor approved for the treatment of advanced metastatic CRC. Although irinotecan has demonstrated only modest activity in PC as single-agent therapy, clinical trials in paired and multiple combinations with gemcitabine (Eli Lilly’s Gemzar), 5-FU (generics) and cisplatin (Bristol-Myers Squibb’s Platinol AQ, generics) are underway. However, because gemcitabine is already approved for the treatment of APC, it is likely that the combination of irinotecan with gemcitabine will prove more successful than the combination of irinotecan with other agents; the following paragraphs therefore report the results of such studies. A Phase III, randomized, multi-center trial in the United States compared the combination of gemcitabine (1,000 mg/m2 ) followed by irinotecan (100 mg/m2 ) on days 1 and 8 of a three-week cycle with gemcitabine alone (1,000 mg/m2 ) administered for seven consecutive weeks on days 1 and 8 of cycle one and days 1, 8, and 15 for all subsequent cycles (Rocha L, 2004). One hundred and eighty patents with APC were randomized to each treatment arm. Extrapolation of the trial results suggested a longer time to progression in patients with locally

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H3C O

O

O N

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O

FIGURE 5. Structure of irinotecan.

advanced disease of 7.7 versus 3.9 months in favor of the combination therapy against gemcitabine alone. Adverse events occurred in both treatment arms and included grade 3–4 neutropenia, diarrhea, vomiting, and thromboembolic events. The occurrence of all side effects was more common in the gemcitabine/irinotecan treatment group. In a Phase II trial of irinotecan in combination with gemcitabine, 60 APC patients—85% of whom were stage IV—were treated with gemcitabine (1,000 mg/m2 on days 1 and 8) plus irinotecan (300 mg/m2 on day 8) for cycles of three weeks. Intention-to-treat analysis identiÞed 1 (1.7%) complete response and 14 (23.3%) partial responses. (Objective response rate 24.7%. Stable disease was achieved in 22 patients [36.7%]). Twenty-three patients (38.3%) had progressive disease. Median duration of response was Þve months; median time to tumor progression was seven months and median overall survival was seven months; one-year survival was 22.5%. Pain improvement was observed in 45% of patients, and improvement in asthenia (weakness) was observed in 43% of patients. Side effects included anemia (grade 2 in 16.7% of patients; grade 3 in 5% of patients); thrombocytopenia (grade 3 in 11.7% of patients); and neutropenia (grades 3 and 4: 45%; two patients died from sepsis). The author’s note that (with a 25% objective response rate) the combination of gemcitabine and irinotecan is active in PC and that toxicity was acceptable for the majority of patients (Stathopoulos GP, 2003). Rubitecan. In March 2004, SuperGen Þled Orathecin (rubitecan), its orally active topoisomerase I inhibitor, for approval in the United States for the management of APC in patients who failed to respond to treatment with at least one prior line of chemotherapy. Filing was based on consolidated data from Phase I, II, and III trials. In total, these trials included more than 1,000 patients who had failed previous chemotherapy; of this population, 600 patients received rubitecan and the remainder received control therapies. Additional Phase III studies are still ongoing in the United States and Europe. These investigations include a trial comparing the efÞcacy of rubitecan with that of gemcitabine in chemotherapy-n¨aive patients with APC. Another Phase III trial

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is comparing the response rates achieved by rubitecan with those of 5-FU in patients who have progressed following treatment with gemcitabine. In the Phase III study included in the FDA Þlling, 409 APC patients who had failed previous chemotherapy were randomized to receive rubitecan or “best choice” alternatives: typically gemcitabine, 5-FU, mitomycin C, capecitabine, or docetaxel (Jacobs AD, 2004; Burris H, 2003). Thirteen of the 196 patients (7%) randomized to the rubitecan arm experienced partial or complete tumor response (tumor shrinkage in excess of 50%) in comparison to one (0.5%) of 211 patients in the control arm. Additionally, median time to progression was signiÞcantly greater in the rubitecan arm (57 days versus 47 days) as was median survival time (109 days versus 94 days) although this result was not statistically significant. Among the patients in the rubitecan arm who experienced complete or partial response, median time to disease progression was 246 days and median survival time was 336 days. Patients in the control arm where permitted to cross over to the rubitecan arm upon treatment failure. One hundred and two patients in the original control arm received this “rescue therapy,” and 109 patients in the original control arm received no rescue therapy. A signiÞcant increase in survival from randomization was achieved by rescue therapy: 137 days versus 59 days, respectively. Less than 5% of patients in either arm discontinued therapy because of adverse events. Side-effect occurrence rates were similar in the rubitecan and “best choice” arms and included asthenia (19% versus 16%), abdominal pain (15% versus 8%), sepsis (5% versus 7%), nausea (12% versus 8%), vomiting (11% versus 5%), leukopenia (19% versus 10%), and anemia (15% versus 7%). Exatecan. Daiichi’s exatecan is a water-soluble synthetic derivative of camptothecin that is undergoing Phase III clinical development in the United States and Europe and Phase I trials in Japan for metastatic PC. The agent is also under investigation for the treatment of advanced ovarian, peritoneal, and non-smallcell lung cancer. In preclinical studies, exatecan proved to be a potent inhibitor of topoisomerase I and tumor growth. At the 2004 ASCO meeting, negative data from two Phase III trials were disclosed. In one study, 349 APC patients were randomized to receive exatecan and gemcitabine or gemcitabine alone (O’Reilly EM, 2004). Median survival time was 6.7 months for the combination arm and 6.2 months for gemcitabine alone. The median time to progression was 3.7 months for the combination arm and 3.8 months for gemcitabine alone. Tumor response rates were 8.2% for the combination and 6.3% for gemcitabine alone. Grade 3–4 toxicity was higher in the combination arm than in the monotherapy arm: neutropenia (30% versus 15%), thrombocytopenia (17% versus 4%), and vomiting (11% versus 5%). The other study randomized 330 APC patients to receive exatecan or gemcitabine (Cheverton P, 2004). The primary endpoint—to detect a 50% improvement in overall survival in the patients receiving exatecan over gemcitabine—was not attained.

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Epidermal Growth Factor Receptor Inhibitors Overview. The epidermal growth factor receptor (EGFR [HER-1]) receptor tyrosine kinase (RTK) normally regulates cellular differentiation and proliferation. PC is associated with consistent upregulated expression of EGFR and its ligands, epidermal growth factor (EGF), and transforming growth factor-alpha (TGF-α). Overexpression of EGFR and its ligands confer a selective growth advantage, which makes EGFR an attractive therapeutic target (Smith JJ, 1987). Mechanism of Action. EGFR is part of the ERBB family of receptors, which consist of an extracellular ligand-binding domain, a transmembrane domain, and an intracellular tyrosine kinase domain. Ligand binding to the receptor generates tyrosine kinase activity and receptor autophosphorylation, which in turn activates tyrosine phosphorylation of the membrane-anchored GTP-binding protein Ras. Ras signaling activates mitogen-activated protein (MAP) kinase, which stimulates the promitogenic transcription factors Fos and Jun, promoting cell proliferation. EGFR activity is also thought to inhibit apoptosis by repression of the nuclear factor-kappa B (nfκB) signaling pathway (Sclabas GM, 2003). Activating mutations of EGF or EGFR stimulate uncontrolled cell proliferation by constitutive activation of Ras and MAP-kinase. The precise mechanism of action of EGFR inhibitors is unknown and complicated by the fact that EGFR expression does not necessarily correlate with anticancer activity. However, researchers believe that drug intervention, either at the EGFR or the intracellular tyrosine kinase, antagonizes receptor-mediated activation of this pathway, inhibiting cell proliferation and slowing tumor growth. Antibodies to EGF or EGFR antagonize ligand binding to the receptor, preventing receptor activation. Small-molecule inhibitors of EGFR inhibit tyrosine kinase activity, preventing autophosphorylation and subsequent activation of Ras following ligand binding. Trastuzumab. Trastuzumab (Genentech/Roche’s Herceptin) is a humanized monoclonal antibody approved for the treatment of HER-2 -positive breast cancer. Trastuzumab binds to the p185HER2 tumor antigen (which is serologically related to EGFR), inhibiting mitogenic stimuli and promoting apoptosis. Although HER2 was believed to be ampliÞed or overexpressed in 20–40% of PC cases, recent data suggest that its ampliÞcation may, in fact, be substantially less common (Potti A, 2003). Trastuzumab is in Phase II development for the treatment of PC in combination with gemcitabine, although data from these trials remain relatively scarce. The results of a small Phase II trial in patients with metastatic PC were reported at ASCO’s annual meeting in 2001 (Safran H, 2001). Twenty-one patients with HER-2 overexpression received gemcitabine (1,000 mg/m2 /week) and trastuzumab (a loading dose of 4 mg/kg, then 2 mg/kg/week). Of 18 evaluable patients, 4 (22%) had partial responses and 9 (50%) had more than 50% reduction in the carbohydrate antigen CA 19-9. Grade 3 and 4 toxicities included neutropenia (n = 3), thrombocytopenia (n = 1), and elevated transaminase (n = 1). Researchers observed a grade 3 decline in left-ventricular ejection fraction in one patient.

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Data from a Phase II trial of trastuzumab in combination gemcitabine and cisplatin in non-small-cell lung cancer suggest that trastuzumab combinations may not be efÞcacious in all cancer settings. In this study, treatment with trastuzumab combined with gemcitabine and cisplatin conferred no clinical beneÞt in patients with HER-2 -positive non-small-cell lung cancer (Gatzemeier U, 2004). Cetuximab. Developed by ImClone Systems and Bristol-Myers Squibb in the United States and Merck KGaA in Europe, cetuximab (Erbitux) is a chimeric monoclonal antibody that binds to the external portion of EGFR, inhibiting ligand-mediated mitogenic stimulation. It is approved for the treatment of irinotecan-refractory CRC in Europe and the United States. The agent is in Phase III clinical trials in the United States and Phase II in Europe for PC. It is also being investigated for a range of other neoplasms, including head and neck cancer and non-small-cell lung cancer. A Phase II study involving 41 patients with HER-1 -positive pancreatic tumors evaluated the use of cetuximab at a loading dose of 400 mg/m2 , followed by a weekly dose of 250 mg/m2 , and standard gemcitabine therapy (Abbruzzese JL, 2001). Five patients (12.5%) achieved a partial response (>50% tumor regression), and an additional 21 patients (52.5%) achieved disease stabilization. Median time to disease progression was approximately 3.5 months. Overall median survival was 6.75 months, with a one-year overall survival rate of 32.5%. The most common adverse events reported were nausea, fatigue, and an acnelike rash. Toxicities associated with the use of gemcitabine did not appear to be exacerbated by the addition of cetuximab. A Phase III, multicenter, randomized, open-label trial comparing gemcitabine alone with gemcitabine plus cetuximab in patients with locally advanced, unresectable, or metastatic disease started patient recruitment in December 2003 (www.cancer.gov). The objectives of the trial include a comparison of overall survival, time to treatment failure, toxicity, total response rate, and quality of life. Investigators will estimate tumor EGFR expression and establish a correlation to overall survival. The South West Oncology Group (SWOG) intends to recruit 704 patients from multiple centers in the United States and randomize them to receive either gemcitabine alone given on days 1, 8, 15, and 22 of course one and days 1, 8, and 15 of all subsequent courses, or gemcitabine plus cetuximab on days 1, 8, 15, and 22 of each course. Courses are given every four weeks and will be continued until disease progression. Patients will be followed for a total of three years. The results of this Phase III trial will be a pivotal factor in the approval of cetuximab for APC in the future. Erlotinib. Erlotinib (Tarceva), a small-molecule EGFR tyrosine kinase inhibitor, is under development by OSI Pharmaceuticals in alliance with Genentech and Roche. Erlotinib is in Phase III clinical trials for APC, alone and in combination with gemcitabine; Phase III studies in combination with gemcitabine were initiated in 450 APC patients at the beginning of 2003. The primary endpoint is a 33% improvement in patient survival, but OSI recently announced that erlotinib is unlikely to meet this goal.

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The NCI initiated a Phase I study of erlotinib with gemcitabine and radiotherapy in patients with APC in July 2003. This nonrandomized, open-label, single-center, dose-escalation study will determine the maximum tolerated dose of erlotinib when given in combination with chemotherapy and radiotherapy. Preliminary efÞcacy data will be generated from the 19–28 patients enrolled in this study. Farnesyl Transferase Inhibitors Overview. The proto-oncogene Ras, which encodes a small GTP-binding protein, is required for both normal intracellular signaling via RTKs (such as EGFR and VEGFR) and appropriate control of cell division. Mutated, oncogenic Ras occurs in up to 90% of PC cases, and substantial research has been invested in identifying inhibitors of Ras function. Farnesyl transferase inhibitors (FTIs) have emerged as potential therapies for the treatment of PC and other neoplastic diseases because they catalyze a critical posttranslational modiÞcation step required for Ras activity. In preclinical models, unmodiÞed Ras molecules are unable to stimulate MAP kinase, inhibiting cell division (McGeady P, 1995). Several companies, including Schering-Plough, Janssen Pharmaceutica, and Bristol-Myers Squibb, are developing FTIs. Mechanism of Action. Ras normally transduces intracellular signals from RTKs; Ras signaling activates MAP kinase, thereby stimulating the promitogenic transcription factors Fos and Jun and promoting cell proliferation. Mutations of Ras stimulate uncontrolled cell proliferation by constitutive activation of MAP kinase. Ras function requires the addition of a farnesyl (lipid) group to a cysteine residue at the carboxy-terminus of the protein. This posttranslational modiÞcation is catalyzed by farnesyl transferase; small-molecule inhibitors of this enzyme antagonize the modiÞcation of Ras, which in turn generates a n¨aive form of Ras that is incapable of stimulating MAP-kinase-associated cell division. Tipifarnib. Tipifarnib (formerly known as R-115777; trade name Zarnestra) is a selective nonpeptideomimetic FTI in development by Janssen Pharmaceutica (a subsidiary of Johnson & Johnson). The agent is in Phase III trials in the United States and Europe for the treatment of metastatic PC; it is also under investigation for the treatment of other carcinomas, including breast cancer. Data from a large-scale, randomized Phase III trial of tipifarnib in combination with gemcitabine in previously untreated APC patients indicate no signiÞcant improvement in efÞcacy relative to gemcitabine plus placebo (Van Cutsem E, 2004). In this trial, 688 previously untreated patients with APC received either weekly gemcitabine (1,000 mg/m2 for seven weeks) followed by one week of rest and a further three weeks of therapy plus placebo or weekly gemcitabine plus tipifarnib (200 mg twice a day) for the duration of the trial. Treatment was given for a median of 85 and 98 days, respectively. No statistically signiÞcant differences were observed in the primary endpoint—median survival rate—between treatment groups receiving tipifarnib and gemcitabine and those receiving gemcitabine plus placebo (193 versus 182 days, respectively).

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SCH-66336. Schering-Plough’s SCH-66336 is an orally active FTI in Phase II development in combination with gemcitabine. In a randomized Phase II trial, 63 metastatic PC patients received either gemcitabine or SCH-66336 (200 mg twice a day) (Lersch C, 2001). The three-month, progression-free survival rate and length of median survival were greater in the gemcitabine group (31% and 4.4 months) than in the SCH-66336 group (23% and 3.3 months). However, SCH66336 was better tolerated than gemcitabine. Although the incidence of nausea, vomiting, and diarrhea was equivalent in the two groups, these side effects were more severe in the gemcitabine group. SCH-6336 patients also experienced less hematological toxicity. In a second trial, 25 evaluable patients with solid tumors received SCH-66336 in combination with gemcitabine. Two achieved partial responses, two had a minor response, and 11 had stable disease. One patient with PC maintained stable disease for more than 16 months. Preliminary data suggested there was no interaction between gemcitabine and SCH-66336 (Hurwitz HI, 2000). Thymidylate Synthase Inhibitors Overview. Inhibition of DNA synthesis is a common mechanism of action for cytotoxic agents. Folic acid antagonists (antifolates)—such as methotrexate (an established agent in chemotherapy regimens used to treat numerous neoplasms)—act by inhibiting the function of dihydrofolate reductase (DHFR), a key enzyme in the thymidylate cycle that is required for the production of the nucleotide thymidine. However, the use of methotrexate is associated with cellular resistance because tumors acquire mutations that allow escape from the cytotoxic effects of methotrexate. Many emerging antifolates are analogues of folates or methotrexate that have been rationally designed to overcome the cellular resistance associated with methotrexate. Some emerging folate antagonists inhibit the function of thymidylate synthase (TS), another component of the thymidylate cycle, which catalyzes the generation of thymidylate from deoxyurydilate. Mechanism of Action. TS inhibitors inhibit the conversion of deoxyuridylate to thymidylate; in the absence of de novo synthesis of thymidine, cells are unable to replicate their DNA prior to cell division and proliferation. Pemetrexed. Pemetrexed (Eli Lilly’s Alimta) (Figure 6) is a multitargeted antifolate (i.e., thymidylate synthase and dihydrofolate reductase inhibitor) undergoing Phase III clinical studies in the United States and Europe for the treatment of APC. The agent exerts its antitumor activity by inhibiting multiple components of the thymidylate cycle: TS, dihydrofolate reductase, and glycinamide ribonucleotide (GAR) formyltransferase. Pemetrexed’s multitargeted action may overcome the resistance acquired through the overexpression of any single enzyme. The compound is approved in the United States–in combination with cisplatin—for the treatment of patients with malignant pleural mesothelemia who cannot undergo surgery; Lilly has also Þled for pemetrexed’s approval as secondline therapy for NSCLC in Europe and metastatic NSCLC in United States.

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FIGURE 6. Structure of pemetrexed.

O CH3 HN

O F

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FIGURE 7. Structure of capecitabine.

Negative Phase III data were disclosed at the ASCO 2004 meeting (Richards DA, 2004). The trial involved 565 patients with APC randomized to receive gemcitabine alone or gemcitabine and pemetrexed (GP). No statistical difference in overall survival (median 6.2 months for GP and 6.3 months for gemcitabine) was evident, although response rate was signiÞcantly better for GP (18.3%) than for gemcitabine monotherapy (9.1%), as was median time to progression (5.2 months for GP versus 3.6 months for gemcitabine). Progression-free survival (3.9 months versus 3.3 months) and one-year survival (21.4% versus 20.1%) were similar in both arms. More grade 3/4 toxicity was seen among PG-treated patients: neutropenia (45.1% versus 12.8%), thrombocytopenia (17.9% versus 6.2%), anemia (13.9% versus 2.9%), febrile neutropenia (9.9% versus 0.4%), and fatigue (15% versus 6.6%). Capecitabine. Capecitabine (Roche’s Xeloda) (Figure 7) is a ßuoropyrimidine. Fluoropyrimidines, which are structural analogues of naturally occurring metabolic products (generally folates, purines, and pyrimidines) needed for the synthesis of nucleic acids, act by inhibiting TS, a key enzyme in DNA synthesis. Capecitabine is marketed for the treatment of metastatic CRC and metastatic breast cancer in the United States and Europe. Phase III trials of capecitabine in combination with gemcitabine are recruiting patients with APC in Europe. Capecitabine is in Phase I/II trials for PC in the United States. A Phase II multicenter European study investigated the efÞcacy of the capecitabine/gemcitabine combination in patients with APC (Stathopoulos G,

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FIGURE 8. Structure of docetaxel.

2004). A total of 53 therapy-n¨aive patients were treated with gemcitabine (1,000 mg/m2 ) on days 1 and 8 and capecitabine (1,300 mg/m2 ) as total dose-perday on days 1 through 14 of a 21-day cycle. An objective partial response was achieved in 10 (18.9%) patients. Stable disease was reported in 22 (42%) patients, and the remaining 15 (28%) patients had progressive disease. The median duration of response was three months, and the median time to progression was 6.5 months. The median overall survival was eight months. One-year survival was reported as 34.8%. Side effects at grade 3 or 4 toxicity levels were recorded for anemia in 9%, thrombocytopenia in 6%, neutropenia in 34%, and febrile neutropenia in 4% of patients. Investigators concluded that the combination of capecitabine and gemcitabine is a well-tolerated regimen that requires further investigation in randomized Phase III trials. Preliminary data were disclosed at the 2004 ASCO meeting (Heinemann V, 2004). The trial randomized 135 patients to receive capecitabine/oxaliplatin, capecitabine/gemcitabine, or gemcitabine/oxaliplatin. EfÞcacy data were not available, but all three regimens were equally well tolerated. Taxanes Overview. Taxanes are potent antitumor agents; members of this class—such as paclitaxel (Bristol-Myers Squibb’s Taxol, generics)—are marketed for numerous forms of cancer, including breast cancer. As single agents and in combination with other chemotherapeutic agents, taxanes have shown activity against a range of solid tumors but have demonstrated variable efÞcacy against PC. Phase II studies have failed to conÞrm the high response rates observed in early studies. Mechanism of Action. Taxanes promote microtubule polymerization and inhibit tubulin depolymerization, arresting mitotic cell division at the metaphase/anaphase transition and inducing cell death. Docetaxel. Docetaxel (Aventis’s Taxotere) (Figure 8) has shown activity against PC. In a Phase II study in 40 patients with unresectable locally advanced or metastatic PC, 6 patients had a partial response and 15 patients had stable disease. The median duration of response was 5.1 months (Rougier P, 2000). Interest has developed in employing docetaxel in combination regimens with gemcitabine.

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In another Phase II combination trial, 40 patients with APC were enrolled to receive gemcitabine (750 mg/m2 ) and docetaxel (35 mg/m2 ) weekly for three out of four weeks for a median of four cycles. An objective response was obtained in 10 (27%) patients with a median duration of 17 weeks. Median survival was seven months and one-year survival was 19.3%. The authors conclude that, although the gemcitabine-docetaxel regimen was well tolerated, its ultimate role in the treatment of PC can only be determined by a randomized Phase III trial (Schneider BP, 2002).

Gene Therapy Overview. The withdrawal from clinical development of Onyx Pharmaceuticals’ Onyx-015—an attenuated adenovirus that preferentially replicates in p53 -deÞcient cells—suggests that gene therapy may not be a viable approach to the treatment of PC, especially given the disease’s complex biological nature. The gene therapies still in development are at relatively early stages. Mechanism of Action. The primary approach of gene therapy is to either replace or activate aberrant tumor-suppressor genes or to inactivate oncogenes by insertion of a gene or antisense DNA using a vector. Alternatively, some approaches deliver genes that confer cytotoxic function to the tumor site. Delivery mechanisms include attenuated viruses (e.g., adenoviruses, selected retroviruses) or encapsulated cell approaches. CYP2B1. Austrianova is developing CYP2B1, an encapsulated cell therapy/ifosfamide combination approach for the treatment of PC. It is undergoing Phase II trials and was granted orphan drug status in the European Union in 2003. CYP2B1 therapy requires angiographic implantation of encapsulated cells modiÞed to express cytochrome P-450 2B1 (CYP-B1). Found naturally in the liver, cytochrome P-450 is a family of enzymes that metabolize the pro-drug ifosfamide into an inactive intermediary compound, which subsequently spontaneously converts into two cytotoxic compounds. Both cytotoxic compounds have short half-lives, limiting the effectiveness of ifosfamide treatment. During therapy, genetically modiÞed cells are injected into an artery feeding the primary tumor. (The encapsulation procedure prevents attack by the host’s immune system.) Low doses of ifosfamide are injected into the patient, and the ifosfamide is locally activated by the genetically modiÞed cells, producing a high concentration of active ifosfamide metabolites at the tumor site. Data from a single-arm Phase I/II trial in 14 patients with inoperable PC indicate the potential of this approach. Ten patients showed stable disease, and median survival of treated patients was twice that of those treated with control therapy. (Salmons B, 2003). One-year survival in the treated group was 36% (twice that of the control group), and four of 13 evaluable patients reported improvements in pain assessment.

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TNFerade. TNFerade, a tumor necrosis factor-alpha (TNF-α)-based gene therapy in development by GenVec, is undergoing Phase II trials for the treatment of PC. TNFerade is a replication incompetent adenovirus vector carrying TNF-α under the transcriptional control of the radiation-sensitive early growth response (EGR)-1 gene promoter. The combination of spatially and temporally restricted control of TNF-α expression and synergistic action with radiation therapy is expected to increase the efÞcacy of this agent and minimize the side effects associated with systemic administration of TNF-α. Data from Phase II trials were presented at the American Society of Clinical Oncology’s First Annual Symposium on Gastrointestinal Cancers in 2004. ThirtyÞve patients with unresectable PC were treated with TNFerade in combination with radiotherapy; after three months, 26 of 35 patients showed stable disease and four of these patients demonstrated tumor regression in excess of 50%. As a result, four patients were able to undergo surgical resections. Transient hypotension, the only therapy-speciÞc toxicity, occurred in one patient. TNFerade was efÞcacious at all doses used and the maximum tolerated dose was not reached (Hanner N, 2004). Rexin-G. Rexin-G, a gene-therapy-based approach targeting cyclin-G1, is in Phase II development in the United States by Epeius Biotechnologies. Rexin-G is an injectable retroviral vector encoding a mutant form of the cell-cycle-control molecule cyclin-G1, which normally regulates the passage of cells from G1 phase to S phase during the cell cycle. Cell-cycle arrest at this phase causes cells to die. Phase II data from three APC patients treated with Rexin-G indicate some efÞcacy. Patients were treated with one of two protocols. In the Þrst protocol, intravenous Rexin-G was administered for eight to ten days followed by a oneweek toxicity evaluation period. Subsequently, the maximum tolerated dose of Rexin-G was administered for eight to ten days. In the second protocol, intravenous Rexin-G was administered for six days followed by eight doses of weekly gemcitabine. All three patients experienced arrested tumor growth without experiencing dose-limiting toxicity. No signiÞcant side effects were reported. Two patients were reported to be alive with stable disease approximately Þve and 14 months from diagnosis. One patient remains alive with progressive disease 20 months from diagnosis (Gordon EM, 2004). REFERENCES Abbruzzese JL, et al. Phase II study of anti-epidermal growth factor receptor (-EGFR) antibody cetuximab (IMC-C225) in combination with gemcitabine in patients with advanced pancreatic cancer. Proceedings of the American Society of Clinical Oncology. 2001. Abstract 518. Ahlgren JD. Epidemiology and risk factors in pancreatic cancer. Seminars in Oncology. 1996;23:241–250. Alberts SR, et al. Gemcitabine and oxaliplatin for metastatic pancreatic adenocarcinoma: a North Central Cancer Treatment Group Phase II Study. Annals of Oncology. 2003;14: 580–585.

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Silverman DT, et al. Diabetes mellitus, other medical conditions and familial history of cancer as risk factors for pancreatic cancer. British Journal of Cancer. 1999;11: 1830–1837. Simmer CM. Miller Johnson Steichen Kinnard Publications. Investment securities: research update. Aphton Corporation. October 31, 2003. Simon B, Printz H. Epidemiological trends in pancreatic neoplasias. Digestive Diseases. 2001;19(1):6–14. Sinicrope FA, et al. Bcl-2 and p53 expression in resectable pancreatic adenocarcinomas: association with clinical outcome. Clinical Cancer Research. 1996;2(12):2015–2022. Smith AM, Watson SA. Review article: gastrin and colorectal cancer. Alimentary Pharmacology & Therapeutics. 2000;14(10):1231–1247. Smith JJ, et al. Production of transforming growth factor-alpha in human pancreatic cancer cell: evidence for super agonist autocrine cycle. Cell Biology. 1987;84:7567–7570. Solomon L. The descriptive and analytical epidemiology of nine cancers. Cambridge University. 2002;1–349. Stat bite: Pancreatic cancer incidence in U.S. blacks and whites, 1973–1999. Journal of the National Cancer Institute. 2002;94:1671. Stathopoulos GP, et al. Treatment of pancreatic cancer with a combination of irinotecan (CPT-11) and gemcitabine: a multi-center Phase II study by the Greek Cooperative Group for Pancreatic Cancer. Annals of Oncology. 2003;14:388–394. Surveillance, Epidemiology, and End Results (SEER). http://training.seer.cancer.gov/ Accessed April module staging cancer/unit03 sec01 part00 sum staging.html. 14, 2004. Swerdlow AJ, et al., eds. Cancer Incidence and Mortality in England and Wales: Trends and Risk Factors. Oxford, England: Oxford University Press; 2001. Tempero M, et al. Randomized Phase II comparison of dose-intense gemcitabine: 30minute infusion and Þxed-dose-rate infusion in patients with pancreatic adenocarcinoma. Journal of Clinical Oncology. 2003;21(18):3402–3408. Thayer SP, et al. Hedgehog is an early and late mediator of pancreatic cancer tumorigenesis. Nature. 2003;425:851–856. Van Cutsen E, et al. Phase III trial of gemcitabine plus tipifarnib compared with gemcitabine plus placebo in advanced pancreatic cancer. Journal of Clinical Oncology. 2004;22(8):1430–1438. Wayne JD, et al. Localized adenocarcinoma of the pancreas: the rationale for preoperative chemoradiation. Oncologist. 2002;7(1):34–45. Wicking C, et al. The hedgehog signaling pathway in tumorigenesis and development. Oncogene. 1999;18(55):7844–7851. World Health Organization. International statistical classiÞcation of diseases and related health problems, tenth revision (ICD-10). Geneva, Switzerland: WHO, 1992. Yanagisawa A, et al. Frequent c-K-ras oncogene activation in mucous cell hyperplasia of pancreas suffering from chronic inßammation. Cancer Research. 1993;53:953–956. Young JL Jr, et al., eds. SEER Summary Staging Manual—2000: Codes and Coding Instructions, National Cancer Institute, Bethesda, MD: NIH Pub. No. 01-4969; 2001. Zalatnai A. Pancreatic cancer—a continuing challenge in oncology. Pathology Oncology Research. 2003;9(4):252–263.

Prostate Cancer

ETIOLOGY AND PATHOPHYSIOLOGY Introduction Typically, tumors of the prostate grow slowly. The vast majority arise in the subcapsular peripheral region of the posterior lobe—a region of the gland that is most sensitive to changes in androgen levels. Tumors generally originate as small, well-differentiated lesions and double in size every two to four years. As a tumor grows, the cells within it become more diverse, and the outlines of the lesion become less visible. By the time the tumor reaches 4–5 cm along its longest axis, the cancer is likely to have spread to other areas of the body and can usually be seen (typically during surgery) extending beyond the prostate gland. The cancer spreads, both locally and to distant sites, over a period of eight to ten years following the appearance of the primary tumor. Local spread typically involves the seminal vesicles, ejaculatory ducts, and pelvic lymph nodes. Undetected, the cancer continues to advance, reaching the liver and lungs; death generally occurs 12–15 years after the development of the primary tumor. Distant spread or metastatic deposits are most commonly in the bone, where the cancer grows at an accelerated pace, fueled by the presence of transferrin, a protein that is abundant in bone tissue (Weinzimer SA, 2001). Transferrin is a insulin-like growth factor (IGF)-binding protein-3 (IGFBP-3). Recent research demonstrates that insulin-like growth factor I (IGF-I) is an important factor for the growth and survival of the LNCaP cell line, while IGF-binding protein-1 (IGFBP-1) induces apoptosis (programmed cell death) (Ngo TH, 2003). A low-fat Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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221

diet and exercise regimen reduce IGF-I and increase IGFBP-1, thereby substantially reducing IGF and prostate cancer (CaP) cell growth. This section discusses key features of CaP that inßuence approaches to medical treatment. Pathophysiology Biology of Prostate Cancer. The development and maintenance of the normal adult prostate are under the hormonal control of androgens acting through the androgen receptor (AR) (Heinlein CA, 2004). The androgens are converted to testosterone in peripheral tissues and the prostate gland. Testosterone diffuses into the epithelial or stromal cells, where it is converted into the functionally active androgen dihydrotestosterone (DHT) by the action of the 5-alpha (−α) reductase enzyme system located on the nuclear membrane (Cheng E, 1993; Russell DW, 1994). DHT binds to the androgen receptor, which undergoes a conformational change and is transported into the nucleus. Inside the nucleus of the cell, the androgen receptor (which is bound to DHT) binds to the target genes and initiates transcription. In this way, the androgen receptor ultimately controls the regulation of the cell cycle, cell growth, and cell differentiation (Cheng E, 1993; Russell DW, 1994). Prostate-Specific Antigen. Prostate-speciÞc antigen (PSA) is a protein produced in the epithelial cells of the prostate (Oesterling JE, 1988). In men younger than age 40, small amounts of PSA (less than 4 ng/mL) circulate in the blood; these levels rise naturally with age, corresponding to an age-dependent increase in prostate size. In general, the higher the PSA level in the bloodstream, the greater the chance of developing CaP. However, some men with CaP do not have high PSA levels, and two-thirds of men with elevated levels of PSA do not have CaP (Schro¨ der FH, 2004). High PSA levels may also indicate nonmalignant conditions, such as benign prostatic hyperplasia. Indeed, it has been suggested that PSA population screening could result in men making the wrong decision and having unnecessary, aggressive treatment (Schro¨ der FH, 2004). In practice, many clinicians believe that PSA velocity or doubling time is more signiÞcant than the absolute PSA level for determining the aggressiveness of the tumor. A recent study found that men with high-risk early-stage CaP (as deÞned by high PSA velocity) have a high risk of death from CaP despite radical prostatectomy (D’Amico AV, 2003). Recent Þndings from a population-based, case-controlled study reported at the annual meeting of the American Society of Clinical Oncology (ASCO) in 2004 suggest that PSA screening among asymptomatic men may substantially reduce the risk of metastatic CaP (Kopec JA, 2004). Further research is needed to conÞrm these results in a randomized trial. A comparison study found that PSA screening of men at ages 40 and 45, then every two years beginning at age 50, would save more lives than the current model—annual screening beginning at age 50 (Ross KS, 2000). The former schedule prevented 3.3 deaths and involved an additional 7,500 tests and 450 prostate biopsies. The standard strategy (annual

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PSA testing beginning at age 50) prevented 3.2 deaths but required an additional 10,500 tests and 600 biopsies. Researchers based these estimates on a PSA level of 4.0 ng/mL for prostate biopsy. The earlier, less frequent schedule would save money and reduce the number of unnecessary biopsies performed each year. The study was primarily in white populations; consequently, these results may not apply to men of other races (Ross KS, 2000). Key applications of PSA as a tumor marker include the following: •

•

•

•

Screening of the general male population. Although widespread, ad hoc screening has clearly resulted in a shift toward the detection of earlier cancers, no prospective, controlled studies of PSA screening have shown that PSA measurement reduces morbidity or mortality (Barry MJ, 2001). Monitoring response to treatment. Failure to achieve PSA of close to 0 ng/mL after local treatment suggests that metastatic disease is already present. Detecting recurrence. A rise in PSA from initially low levels achieved following local treatment such as radiotherapy or radical prostatectomy (RP) likely reßects a recurrence of the tumor (Liao Z, 2004). The availability of the PSA test to detect recurrence (unique among solid tumors) has engendered a huge shift toward early treatment; patients are increasingly treated upon PSA rise rather than waiting several years until metastases are visible on imaging studies. Evaluation of response to novel therapies. The PSA Working Group has recommended a standardized method for reporting PSA response in Phase II trials: trials should report a decline of 50% or greater, versus less than 50%. The group based this standard on the Þndings of several small trials that statistically associated a 50% or greater decline with improved survival (Scher HI, 1999).

Research is ongoing to enhance the sensitivity of PSA testing by using one of the following markers as an adjunct: • • •

Human glandular kallikrein-2. Insulin growth factor-1. Binding protein-3.

Staging. In common with most cancers, prostate tumors are staged according to their degree of metastasis. The most commonly used staging scheme is the primary tumor, regional lymph node, and distant metastases (TNM) system, which is replacing the older Jewett-Whitmore system. Table 1 describes the TNM system in detail. The American Joint Committee on Cancer (AJCC) and the International Union Against Cancer (UICC) adopted the revised TNM staging system in 1992. This system was revised again in 1997 to include reference to tumor grade (the degree of abnormality of cancer cells compared with normal cells). The TNM system

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TABLE 1. TNM Staging Classification System for Prostate Cancer Primary Tumor T1a T1a T1b T1c T1 T2 T2a T2b T3 T4 Any T Any T

Regional AJCC/UICC Lymph Distant Histopathological AJCC/UICC (1997) AJCC (2002) Nodes Metastasis Grade (1992) Stage Stage Stage N0 N0 N0 N0 N0 N0 N0 N0 N0 N0 N1 Any N

M0 M0 M0 M0 M0 M0 M0 M0 M0 M0 M0 M1

G1 G2, 3-4 Any G Any G Any G Any G Any G Any G Any G Any G Any G Any G

I I I I II II II III IV IV IV

I II II II II II II II III IV IV IV

I II II II II II II II III IV IV IV

TNM = Tumor, node, and metastasis. Note: The sixth edition (AJCC 2002) is for cancer cases diagnosed beginning in January 2003. Source: American Joint Committee on Cancer (AJCC) and the International Union Against Cancer (UICC).

now uses broad tumor stage categories, including a stage to describe patients whose only sign of CaP is an abnormal PSA level, and assigns cancers to stages I through IV. TNM was further revised in 2002 to include new T subcategories of prostate tumors for cancer cases diagnosed from January 2003. Figure 1 shows the timescale for CaP’s progression through the disease stages. Grading. Histologically, nearly all CaP are adenocarcinomas, which are further classiÞed by grade. The Gleason scoring system (the most commonly used measure for grading CaP) recognizes that CaP is a multifocal disease with heterogeneous distribution within the gland. Thus, two individual scores, each ranging from 1 to 5, are given to the two most predominant histological patterns of CaP in a biopsy sample. The two scores are added together to give the Gleason sum. In the 1997 AJCC cancer staging manual, sums of 2–4 represent well-differentiated disease; 5–7, moderately differentiated disease; and 8–10, poorly differentiated disease. This system is slightly different in the 2002 edition, in which sums of 5–6 represent moderately differentiated disease and 7–10, poorly differentiated disease (AJCC, 2002). However, the Gleason score is emphasized as the grading system of choice, not the deÞnitions of the scores. Physicians collect the sample via a needle biopsy of the prostate, and the scores are based on morphological appearances corresponding to the size and pattern of the tumor. The Gleason score can help predict the chance of cancer spread. For example, published reports indicate risk of lymph node metastasis in the range of 2%, 13%, and 23% for patients with Gleason scores of Þve, six, and eight, respectively. Gleason scores based on samples taken after radical prostatectomy, rather than on needle biopsy samples, are more accurate and precise; these postprostatectomy samples are assigned Gleason scores to aid in decisions about adjuvant therapy (usually radiation).

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Timescale Localized disease (treated with surgery, radiotherapy, or "watchful waiting") 3 years Rising PSAa 7.5 years Metastases visible on imaging studies (hormone-responsive) 5.5 years Hormone-refractory disease 1 years Death

a. Micrometastatic disease or metastatic disease is present; disease usually responds initially to hormone therapy. Occurs in nearly half of patients treated with local therapy.

FIGURE 1. Prostate cancer disease progression.

Bone Metastases. The skeleton is the major site of CaP metastases. Approximately 70% of CaP metastases involve the skeleton, and the major cause of hospitalization of CaP patients is for the relief of bone pain (Coleman RE, 1987). CaP usually forms osteosclerotic lesions characterized by increased osteolysis (bone destruction) and increased bone formation (osteogenesis) around tumor deposits. Osteolytic metastases can cause severe pain, pathological fractures, lifethreatening hypercalcemia, spinal cord compression, and other nerve-compression syndromes (Roodman GD, 2004). Growth factors potentially involved in increased bone formation include transforming growth factor-beta-2, basic Þbroblast growth factors-1 and -2, bone morphogenetic protein, PSA, and endothelin-1. A recent study found that patients with PSA 20 ng/mL or greater, locally advanced disease (T3 and T4), or Gleason score eight or higher are at high risk for bone metastases; such patients should therefore be considered for bone scan (Abuzallouf S, 2004). Agents that block bone resorption such as the bisphosphonates ease bone pain, spinal cord compression, and the risk of pathological fractures (Charhon SA, 1983; Oades GM, 2002). Bisphosphonates are not covered here because of their lack of direct antitumor effect in CaP; these agents are used only for symptom relief.

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TABLE 2. Combined-Modality Risk Stratification for Prostate Cancer Risk Group Low risk Intermediate risk High risk

Diagnostic Markers PSA ≤10 ng/mL and biopsy Gleason score ≤6 and AJCC T1c or T2a PSA >10–20 ng/mL or biopsy Gleason score of 7 or AJCC T2b PSA >20 ng/mL or biopsy Gleason score of 8–10 or bilateral disease

AJCC = American Joint Committee on Cancer; PSA = Prostate-specific antigen. Source: Adapted from D’Amico AV, et al. Vital statistics following surgery or radiation for patients with clinically localized prostate cancer managed during the PSA era. Proceedings of the American Society of Clinical Oncology. 2003. Abstract 1528.

Prognostic Factors. The probability that an apparently early-stage CaP will recur after local therapy depends on the three most important clinical prognostic factors: the clinical stage of the cancer, its grade, and the PSA level before treatment. Preoperative or postoperative nomograms or “Partin tables” have been developed for localized cancers. These tables combine clinical stage, Gleason score, preoperative PSA levels, surgical margin status, lymph node status, and prostatic cap invasion to aid physicians in evaluating prognosis (Graefen M, 2002). Table 2 shows how PSA stage, AJCC stage, and Gleason score can be combined to stratify risk. When the cancer is conÞned to the prostate gland (stages I or II), median survival is likely to exceed Þve years. Patients with locally advanced cancer (stage III) usually are not curable, and most will eventually die of their tumor, although median survival may be as long as Þve to seven years. If CaP has spread to distant organs (metastatic stage IV disease), current therapies will not cure it, and median survival is usually one to three years. However, even in this last group of patients, some cancers spread slowly and the patient lives for many years with the disease. This scenario highlights the need for better prognostic indicators. In the 1997 AJCC cancer staging manual, poorly differentiated (Gleason score of eight or more) tumors are more likely to have metastasized by the time they are diagnosed and are associated with a worse prognosis. Furthermore, in the majority of studies, ßow cytometry has shown that nuclear DNA ploidy is an independent prognostic indicator for progression and for cause-speciÞc survival in some CaP patients. Diploid tumors have a more favorable outcome than either tetraploid or aneuploid tumors. Among patients with hormone-refractory CaP, factors affecting prognosis include performance status, hemoglobin level, and serum levels of lactate dehydrogenase and alkaline phosphatase (George DJ, 1999). Potential new markers predictive of early relapse (for testing tumor samples after radical prostatectomy) include the following (Kumar-Sinha C, 2003): • • • • •

E-cadherin Microvessel density Ki-67 proliferation index Neuroendocrine differentiation Bcl-2 status

226 • • • • •

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HER2 status CDK1 Caveolin-1 P27 TGF-β

In each case, more studies are needed because available evidence of the predictive power of these markers is either conßicting or preliminary. Etiology The precise etiology of CaP is unknown, although studies suggest that CaP occurs as a result of complex interactions between exposure to environmental toxins and associated genetic changes and predisposition (Abate-Shen C, 2000). CaP appears in the prostate epithelial cells, where precancerous histological changes are common in men in their 20s and 30s; however, CaP does not become evident in most cases until after age 50. Genetic Influences. Genetic epidemiology of CaP remains in its infancy. Most studies have examined the effect of single genes/polymorphisms. Conßicting results have been reported for several of the genetic variations. Table 3 brießy describes the oncogenes and tumor suppressor genes (TSGs) implicated in CaP and their clinical signiÞcance. One key goal in treating CaPl is to map genes responsible for high-grade disease and to Þnd more-sensitive biological markers. A genome-wide linkage analysis of 326 affected sibling pairs, stratiÞed on the basis of Gleason score, has highlighted certain chromosomal regions as containing potential loci important in the development of aggressive disease (Witte JS, 2000). A second goal in understanding the microbiology of CaP is to explain how CaP cells protect themselves against the cytotoxic effects of chemotherapy. One hypothesis suggests that the ability of CaP cells to up-regulate the antiapoptotic proteins Bcl-2 and Bcl-xL is crucial. Bcl-2 blocks the proapoptotic release of cytochrome c from mitochondria; cytochrome c is increasingly expressed in CaP as a function of stage and grade (McDonnell TJ, 1992). All CaPs express Bcl-xL. Blocking the action of these proteins may explain the efÞcacy of taxanes in treating CaP (see “Current Therapies”). Chemosensitizing agents in development for CaP are also targeting these proteins. Androgen-Receptor Mutations. Withdrawal of androgens induces apoptosis in normal prostate epithelial cells (Kyprianou N, 1988). In CaP cells, scientists recognize three phenotypes: androgen-dependent, androgen-sensitive, and androgen-independent. Androgen-dependent tumor cells, like normal cells, die in the absence of androgen. In contrast, androgen-sensitive cells survive in the absence of androgen but grow faster if exposed to the hormone. The growth rate of androgen-independent (also called hormone-refractory) cells is unaffected by the presence of androgen. Androgen-independent cells predominate only in latestage, hormone-refractory, metastatic disease. (Figure 2 shows the sites of action

ETIOLOGY AND PATHOPHYSIOLOGY

227

TABLE 3. Genetic Mutations in Prostate Cancer Gene/Protein

Function

Tumor suppressor genes (TSGs) p53 The p53 protein normally prevents the propagation of DNA damage by causing cell arrest at the G1/S checkpoint and inducing apoptosis. Mutations in p53 remove this regulatory control, allowing mutation-carrying cells to progress through the cell cycle. An intact p53 regulatory mechanism is thought to be necessary for the activity of DNA-damaging cytotoxic agents. CHEK2

RB1

PTEN/MMAC1

Glutathione S-Transferase 1 (GSTP1)

Comments

p53 is mutated in up to 75% of CaP bone metastases (Navone NM, 1999). Introduction of a wild-type p53 suppresses growth of human CaP cell that contains mutant p53 alleles, suggesting a functional role in suppressing prostate tumorigenesis (Isaac WB, 1991). Additionally, p53 protein expression is an independent prognostic marker for disease-free survival after radical prostatectomy (Bauer JJ, 1995). The cell-cycle checkpoint kinase 2 Studies of CHEK2 have shown (CHEK2) encodes an upstream germline mutations in patients regulator of p53 in the DNA with sporadic and hereditary damage signaling pathway. CaP. Most CHEK2 mutations detected were not present in 423 unaffected men (Dong X, 2003). The retinoblastoma tumor Reduced pRb expression is suppressor gene (Rb) encodes a associated with androgenprotein (pRb) that acts stimulated proliferation in the downstream of p53, also human CaP cell line causing cell arrest at the G1/S (LNCaP) (Taneja SS, 2001). The checkpoint. The p16 gene replacement of a mutated Rb regulates pRb function by gene with a normal wild-type inhibiting CDK4 enzyme activity. into CaP cell line (DU145) Inactivation of this gene, suppresses tumorigenecity in therefore, has the downstream nude mice (Bookstein R, 1990). effect of disrupting pRb-mediated cell-cycle control. Phosphatase and tensin homolog Loss of PTEN expression is (mutated in multiple advanced correlated with higher Gleason cancers) is a new TSG that has score and advanced stage been located on chromosome (McMenamin ME, 1999). A 10q23.3. PTEN phosphatase recent study also found that loss negatively regulates cell of PTEN and p27 is correlated interactions with the extracellular with an increased risk of matrix (Tamura M, 1998). recurrence (Halvorsen OJ, 2003). GSTP1 is involved in detoxifying GSTP1 is a promising screening metabolites of environmental marker for detecting carcinogens and protecting DNA organ-confined disease (Harden from oxidative damage. SV, 2003). Hypermethylation of GSTP1 gene is the most common (>90%) genetic alteration in CaP (Harden SV, 2003).

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TABLE 3. (continued) Gene/Protein

Function

Comments

BRCA1 and BRCA2 BRCA1 and BRCA2 are TSGs that Mutations of BRCA1 or BRCA2 regulate proliferation and have been reported in more than differentiation of breast epithelial 2% of Ashkenazi Jews; there cells. BRCA1 and BRCA2 have was an elevated risk of been implicated as a CaP developing CaP as well as breast susceptibility locus. and ovarian cancers (Struewing JP, 1997). RNASEL/HPC1 2’-5–oligoadenylate In small population studies, the (2-5A)-dependent ribonuclease L RNASEL allele with a termination (RNASEL) gene encodes a codon at amino acid position widely expressed latent 265 was detected in 0.54% of endoribonuclease involved in white men without CaP. The degradation of viral and cellular same mutation was detected in RNA. RNASEL is involved in the 4.3% of Finnish men with CaP innate immune response to and only 1.8% of control men. pathogens. The RNASEL gene Among Ashkenazi Jews, the has been linked to hereditary allele was present in 6.9% of prostate cancer 1 (HPC1). men with CaP and 2.9% of Evidence for RNASEL elderly men without CaP involvement in CaP is conflicting. (Rennert H, 2002). One study found no association between mutant RNASEL alleles and CaP. MSR1 MSR1 encodes a macrophage Germline MSR1 mutations have scavenger receptor that can bind been linked to CaP in some to many different ligands, families with hereditary CaP. including oxidized lipoproteins, Among men of European apoptotic cells, and bacteria. descent, MSR1 mutations were MSR1 is involved in the innate found in 4.4% of individuals immune response to pathogens. affected with sporadic CaP but Expression of MSR1 appears to only 0.8% of unaffected men (Xu be restricted to macrophages in J, 2002). Among the prostate that are abundant at African-American men, these sites of inflammation. values were 12.5% and 1.8%, respectively, indicating that MSR1 may be important in susceptibility to CaP in men of both African-American and European descent (Xu J, 2002). KLF6 Kruppel-like factor 6 (KLF6) is a Early LOH analyses have found zinc-finger transcription factor of that one KLF6 gene allele is unknown function that has been deleted in 77% of primary proposed as a candidate TSG prostate tumors; the retained that plays a role in the allele is mutated in 71% of those development of the majority of tumors (Narla G, 2001). Unlike CaPs (Narla G, 2001). wild-type KLF6 (which upregulates p21 independently of p53), tumor-derived mutants fail to reduce cell proliferation.

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TABLE 3. (continued) Gene/Protein Oncogenes HER-2/neu (C-erbB-2)

Function

Comments

The human epidermal growth C-erbB-2 is overexpressed in up to factor HER2 is a transmembrane 60% of patients with glycoprotein with intrinsic hormone-refractory CaP. receptor tyrosine kinase activity Overexpression of C-erbB-2 involved in epithelial cell growth correlates with shorter survival and division. Overexpression of (Jorda M, 2002). High C-erbB-2 the growth receptor on the expression is associated with surface of cells enhances antiandrogen therapy and may metastatic potential. contribute to androgen-independent CaP (Shi Y, 2001). EZH2 polycomb The polycomb group protein EZH2 mRNA and protein are (enhancer of zeste homologue 2) significantly increased in is activated in advanced CaP. malignant CaP compared with EZH2 encodes the EZH2 protein, benign prostate; higher which represses transcription concentrations of E ZH2 mRNA (Karayi MK, 2004). and protein in clinically localized CaP predict poor prognosis (Varambally S, 2002). Other relevant genes and events in CaP E-cadherin and its The presence of E-cadherin and all Decreased expression of E-cad associated catenins are essential for full has been found in high-grade intracellular adhesive function of the cell prostate tumors. It is associated molecules (Pirinen RT, 2001). with recurrence after radical catenins prostatectomy (Kallakury BV, 2001). Androgens Androgens play a major role in the A gene-gene interaction study development of CaP. Androgens found that the combined effect augment the effect of of a high-risk polymorphism in carcinogens present and the AR CAG repeat and a stimulate cell division via the high-risk polymorphism in PSA androgen receptor (AR) (Nelson conferred a higher risk of CaP KA, 2002). The AR is a member than either polymorphism alone of the nuclear receptor (Xue W, 2000). Men who were superfamily of ligand-activated homozygous for the PSA G allele transcription factors. and had short AR CAG repeats Polymorphisms that may play a were at increased risk of CaP role in androgen metabolism and advanced disease. include AR, CYP17, CYP19, SRD5A2, and HSD3B2 (Ntais C, 2003). SRD5A2 SRD5A2 encodes the steroid Studies have shown that 5-a-reductase type 2 enzyme. populations with low This enzyme catalyses the 5-a-reductase activity have a conversion of testosterone into lower incidence of CaP (Ross the main prostatic androgen, RK, 1992). One study found that dihydrotestosterone (DHT). young Japanese men have lower 5-α-reductase activity than do young Caucasian-American and African-American men.

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TABLE 3. (continued) Gene/Protein Vitamin D

HPC2/ELAC2

Function

Comments

Vitamin D is a steroid hormone that Researchers have proposed that plays a role in regulating body vitamin D may influence CaP levels of calcium and risk. Several vitamin D receptor phosphorus, and in polymorphisms have been mineralization of bone. Its action investigated for their role in CaP, is mediated by its receptor. but results from reported studies have been inconsistent. In the case of several polymorphisms (e.g., the poly-A long allele), some studies have shown an increased risk is associated with vitamin D (Ingles SA, 1997), while others showed a protective effect. HPC2/ELAC2 is a candidate CaP In the general population, men who susceptibility gene on carry both of two common chromosome 17p that explains polymorphisms in the ELAC2 hereditary patterns of CaP in gene experience a modest families. HPC2/ELAC2 is increase in risk for CaP. suspected to be associated with Researchers estimate that these an increased risk of developing polymorphisms may play a role the disease. in 2–5% of CaP cases. Researchers found evidence that a mutated version of the gene raised CaP risk in 2 of 33 families who showed an inherited predisposition (Tavtigian SV, 2001).

LOH = Loss of heterozygosity. mRNA = messenger RNA.

of androgen-blocking therapies.) Additionally, androgen-independent progression from an androgen-dependent stage is associated with mutations or ampliÞcation of the androgen-receptor (AR) gene and activation of intracellular signal transduction pathways that stimulate the androgen-receptor gene. Scientists believe that androgen-independent cells come to predominate because androgen depletion exerts selection pressure that favors the growth of preexisting, androgen-independent clones (Craft N, 1999). This progression to androgen independence appears to involve adaptive up-regulation of genes that help cells survive and grow after androgen depletion, together with mutation of androgen receptors. Also contributing are loss-of-function mutations in the genes that code for components of apoptotic pathways. Recent research has demonstrated that in the absence of androgen, the androgen-receptor gene may be activated by protein kinase A and other nonhormonal growth factors (Sadar MD, 1999). Protein kinase A is blocked by certain antiandrogens (e.g., bicalutamide) but not others (e.g., ßutamide); thus, switching from one antiandrogen to another can elicit a response in some patients.

ETIOLOGY AND PATHOPHYSIOLOGY

231

Hypothalamus

LHRH Inhibitors of LH secretion: estrogens, LHRH analogues, progestins

Pituitary

LH Testes

Inhibitors of steroid synthesis: ketoconazole

Inhibitors of ACTH secretion: predinisone ACTH Adrenal

Testosterone

Testosterone Dihydrotestosterone

Prostate cancer cell

5-α reductase inhibitors: finasteride

Androgen receptor Antiandrogens: flutamide, bicalutamide, nilutamide

ACTH = Adrenocorticotropic hormone. LH = Luteinizing hormone. LHRH = Luteinizing hormone-releasing hormone. FIGURE 2. Control of normal prostate cell growth and sites of action of drug therapies.

Resistance to androgen-depletion therapies develops not because of a loss of androgen receptors, which continue to be expressed, but because mutations occur in the androgen-receptor gene. These mutations probably provide a selective growth advantage to a preexisting clone of androgen-independent cells, which can begin to spread under conditions of androgen deprivation. Scientists believe that ampliÞcation of androgen-receptor expression is one mechanism through which cells might become androgen-independent. Overexpressed androgen receptors may be constitutively active (i.e., they stimulate cell growth even in the absence of androgen) or maximally stimulated by very low levels of androgen (if, for example, androgen blockade is incomplete). Risk Factors Age. A man’s age is his most important risk factor for developing CaP (Crawford ED, 2003). CaP incidence increases faster with age than does the incidence of any other major cancer. The probability of developing clinically signiÞcant CaP

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is less than 1 in 10,000 for men younger than age 39, but it skyrockets to 1 in 8 for men aged 60–79. Approximately 80% of men older than 80 have some histological, if not clinical, evidence of CaP. Race and AR-CAG Repeat Status. Epidemiological studies consistently show a higher incidence of CaP in Western Europe and the United States and lower rates in Asia. Black men in the United States have the highest incidence rates worldwide. These Þndings suggest that some racial groups are more susceptible to cancer-promoting events or are more exposed to cancer-promoting agents. CaP risk increases in Asian men when they emigrate to the United States (Whittemore AS, 1995). Studies of the racial-ethnic distribution of a CAG triplet repeat (repetitive DNA sequences) present in exon 1 of the androgen-receptor gene (AR-CAG) identiÞed a link between the size of this microsatellite, the relative risk of developing CaP, and the racial prevalence of the disease. The CAG sequence varies in length from 11 to 31 repeats in healthy men (Crawford ED, 2003). Short AR-CAG alleles (22 repeats or less) are common in African-American men but less so in Asian and Caucasian males. A small study in Caucasians found that the short AR-CAG genotype was associated with a two to three times higher risk of advanced disease (Ingles SA, 1997). However, subsequent epidemiological studies examining the link between short AR-CAG and CaP risk have had mixed results (Crawford ED, 2003). Family History. Studies have shown that slightly more than 40% of CaP cases have a heritable component. Accordingly, family history of CaP is a very important risk factor for the disease. In 1990, investigators reported that men with family members diagnosed with CaP are at signiÞcantly increased risk of developing the disease themselves (Steinberg GD, 1990). The presence of one, two, or three affected family members increased the risk of CaP by a factor of 2, 5, and 11, respectively. Inherited forms of CaP tend to develop at an earlier age than sporadic cases. A recent study of cancer risk among 45,000 twin pairs in Scandinavia found that 42% of CaP cases are inherited, while remaining cases are attributable to environmental factors (Lichtenstein P, 2000). Diet. A case-control study involving 1,655 CaP patients and 1,645 controls (matched by age, ethnicity, and region of residence) found a positive association of CaP risk and total fat intake for all ethnic groups combined (Whittemore AS, 1995). Saturated fat intake was associated with higher risks for Asian-Americans than for blacks and whites. Importantly, among foreign-born Asian-Americans, the risk increased with years of residence in North America and with saturated fat intake. Investigators estimate that differences in saturated fat intake account for 10% of black-white differences and 15% of white-Asian-American differences in CaP incidence (Whittemore AS, 1995). Investigators at the Fred Hutchinson Cancer Center found that men consuming 28 or more servings of vegetables per week had a 35% reduced risk of CaP when compared with those eating fewer than 14 servings per week (Cohen JH, 2000). Some studies have found that

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233

CaP risk is reduced by dietary factors including antioxidants such as selenium, vitamin E, and carotenoid lycopene found in tomatoes (Chen L, 2001; Gann PH, 1999). Generally, the results of studies examining the effect of fruit and vegetable consumption on CaP risk have been inconsistent. Other Risk Factors. Researchers have examined occupation, smoking history, and sexual activity as potential risk factors for CaP, but none has been consistently associated with increased risk. However, smoking appears to elevate the risk of death in men with CaP (Roberts WW, 2003). CURRENT THERAPIES The main treatment modalities for prostate cancer (CaP) include “watchful waiting”; local therapy (prostatectomy or radiotherapy, either external beam radiation therapy [EBRT] or brachytherapy); hormonal therapy; and chemotherapy. Watchful waiting is generally reserved for elderly men, who, because of short life expectancy or slowly progressing disease, are likely to die with CaP rather than because of CaP. Local therapies alone can often cure patients diagnosed with early-stage (I or II) prostate-conÞned disease. Hormonal therapy is used primarily to delay disease progression when local therapies have failed. Chemotherapy is generally reserved for hormone-refractory disease to palliate symptoms. A growing trend in CaP treatment is the use of intermittent therapy. Hormonal therapies are often administered for three years or more as adjuvant therapy. Although their sideeffect proÞle is mild compared with that of many chemotherapy agents, they do have several undesirable effects (e.g., hot ßashes, sexual dysfunction, gynecomastia [excessive development of mammary glands]). To reduce these side effects and improve their quality of life, patients are increasingly requesting breaks from treatment. To establish the role and efÞcacy of intermittent hormonal therapy, the U.S. National Cancer Institute is sponsoring several trials, including SWOG-9346, CALGB-9594, and EU-99013. Estrogen and progestin were among the Þrst hormone agents used to treat CaP. These sex hormones have numerous side effects when prescribed to men, including gynecomastia (breast enlargement and tenderness), hot ßashes, and ßuid retention. More importantly, they have a tendency to boost cholesterol to levels that put patients at risk of cardiovascular disease. These have now been replaced by safer, more-effective therapies. The adrenal inhibitors aminoglutethimide (Novartis’ Orimeten/Cytadren) and ketoconazole (Janssen-Pharmaceutica’s Nizoral) have shown some evidence of a beneÞcial effect in hormone-refractory CaP. Aminoglutethimide is marketed for the palliative treatment of metastatic CaP on the basis of subjective improvements in symptoms and pain relief but has not demonstrated any beneÞt in terms of survival. Ketoconazole is not marketed for the treatment of CaP despite numerous clinical trials demonstrating a marginal beneÞt. These agents are not discussed in detail here because recent treatment advances have rendered them marginal to mainstream treatment choices.

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Table 4 summarizes the leading therapies available to treat CaP. Table 5 compares their efÞcacy, advantages, and disadvantages. Luteinizing Hormone-Releasing Hormone Analogues Overview. Luteinizing hormone-releasing hormone (LHRH) analogues are central to the treatment of all patients with hormone-sensitive CaP. LHRH analogues are an effective alternative to surgical castration (orchiectomy), a treatment to which very few men consent. To be approved, all LHRH analogues demonstrated, in open-label, randomized trials, testosterone suppression equivalent to that of orchiectomy. Testosterone suppression removes the main tumor growth stimulus and leads to tumor cell death. Regulatory authorities in the seven major pharmaceutical markets (United States, France, Germany, Italy, Spain, United Kingdom, and Japan) have determined that any agent that suppresses testosterone to the same extent as orchiectomy will have the same antitumor effect as orchiectomy. Mechanism of Action. As illustrated in (Figure 2) (the major sites of hormone blockade), LHRH analogues exert their action on the pituitary. Initial administration of an LHRH analogue increases LHRH release from the pituitary, thereby triggering the production of testosterone from the testes and causing an initial tumor growth spurt (tumor ßare). This initial increase in testosterone production lasts approximately two to four weeks. Subsequently, the LHRH analogue blocks the effect of endogenous LHRH on the pituitary, causing a decline in body testosterone levels similar to those achieved with castration. However, unlike castration, the effects of LHRH analogues are reversible upon withdrawal of therapy. LHRH analogues suppress only androgen production by the testes and have no effect on androgen production of the adrenal glands. The fall in body testosterone achieved by androgen suppression alone is sufÞcient to cause cell death of androgen-dependent tumor cells. Formulation. To overcome their short duration of action, the majority of LHRH analogues are formulated as depot preparations. Numerous formulations are available that can provide a continuous plasma level of LHRH sufÞcient to suppress testosterone for 1–12 months. The majority of leuprolide preparations (including Tap Pharmaceutical’s Lupron, Takeda’s Enantone/Trenantone, Abbott’s Procrin/Ginecrin, and Wyeth’s Prostap) and triptorelin (Ferring/Ipsen’s Decapeptyl, PÞzer’s Trelstar) are formulated as microspheres, which are mixed with a viscous vehicle prior to administration. The resulting suspension is administered either subcutaneously or intramuscularly to form a drug depot. The active drug is released from the depot at a controlled rate into the circulation to achieve the desired drug levels for periods of one, three, four, or six months. Goserelin (AstraZeneca’s Zoladex) (Figure 3) is formulated into a solid implant that is administered subcutaneously into the abdominal wall via a preÞlled syringe. The implant disintegrates at a

TABLE 4. Current Regimens/Classes Used for Prostate Cancer Regimen or Class

Regimen Components Agent

Availability

Dose

Common Toxicities Impotence, gynecomastia, hot flashes, sweating, peripheral edema, gastrointestinal disturbances, mood changes, decreased libido Renal/metabolic disorder, central nervous system disorder

LHRH analogues

Leuprolide acetate (leuprorelin)

Goserelin Triptorelin

TAP Pharmaceutical’s Lupron, Takeda’s Enantone/Trenantone, Abbott Laboratories’ Procrin/Ginecrin, Wyeth’s Prostap, generics Bayer’s Viadur AstraZeneca’s Zoladex

US, F, G, I, S, UK

22.5 mg SC monthly/30 mg SC three times monthly/65 mg SC six times monthly

US US, F, G, I, S, UK, J

Pfizer’s Trelstar, Ferring/Ipsen’s Decapeptyl

US, F, G, I, S, UK

65 mg SC every 12 months 3.6 mg SC monthly/10 mg SC three times monthly 3 mg SC monthly/11.5 mg SC three times monthly

Buserelin

235

Sanofi-Aventis’s Superfact/Profact, Sanofi-Aventis/Mochida’s Supurecur, generics Nonsteroidal antiandrogens

F, G, I, S, UK, J

0.5 mg SC q8h for seven days, then intranasal six times each day

Bicalutamide

AstraZeneca’s Casodex

US, F, G, I, S, UK, J

50/150 mg

Flutamide

Schering-Plough’s Eulexin/Eulexine/Drogenil, Nippon-Kayaku’s Odyne, generics

US, F, G, I, S, UK, J

750 mg qd

Pain, headache, dizziness, itching, mental depression, paranoia Headache (in the case of nasal administration) Impotence, hot flashes, fatigue, gastrointestinal disturbances, cardiovascular disorder, renal toxicity, gynecomastia Neurological disorder, pain, infection, alopecia, rash Liver toxicity, depression

236

TABLE 4. (continued) Regimen or Class

Regimen Components Agent

Availability

Dose

Common Toxicities

Steroidal antiandrogens Cyproterone Schering’s Androcur/Cyprostat, acetate Jenapharm’s Androcur, generics

F, G, UK

200–300 mg

Benign and malignant liver changes, adrenal-cortical suppression, thromboembolism, osteoporosis

Combined androgen blockade An LHRH analogue A nonsteroidal antiandrogen

US, F, G, I, S, UK, J

monthly/three times monthly 50/150 mg qd

Impotence, gynecomastia, hot flashes, sweating, peripheral edema, gastrointestinal disturbances, mood changes, decreased libido

US, F, G, I, S, UK, J

280 mg po tid for eight weeks, then 140 mg po tid 75 mg/m2 IV every 21 days plus 5 mg of prednisone daily continuously

Gastrointestinal toxicity

Docetaxel 70 mg/m2 on day one; estramustine 280 mg po q6h for five doses

Neutropenia, thrombocytopenia, fatigue, diarrhea, nail changes, edema, asthenia

12 mg/m2 on day 1 of a 21-day cycle; prednisone 5 mg po qd continuously

Neutropenia

Chemotherapy regimens Estramustine Pfizer’s Emcyt/Estracyt, Nihon (single agent) Kayaku’s Estracyt, generics Docetaxel/ Sanofi-Aventis’ Taxotere prednisone

Docetaxel/ estramustine

Mitoxantrone/ prednisone

US, F, G, I, S, UK, J

Merck’s Decortin generics Sanofi-Aventis’s Taxotere

US, F, G, I, S US, F, G, I, S, UK, J

Pfizer’s Emcyt/Estracyt, Nihon Kayaku’s Estracyt, generics Serono/Wyeth Lederle’s Novantrone, Baxter’s Onkotrone

US, F, G, I, S, UK, J

Merck’s Decortin, generics

US, F, G, I, S, UK, J

Neutropenia, bone pain, infection, fatigue, diarrhea

US, F, G, I, S

US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan. IM = Intramuscular; IV = Intravenous; LHRH = Luteinizing hormone-releasing hormone; po = Orally; q8h = Every eight hours; qd = Once each day; SC = Subcutaneous; tid = Three times daily. Note: In most European countries, prednisolone is used in place of prednisone, which is used primarily in the United States.

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controlled rate, thus providing a constant plasma level of goserelin for one or three months depending on the preparation used. Buserelin (SanoÞ-Aventis’s Superfact/Profact, SanoÞ-Aventis/Mochida’s Supurecur, generics) is the only product formulated into a nasal spray. Owing to LHRH analogues’ short duration of action, Superfact must be administered six times a day. In contrast, Bayer’s Viadur is a nonbiodegradable implant with 12-month activity. Viadur is a titanium alloy reservoir of leuprolide that relies on osmosis to pump the active drug into the systemic circulation at a rate of 120 micrograms each day. After 12 months, the spent reservoir must be removed (Alza, 2002). Viadur requires comparatively complex surgical implantation and removal. Leuprolide. Leuprolide (Figure 4) is formulated into Þve branded products, all of which vary slightly in their presentation, but their clinical effect is identical. TAP Pharmaceutical markets leuprolide as Lupron in the United States for the treatment of both CaP and breast cancer (CaB), while Takeda markets it as Enantone/Trenantone in France, Germany, and Italy for the same indications. It is available in a one-, three-, and four-month depot preparations formulated into a suspension for subcutaneous (SC) injection. Wyeth’s Prostap is marketed in the United States and the United Kingdom for the treatment of advanced CaP. Prostap is available in both one- and three-month preparations formulated into an aqueous suspension for SC administration. Abbott Laboratories’ Procrin/Ginecrin is marketed for CaP in Spain, where it is available in one- and four-month doses. Bayer’s Viadur was launched in the United States in 2000; it is the only product marketed with a 12-month dose duration. Alza is the patent holder and sole manufacturer of Viadur, but in 2000, the company signed a commercialization agreement with Bayer to market Viadur in the United States. Viadur is marketed only for the treatment of CaP. Treatment with leuprolide stimulates testosterone secretion for the Þrst month and may cause tumor ßare, but the concurrent administration of an antiandrogen (discussed in the sections “Nonsteroidal Antiandrogens” and “Steroidal Antiandrogens”) will prevent tumor ßare. Goserelin. AstraZeneca’s Zoladex is marketed for the treatment of CaP, CaB, endometriosis, uterine Þbroids, and assisted reproduction. Goserelin is available in one- and three-month implant dosage forms. An analogue of endogenous LHRH, goserelin, like leuprolide, stimulates testosterone production for the Þrst month, but chronic goserelin administration completely blocks the production of endogenous LHRH. Because goserelin is formulated into a disintegrating pellet to achieve an SC depot, the gauge of the syringe required to deliver the intact pellet is large and therefore unpopular among some patients. It is also recommended that the SC injection be given only into the abdominal wall. Triptorelin. Triptorelin (PÞzer’s Trelstar, Ferring/Ipsen’s Decapeptyl) is marketed for the treatment of advanced CaP only. Ferring markets triptorelin in

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TABLE 5. Comparison of Current Therapies for Prostate Cancer Treatment LHRH analogues

Nonsteroidal antiandrogens

Steroidal antiandrogens Combined androgen blockade

Estramustine (single agent)

Estramustine/ docetaxel

Docetaxel/ prednisone

Mitoxantrone/ prednisone Watchful waiting

Advantages • Reversible suppression of testosterone. • More acceptable to men than castration. • 1-, 3-, 4-, and 12-month depot injections available.

Disadvantages • Chronic therapy causes testicular atrophy and is associated with decreased libido. • Loss of bone mineral density.

• No suppression of testosterone production. • Oral therapy.

• Significant risk of cardiac disorders.

• Oral therapy.

• Numerous side effects associated with adrenocortical suppression.

• Total androgen deprivation removes tumor growth stimulus. • Removal of tumor-flare risk associated with initial LHRH therapy.

• Increased number and frequency of side effects compared with single-agent treatment.

• Cytotoxic agent that also causes testosterone suppression.

• No survival advantage provided by therapy. • Significantly more toxic than androgen therapy.

• Provides a three-month survival advantage to patients with hormone-refractory disease.

• More toxic than single-agent cytotoxic therapy. • Significantly more toxic than endocrine therapy.

• Provides a survival advantage to patients with hormone-refractory disease. • Less toxic than combination chemotherapy.

• Significantly more toxic than endocrine therapy.

• Well tolerated therapy hormone-refractory disease.

• No proven survival advantage.

for

• No side effects and noninvasive.

• Appropriate only for elderly patients with slow-growing tumors. • Requirement for regular PSA testing. • Survival disadvantage for men with life expectancy longer than 15 years.

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TABLE 5. (continued) Treatment

Advantages

Radical prostatectomy

• Effective treatment of young patients. • Associated with the lowest risk of relapse of all treatments (Oesterling J, 1997).

Disadvantages

• Surgery associated with significant morbidity, including incontinence, impotence, urethral stricture, and rectal injury.

Radiation therapy • Effective treatment for inoperable tumors. • Local therapy with limited toxicity.

• Associated with morbidity caused by damage to surrounding tissue, including impotence, incontinence, proctitis, cystitis, rectal and urethral strictures, and bowel obstruction.

LHRS = Luteinizing hormone-releasing hormone.

H-5-OxoPro-His-Trp-Ser-Tyr-D-Ser(t-Bu)-Leu-Arg-Pro-NHNHCONH2 FIGURE 3. Structure of goserelin.

H-5-OxoPro-His-trp-Ser-Tyr-D-Leu-Leu-Arg-ProNHCH2CH3 CH3COOH FIGURE 4. Structure of leuprolide.

Germany; Ipsen is responsible for marketing in France, Italy, Spain, and the United Kingdom; and PÞzer owns marketing rights in the United States. An analogue of endogenous LHRH, triptorelin, like leuprolide and goserelin, stimulates testosterone production for the Þrst month, but chronic administration of the compound completely blocks the production of endogenous LHRH. Unlike other LHRH analogues, triptorelin’s marketing approval prevents the drug from being prescribed to men with bone metastases who are at risk of spinal cord compression caused by tumor ßare. Buserelin. Buserelin (SanoÞ-Aventis’ Superfact/Profact, SanoÞ-Aventis/ Mochida’s Supurecur, generics) is marketed in Europe and Japan for the treatment of advanced CaP. It is not available in the United States. The compound is available as a subcutaneous (SC) injection and nasal spray. An analogue of endogenous LHRH, buserelin stimulates testosterone production for the Þrst month. Chronic administration of the compound then completely blocks the production of endogenous LHRH.

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Buserelin is not available in depot formulation. For the Þrst seven days, patients are administered an SC injection every eight hours. Patients are then switched to the less invasive nasal spray, which must be administered six times a day. Luteinizing Hormone-Releasing Hormone Antagonists Overview. LHRH antagonists are the most recent class of hormonal therapy to enter the CaP armamentarium. Mechanism of Action. Compared with LHRH analogues (e.g., goserelin, leuprolide acetate), which produce their effect by activating and then desensitizing androgen-producing cells to LHRH, LHRH antagonists directly block the effect of the releasing hormone. Abarelix. NOTE: This drug has been discontinued. The LHRH antagonist abarelix (Praecis Pharmaceuticals/Schering AG’s Plenaxis) received FDA approval in 2003 for the palliative treatment of men with advanced, symptomatic CaP for whom the temporary testosterone “ßare” associated with LHRH analogue treatment could be dangerous and who refuse orchiectomy. However, in 2005, Praecis Pharmaceuticals has voluntarily discontinued selling Plenaxis to new patients in the United States for economic/commercial reasons. In fact, clinicians interviewed say, ßare is easily prevented by combining an antiandrogen with an LHRH analogue and that abarelix never fulÞlled an urgent unmet need. Nonsteroidal Antiandrogens Overview. The three marketed nonsteroidal antiandrogens have similar clinical activity and differ only in their marketing authorizations. The following paragraphs proÞle the two most widely used antiandrogens: bicalutamide (AstraZeneca’s Casodex) (Figure 5) and ßutamide (Schering-Plough’s Eulexin/Eulexine/Drogenil, Nippon-Kayaku’s Odyne, generics) (Figure 6). Nilutamide (SanoÞ-Aventis’s Nilandron) is indicated only for the treatment of metastatic (stage IV) CaP in combination with surgical castration and is unavailable in most of Europe. Like the LHRH analogues, the antiandrogens have no direct antitumor activity and rely on androgen deprivation to kill androgendependent tumor cells. Unlike the case for the LHRH analogues, their clinical efÞcacy has been proven in placebo-controlled clinical trials assessing tumor response. Mechanism of Action. Nonsteroidal antiandrogens prevent binding of testosterone and dihydrotestosterone (DHT) (androgens) to the androgen receptor within normal and cancerous prostate cells (Figure 2). In this way, they stop prostate cell growth. Nonsteroidal antiandrogens have a higher antiandrogenic activity than steroidal antiandrogens (discussed in more detail in the next section). Because they do not suppress testosterone production, they are not as frequently

CURRENT THERAPIES

HO O F

CH3 O

S

241

CF3

HN

CN

O FIGURE 5. Structure of bicalutamide.

CH3 H N H3C

CF3

O NO2 FIGURE 6. Structure of flutamide.

associated with impotence or atrophy of the testis. In addition, they do not have the side effects associated with steroidal antiandrogens. Nonsteroidal antiandrogens are prescribed a few days before the initiation of LHRH analogue therapy. By preventing the initial surge in testosterone levels that can cause a tumor growth spurt, they prevent tumor ßare. Formulation. Currently available nonsteroidal antiandrogens are administered orally. Bicalutamide (AstraZeneca’s Casodex) is administered once daily, while ßutamide (Schering-Plough’s Eulexin/Eulexine/Drogenil, NipponKayaku’s Odyne, generics) must be administered three times a day. Bicalutamide. Bicalutamide (AstraZeneca’s Casodex) is marketed for the treatment of stage IV CaP in combination with LHRH therapy (combined androgen blockade, discussed later) or surgical castration. High-dose bicalutamide (150 mg daily) is also marketed for the treatment of locally advanced CaP as an adjuvant to prostatectomy or radiotherapy. Patients with locally advanced CaP who are not candidates for surgical castration or other interventions (including prostatectomy, radiotherapy) can be treated with bicalutamide monotherapy. In clinical trials, bicalutamide as a single-agent therapy demonstrated a survival advantage in patients with locally advanced CaP. However, in men with metastatic disease, bicalutamide must be combined with an LHRH analogue to provide a survival advantage over castration. Two multi-center trials with identical protocols randomized 1,453 patients with locally advanced (tumor, node and metastasis [TNM] stage T3/T4 M0) or metastatic disease (TNM stage T4 M1) to either 150 mg bicalutamide each day or castration (Tyrrell CJ, 1998). Pooled analysis at a median follow-up of 100 weeks for both studies found bicalutamide was less effective than castration in patients with metastatic disease. Castration improved median survival by six weeks over bicalutamide. Bicalutamide provided a signiÞcant improvement only in subjective response: 70% versus 58% over castration in men with

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symptomatic metastatic CaP. Quality-of-life questionnaires found bicalutamide treatment offered an advantage in terms of sexual interest and physical capacity and a substantially lower incidence of hot ßashes compared with castration. Pooled analysis of two trials involving patients with nonmetastatic CaP was completed after four years of follow-up (Iversen P, 1998). A total of 480 patients with TNM stage T3/T4 nonmetastatic disease were randomized to bicalutamide 150 mg each day or castration (either orchiectomy or goserelin). Bicalutamide monotherapy was statistically equivalent to castration in the pooled survival analysis. Different end points for time to treatment failure and objective progression between the two trials prevented pooled analysis of these end points. However, one trial favored bicalutamide and the other favored castration. Men treated with bicalutamide maintained a higher sexual interest and capacity than those who were castrated. Because of its poor clinical performance, bicalutamide should not be used as monotherapy in patients with metastatic disease. Gynecomastia (breast enlargement and tenderness) occurs in up to 10% of patients. Hot ßashes, depression, decreased libido, and weight gain occur less frequently. Flutamide. Flutamide (Schering-Plough’s Eulexin/Eulexine/Drogenil, NipponKayaku’s Odyne, generics) is marketed for the treatment of advanced (stage IV) CaP in combination with an LHRH analogue or as monotherapy in castrated patients. Patients who have failed, or are not able to tolerate, other forms of hormone manipulation can also be treated with ßutamide. Flutamide in combination with an LHRH analogue is also marketed for the initial treatment of bulky tumors conÞned to the prostate or those that extend beyond the prostate capsule. Clinical data do not support ßutamide monotherapy in noncastrated patients. Flutamide as a treatment for metastatic CaP is discussed in the upcoming section “Combined Androgen Blockade.” Flutamide must be administered orally every eight hours, which is considerably more inconvenient than bicalutamide’s oncedaily administration. It is associated with hepatic injury—mainly transaminase abnormalities, cholestatic jaundice, hepatic necrosis, depression, and hepatic encephalopathy. The hepatic injury can be fatal. Steroidal Antiandrogens Overview. Chronic administration of steroidal androgens can suppress adrenocortical function (i.e., interfere with the body’s ability to regulate endogenous steroid production). These agents have been replaced by the nonsteroidal antiandrogens, which lack this complicating side effect. Mechanism of Action. Steroidal antiandrogens prevent binding of testosterone and dihydrotestosterone (DHT) (androgens) to the androgen receptor within normal and cancerous prostate cells (Figure 2). In this way, they stop prostate cell growth. Steroidal antiandrogens have less antiandrogenic activity than nonsteroidal antiandrogens, but because they have a progestin-like effect on the pituitary, they can reduce the secretion of luteinizing hormone from the pituitary.

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FIGURE 7. Structure of cyproterone acetate.

As a result, steroidal antiandrogens can reduce testosterone secretion to levels similar to those achieved by castration (Figure 2). Cyproterone Acetate. Cyproterone acetate (Schering’s Androcur/Cyprostat, Jenapharm’s Androcur, generics) (Figure 7) is marketed in France, Germany, and the United Kingdom for the treatment of CaP. It is approved for the suppression of tumor ßare with initial LHRH analogue therapy, for the treatment of metastatic CaP when alternative treatment interventions are contraindicated or not tolerated, and for the treatment of hot ßashes in castrated men receiving LHRH analogues. Numerous trials have been conducted to elucidate cyproterone’s activity in the treatment of locally advanced or metastatic CaP. The pivotal trial in this quest was conducted by the Urological Group of the European Organization for Research on Treatment of Cancer. The Phase III trial randomized 210 patients to receive cyproterone, medroxyprogesterone, or diethylstilbestrol therapy (Pavone-Macaluso M, 1986). Of the three drugs studied, patients treated with medroxyprogesterone had a signiÞcantly shorter duration of time to progression and survival than did those receiving diethylstilbestrol or cyproterone. There was no signiÞcant difference between diethylstilbestrol and cyproterone in terms of overall survival, time to progression, and tolerability. A review of the aforementioned trial and the drugs available in 1988 concluded that the safest and most cost-effective endocrine treatment for metastatic CaP was orchiectomy (Bollack C, 1988). Cyproterone’s steroidal effects are responsible for the majority of its side effects, of which the most serious are benign and malignant liver changes. Adrenocortical suppression can occur, so patients must be monitored for the duration of treatment. Thromboembolism is a considerable risk for patients with a history of thrombosis or diabetics with microvascular disease. Diabetics must be monitored continually because cyproterone can change carbohydrate metabolism. The compound is also linked to osteoporosis. Combined Androgen Blockade Overview. Combined androgen blockade (CAB)—sometimes called androgendeprivation therapy—is the simultaneous administration of an LHRH analogue

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and an antiandrogen. A huge number of randomized, controlled trials have been undertaken to assess the beneÞt of adding an antiandrogen to LHRH therapy. Mechanism of Action. The individual components of this regimen contribute the following mechanisms to achieve the overall regimen activity: •

•

LHRH analogues competitively bind to the pituitary LHRH receptors. After initial stimulation of testosterone production, total block occurs (Figure 2). LHRH analogues suppress testosterone levels to levels achieved by castration. Nonsteroidal antiandrogens prevent binding of testosterone and dihydrotestosterone (DHT) (androgens) to the androgen receptor within normal and cancerous prostate cells (Figure 2). In this way, they stop prostate cell growth.

The combination of blocking androgen production via the pituitary and blocking the effect of androgens produced from other sources, including the adrenal glands, completely removes the endocrine growth stimulus on the tumor cells. An additional beneÞt of combining treatment is prevention of the initial tumor ßare that occurs in the Þrst month of LHRH analogue therapy. The initial surge in testosterone production has no growth effect on tumor cells because the antiandrogen blocks the androgen receptors. Clinical Performance. A National Cancer Institute (NCI) intergroup trial randomized 600 patients to receive either leuprolide or leuprolide plus ßutamide (Crawford ED, 1989). Initially, patients receiving CAB appeared to have a longer time to disease progression (16.5 versus 14 months) and improved survival (37 versus 28 months), but subsequent meta-analyses showed mixed results. For example, the most recent update of the CaP Trialists’ Collaborative Group metaanalysis (CaP Trialists’ Collaborative Group, 2000) found that outcome depends on the antiandrogen used. In this meta-analysis, which comprised 27 clinical trials and 8,000 patients, CAB with nonsteroidal antiandrogens was associated with an 8% reduction in the risk of death and a 2.9% beneÞt in Þve-year survival; in contrast, CAB with the steroidal antiandrogen cyproterone acetate was associated with a 13% increased risk of death and a 2.8% reduction in Þve-year survival. The aforementioned meta-analysis implies that differences among antiandrogens may account for the discrepancy in CAB’s efÞcacy. One hypothesis suggests that activation of the androgen receptor induced by protein kinase A is differentially blocked by various nonsteroidal androgens (Klocker H, 1999; Sadar MD, 1999). Androgen-independent tumor growth may occur because growth factors such as protein kinase A activate the androgen receptor. The achievement of second-line tumor responses by a change in antiandrogen (e.g., from ßutamide to bicalutamide) supports this hypothesis. Estramustine (Single Agent) Overview. Until recently, estramustine (PÞzer’s Emcyt/Estracyt, Nihon Kayaku’s Estracyt, generics) (Figure 8) was one of two chemotherapeutic agents marketed for the treatment of CaP. Estramustine was speciÞcally designed to

CURRENT THERAPIES

O H3C

O

P

245

O− Na+ O− Na+

H O H

H

Cl N

O

Cl FIGURE 8. Structure of estramustine.

combine antiandrogen activity with cytotoxicity. Its poor clinical performance as a single agent prompted researchers to assess estramustine in combination with other cytotoxics. One such study is discussed in the upcoming section “Docetaxel/Estramustine.” Mechanism of Action. Estramustine’s exact mechanism of action is unknown. Researchers postulate that it has a dual mechanism of action. Prior to metabolism, the intact molecule has an antimitotic action, possibly achieved by the stabilization of microtubule dynamics by binding to a novel site in tubulin (Panda D, 1997). Additionally, some of the metabolites exert an antigonadotrophic effect owing to their estrogen-like activity. This antigonadotrophic effect suppresses testosterone. Clinical Performance. In a randomized, Phase III trial conducted by the Urological Group of the European Organization for Research and Treatment of Cancer, 227 patients with T3 or T4 CaP were randomized to estramustine or diethylstilbestrol therapy (Smith PH, 1986). The patients in the estramustine arm received a dose of 280 mg twice a day for the Þrst eight weeks, after which the dose was reduced to 140 mg twice a day. Patients randomized to the diethylstilbestrol arm were treated with 1 mg three times each day. Duration of response was made by assessment by palpation of local tumor response. Patients treated with diethylstilbestrol had a signiÞcantly better local tumor response, but the investigators found no signiÞcant difference between the two drug treatments in response rate of metastases, interval to local and distant progression, and overall survival. Diethylstilbestrol was signiÞcantly more cardiotoxic than estramustine, while estramustine caused gastrointestinal toxicity in 25 patients, six of whom stopped treatment. Docetaxel/Prednisone Overview. In May 2004, the FDA approved the regimen of docetaxel (SanoÞAventis’s Taxotere) (Figure 9) plus prednisone (Merck’s Decortin, generics) (Figure 10) for the treatment of hormone-refractory CaP. This regimen’s apparent efÞcacy is prompting further research in the use of docetaxel in combination with other chemotherapy agents in the hope of improving overall survival for

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FIGURE 9. Structure of docetaxel.

CH2OH C O

H3C

O OH

H3C

O FIGURE 10. Structure of prednisone.

this indication. In November 2004, docetaxel was approved in Europe for the treatment of metastatic, hormone-refractory CaP. Mechanism of Action. The individual components of the docetaxel/ prednisone regimen contribute the following mechanisms to achieve its overall activity: •

•

Docetaxel is a taxane. Taxanes inhibit the dynamic reorganization of microtubule networks, which is essential to vital interphase and mitotic cellular functions. Docetaxel, which achieves selective toxicity against rapidly proliferating cells, is mainly active in the S phase of the cell cycle. Prednisone is a glucocorticosteroid. This class of drugs reduces the inßammatory response to tumor tissue by preventing white blood cells from functioning to produce swelling and pain around the tumor site. Glucocorticosteroids induce lysis of peripheral lymphocytes and slow lymphocyte production. The addition of glucocorticosteroids to chemotherapy regimens may augment their efÞcacy.

Clinical Performance. The TAX-327 study stratiÞed 1,006 patients with histologically proven, hormone-refractory, metastatic CaP by pain and performance status and randomized the patients to one of three treatment arms (Tannock IF, 2004). The trial’s primary end point was survival, and secondary end points were PSA response, pain, and toxicity. Patients in arm A received prednisone (5 mg orally twice daily) and docetaxel (75 mg/m2 every three weeks for ten cycles); arm B patients received the same dose of prednisone plus docetaxel 30 mg/m2 weekly for Þve weeks out of six for Þve cycles; and arm C patients received

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the same dose of prednisone and 12 mg/m2 mitoxantrone every three weeks for ten cycles. Grade 3 or 4 toxicities occurred in the following percentages of patients in arms A, B, and C, respectively: neutropenia 32%, 1.5%, and 21.7%; bone pain 7.8%, 7.3%, and 9.9%; infection 5.7%, 5.5%, and 4.2%; fatigue 4.5%, 5.5%, and 5.1%; and diarrhea 2.1%, 4.8%, and 1.2%. Although the investigators stated that baseline characteristics were similar in each arm, they did not provide information regarding these characteristics (e.g., median age, baseline PSA value, performance status). The median survival times in arms A, B, and C were 18.9 months, 17.4 months, and 1.5 months, respectively. A p-value for the difference in median survival was not presented, but the hazard ratio for patients treated with docetaxel every three weeks (arm A) relative to mitoxantrone (arm C) was signiÞcantly reduced: hazard ratio 0.76 (95% conÞdence interval [CI] 0.62–0.94), p = 0.009. The hazard ratio for the docetaxel arms combined remained signiÞcantly reduced—hazard ratio 0.83, 95% CI 0.7–0.99, p = 0.04—but the hazard ratio for weekly docetaxel (arm B) relative to the mitoxantrone arm (arm C) was not signiÞcantly reduced. Patients with baseline pain intensity greater than two were assessed for their pain response, and patients with baseline PSA greater than 20 ng/mL were assessed for their PSA response. Again, patients in arm A achieved a signiÞcantly higher rate of pain response than did those in arm C (35% versus 22%), p = 0.01, and a signiÞcantly higher PSA response (45% versus 32%). Because of its impact on survival, arm A (prednisone with thrice-weekly docetaxel) is deemed the most effective treatment for metastatic, hormonerefractory CaP. Docetaxel/Estramustine Overview. Researchers have investigated estramustine in combination with many chemotherapy agents. For example, the combination of estramustine with docetaxel (SanoÞ-Aventis’ Taxotere) has achieved promising results. However, the FDA’s May 2004 approval of docetaxel/prednisone for the treatment of hormone-refractory CaP means that the docetaxel/estramustine combination is unlikely to be widely used. Mechanism of Action. The individual components of the docetaxel/ estramustine regimen contribute the following mechanisms to achieve its overall activity: •

•

Docetaxel is a taxane. Taxanes inhibit the dynamic reorganization of microtubule networks, which is essential to vital interphase and mitotic cellular functions. Docetaxel, which achieves selective toxicity against rapidly proliferating cells, is mainly active in the S phase of the cell cycle. Estramustine’s exact mechanism of action is unknown. Researchers postulate that it has a dual mechanism of action. Prior to metabolism, the intact molecule has an antimitotic action, possibly achieved by the stabilization

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H N OH

O

HN

OH

O

HN

OH

OH N H

FIGURE 11. Structure of mitoxantrone.

of microtubule dynamics by binding to a novel site in tubulin (Panda D, 1997). Additionally, some of the metabolites exert an antigonadotrophic effect owing to their estrogen-like activity. This antigonadotrophic effect suppresses testosterone. Clinical Performance. At the 2004 American Society of Clinical Oncology (ASCO) Conference, the Southwest Oncology Group (SWOG-9916) presented the results of a trial comparing docetaxel plus estramustine with mitoxantrone plus prednisone in 770 patients with hormone-refractory CaP (Petrylak DP, 2004). All patients were premedicated with 60 mg dexamethasone. Patients in the docetaxel/estramustine arm received 60 mg/m2 docetaxel by intravenous (IV) infusion on day 2 plus 280 mg estramustine orally days 1–5. Patients in the mitoxantrone/prednisone arm received mitoxantrone 12 mg/m2 IV on day 2 plus 5 mg prednisone orally, twice each day. Cycles were given every 21 days. If no grade 3 or 4 toxicities occurred, the patient’s dose was escalated to 70 mg/m2 docetaxel and 14 mg/m2 mitoxantrone. The study was designed to detect a 33% difference in survival. Arms were balanced for age, race, performance status, PSA, and symptoms. The median survival of men treated with docetaxel versus mitoxantrone was 18 months versus 15 months (p = 0.008). Median time to progression was also higher in the docetaxel arm (six months versus three months, p < 0.0001). Response rates in measurable disease were not signiÞcantly different. Grade 3 or 4 toxicities were reported in 54% of patients treated with docetaxel and 34% of patients treated with mitoxantrone. This difference arose from higher rates of gastrointestinal and cardiovascular toxicity. There was no signiÞcant difference in deaths due to toxicity (2% in the docetaxel arm versus 1% in the mitoxantrone arm). Mitoxantrone/Prednisone Overview. Mitoxantrone (Serono/Wyeth Lederle’s Novantrone, Baxter’s Onkotrone) (Figure 11) is marketed for the treatment of hormone-refractory metastatic CaP in combination with prednisone. Despite mitoxantrone’s inability to improve overall survival, it is well tolerated and thus an appropriate palliative treatment.

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Mechanism of Action. The individual components of the mitoxantrone/ prednisone (MP) regimen contribute the following mechanisms to achieve their overall activity: •

•

Mitoxantrone is an antitumor antibiotic. It is an anthracenedione and structural analogue of the antibiotic anthracyclines. Mitoxantrone’s exact mechanism of action is unknown, but it intercalates with DNA to cause interstrand and intrastrand cross-linking. Mitoxantrone also causes DNA strand breaks by binding with the phosphate backbone of DNA. The agent is cell-cycle-phase-nonspeciÞc. Mitoxantrone also impairs the strandreunion reaction of topoisomerase II. These effects result in the production of protein-linked, double-stranded DNA breaks. Although mitoxantrone is cytotoxic to cells throughout the cell cycle, cells in late S phase are most sensitive. Prednisone is a glucocorticosteroid. This class of drugs reduces the inßammatory response to tumor tissue by preventing white blood cells from functioning to produce swelling and pain around the tumor site. Glucocorticosteroids induce lysis of peripheral lymphocytes and slow lymphocyte production. The addition of glucocorticosteroids to chemotherapy regimens may augment their efÞcacy.

Clinical Performance. Many randomized trials have conÞrmed mitoxantrone’s palliative activity. In one study, 161 hormone-refractory, metastatic CaP patients with pain were randomized to receive either mitoxantrone 12 mg/m2 IV on day 1 of a 21-day cycle plus 5 mg of prednisone orally each day continuously, or prednisone 10 mg daily (Tannock IF, 1996). Patients who failed to respond to prednisone could switch to mitoxantrone. The primary endpoint was a reduction in pain by a scale factor of two in a six-point scale for the duration of two evaluations (which were at least three weeks apart) without an increase in analgesic use. The secondary endpoints were a reduction of 50% or more in duration of pain response and the use of analgesic medication without an increase in pain, and an increase in overall survival. Palliative responses were observed in 29% of patents treated with MP versus 12% (p = 0.01) of patients who received prednisone alone. An additional seven patients in each treatment arm were able to reduce analgesia requirements without an increase in pain symptoms. The median duration of palliation was longer in the MP arm: 43 weeks versus 18 weeks (p = 0.001). Eleven patients who were initially randomized to single-agent prednisone responded after mitoxantrone was added. There was no difference in overall survival. Treatment was well tolerated; only Þve incidences of cardiac toxicity occurred in a total of 130 patients who received mitoxantrone. Most responding patients experienced a reduction in PSA levels from base line. Nonpharmacological Approaches Watchful Waiting. Watchful waiting (no treatment other than observation with PSA testing) is a standard approach for men who choose to postpone or avoid

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treatment, with its associated side effects, until the cancer begins to affect quality of life (by causing pain or interfering with urinary function) or PSA velocity increases. Watchful waiting is appropriate for men with a life expectancy of less than ten years and slow-growing tumors (Gleason score of four or less and PSA level of 5 ng/mL or less). (See the section “Grading” in “Etiology and Pathophysiology” for further explanation of the Gleason score.) Between 1989 and 1999, nearly 700 men with clinically detected, well- or moderately well-differentiated stage T1b, T1c, or T2 CaP were randomly assigned to undergo watchful waiting or radical prostatectomy. The primary endpoint was death from CaP; secondary end points were overall mortality, metastasis-free survival, and local progression (Holmberg L, 2002). At a median follow-up time of 6.2 years, investigators observed no statistically signiÞcant difference in overall mortality, although a higher rate of death from CaP was seen in the watchful waiting group—8.9% for the watchful waiting group versus 4.6% for the prostatectomy group (p = 0.02). (Note that this study involved men with clinically detected disease. The difference in the two treatment outcomes may be less clear among men with smaller, PSA-detected tumors.) Investigators recently published updated results of this trial (Johansson JE, 2004). The majority of cancers in the studied cohort had an indolent course during the Þrst 10–15 years. Investigators found that after 15 years, local progression and aggressive metastatic disease are more likely to occur. The researchers concluded that patients with a life expectancy of 15 years or more should be treated with early radical therapy. Radical Prostatectomy. Radical prostatectomy (removal of the prostate, including the seminal vesicles [RP]) has been a treatment for CaP since the late 1800s. It is Þrst-line therapy for stage I and II CaP in otherwise healthy men younger than age 70 and may be undertaken as an elective procedure in robust older men. RP is rarely performed for advanced disease because of the difÞculty of excising enough tissue to secure negative margins. Although surgical mortality for CaP is extremely low (0.4–0.7%), side effects are common and include impotence and incontinence in up to 50% of patients. RP increased in popularity during the 1970s, 1980s, and early 1990s (Wingo PA, 2000), most likely as a result of reÞnements in surgical techniques (so-called nerve-sparing surgery) that reduced the frequency and severity of side effects. Transurethral resection of the prostate (TURP) is an alternative procedure available for older, less robust patients. Radiation Therapy External Beam Radiation Therapy. External beam radiation therapy (EBRT) is one of the few options for inoperable disease or for early-stage disease when comorbidities preclude surgery. EBRT reduces PSA to less that 1 ng/mL or halts PSA increase in 65–85% of cases (Oesterling JE, 1997). Radiation is usually delivered to the prostate gland, periprostatic tissue, and pelvic lymph nodes in Þve doses per week for six to eight weeks.

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The efÞcacy of EBRT may be improved if given in conjunction with hormonal therapy. In a study comparing EBRT alone with combined hormonal therapy and EBRT, the latter group had signiÞcantly better overall survival (79% versus 62%) at Þve years. Among survivors, Þve-year disease-free survival was also signiÞcantly higher in the EBRT/chemotherapy group (85% versus 48%) (Bolla M, 1997). In a retrospective study of 1,580 men treated with EBRT or EBRT plus adjuvant hormonal therapy, men with intermediate- and high-risk tumors receiving the adjuvant therapy had 20% and 40%, respectively, the risk of local recurrence at Þve years compared with men receiving EBRT alone (D’Amico AV, 2000). Brachytherapy. Interstitial brachytherapy is the transperineal placement of needles that contain permanent or temporary radiation implants, or “seeds,” into tumor and adjacent tissue. The surgeon places the needles generally with the guidance of an imaging modality such as magnetic resonance imaging (MRI), transrectal ultrasound (TRUS), or computed tomography (CT). The seeds (either slow-releasing iodine I-125 or fast-releasing palladium Pd-103) diffuse radiation into the adjacent tumor tissue, resulting in maximal doses of radiation to the tumor but, in most cases, minimal damage to much of the surrounding normal tissue (bladder, rectum). Brachytherapy is usually an outpatient procedure that requires only one visit to the hospital in stark contrast with EBRT’s 25–30 visits. Among patients with localized disease and low combined Gleason scores (2–4), brachytherapy has yielded results comparable to those of radiotherapy and/or prostatectomy: 93% of patients have negligible PSA at Þve years according to some studies (Wallner K, 1996). In a retrospective study of 1,870 men treated with RP, EBRT, or brachytherapy, investigators found no signiÞcant difference in PSA outcomes at Þve years for patients with low-risk disease, though patients at intermediate or high risk (Gleason sum of Þve or more) for disease progression experienced better outcomes with either RP or EBRT (D’Amico AV, 1998). In a retrospective review of 1,050 patients treated with brachytherapy between 1991 and 1999, 92% had negative biopsies at two years; here, too, results were best for patients with low Gleason scores and PSA levels (i.e., less than 10 ng/mL) at diagnosis (Sharkey J, 2000). Orchiectomy. The testes produce 85–90% of naturally occurring androgens, including testosterone; the remaining androgens are produced in the adrenal glands. Removing androgen from the system either pharmacologically (with antiandrogens or hormonal ablation) or with surgery (orchiectomy) shrinks the tumor and lowers PSA in approximately 80% of cases. However, because the resulting cell-death process fails to eliminate the entire malignant cell population (for still unknown reasons), this procedure does not result in a cure. Bilateral orchiectomy (surgical removal of both testes) is a straightforward procedure that is relatively inexpensive when compared with long-term drug therapy. However, because of the psychological impact of this procedure, it is generally reserved for highly select patient groups—those for whom permanent impotence is not

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a signiÞcant concern and those with life-threatening complications. Orchiectomy can be used as monotherapy or may be followed up with antiandrogen treatment for maximum androgen blockade. Antiandrogen Withdrawal. Despite the proven effects of hormonal therapy with CAB or an antiandrogen alone, resistance usually develops after two to three years and PSA levels begin to rise. Withdrawing antiandrogen therapy after long-term treatment is a widely adopted second-line approach that can actually lower PSA and stall disease progression (Dawson NA, 1995; Scher HI, 1993). Accordingly, antiandrogen withdrawal can be used as a second-line hormonal therapy. Twenty-Þve percent or more of patients respond, although the duration of effect is typically only a few weeks. EMERGING THERAPIES Prostate cancer (CaP) R&D is an extremely active Þeld; several diverse agents will likely be launched by the end of 2006, and more than 90 agents are in Phase II trials. The fact that the majority of patients have historically received treatment only from urologists has presented a major hurdle for drug developers because urologists typically lack the facilities to administer chemotherapy. This hurdle is set to disappear for two reasons. First, the recent FDA approval of docetaxel for advanced CaP will ensure that more patients are referred to oncologists in the hopes of realizing a modest survival gain. Second, the advent of nonhormonal oral therapies is enabling increasing numbers of ofÞce-based urologists to enter patients into clinical trials. The availability of prostate-speciÞc antigen (PSA) testing creates a development strategy dilemma for companies investigating agents to treat CaP. They may choose to test agents in late-stage disease where unmet need is greatest and regulatory requirements are less onerous. Alternatively, they may take advantage of the opportunity afforded by the availability of PSA testing and conduct trials of agents in the much larger group of patients who have the minimum residual disease burden (i.e., those with rising PSA levels). Although approval for patients with rising PSA is likely to be very lucrative, the long survival times (a median of eight years from rising PSA to death) make studies extremely long and expensive. Although a reduction in PSA has become a standard end point in late-stage CaP trials and PSA response may be a useful tool for making commercial development decisions, no agent has been approved on the basis of PSA response alone. Improvement in time to disease progression without proven overall survival beneÞt has formed the basis of drug approvals in Europe. At the American Society of Clinical Oncology (ASCO) conference in 2004, the FDA ran workshops to discuss CaP trial endpoints but the agency reportedly remains undecided whether surrogate end points will be considered adequate for drug approval. Table 6 summarizes the drug therapies in development for CaP. This section discusses the most promising agents in Phase II development and all drugs in

EMERGING THERAPIES

TABLE 6. Emerging Therapies in Development for Prostate Cancer Compound

Development Phase

Ixabepilone (BMS-247550) United States Europe Japan Patupilone (EPO-906) United States Europe Japan Satraplatin (JM-216) United States Europe Japan Irofulven (MGI-114) United States Europe Japan Atrasentan (Xinlay) United States Europe Japan Provenge (formerly APC-8015) United States Europe Japan GVAX United States Europe Japan Onyvax-P United States Europe Japan MLN-2704 (ATG-J591 DM) United States Europe Japan MLN-591 RL United States

Marketing Company

II — —

Bristol-Myers Squibb — —

II — —

Novartis — —

III III —

GPC Biotech GPC Biotech —

II II —

MGI Pharma MGI Pharma —

III — —

Abbott Laboratories — —

III — —

Dendreon — —

III — —

Cell Genesys — —

— II —

— Onyvax —

II — —

Millennium — —

II

Cornell University/BZL Biologics/ Millennium — —

Europe — Japan — Bortezomib (Velcade, PS-341, MLN-341, LDP-341) United States I/II Millennium Europe — — Japan — — Bevacizumab (Avastin) United States II Genentech/Roche Europe — — Japan — —

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TABLE 6. (continued) Compound

Development Phase

Thalidomide (Thalidomid) United States II Europe — Japan — CC-4047 (Actimid) United States II Europe — Japan — Oblimersen (Genasense, formerly G-3139) United States II Europe — Japan — Trastuzumab (Herceptin) United States II Europe — Japan — Gefitinib (Iressa, formerly ZD-1839) United States II Europe II Japan II Finasteride (Proscar) United States II Europe — Japan — Histrelin hydrogel implant (Vantas) United States R Europe — Japan —

Marketing Company Celgene — — Celgene — — Genta — — Genentech/Roche/Chugai — — AstraZeneca AstraZeneca AstraZeneca Merck Merck Merck Valera Pharmaceuticals — —

PR = Preregistered; R = Registered. Note: Development phase refers to development for prostate cancer.

Phase III or later. The single exception is DebioPharm’s PL-14, for which published data concerning its mechanism of action and activity are lacking. Cytotoxic Agents Overview. CaP has historically been perceived as chemotherapy-resistant, but the FDA’s recent approval of docetaxel (Aventis’ Taxotere) for hormonerefractory CaP has reversed that perception, and docetaxel is rapidly becoming standard of care for hormone-refractory disease. To achieve success, cytotoxic agents will need to (1) improve upon docetaxel’s toxicity proÞle and/or (2) achieve increased survival, either as a monotherapy or in combination with docetaxel. The following sections proÞle the main candidates. Other cytotoxic agents in development (all Phase II) for which few data are available include amonaÞde dihydrochloride (ChemGenex Therapeutics’ Quinamed); amonaÞde malate (Xanthus’ XanaÞde, previously XLS-001); and Cell Therapeutic’s BBR3576, an aza-anthrapyrazole.

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Mechanism of Action. Cytotoxic agents’ fundamental aim is to exert selective toxicity toward cancer cells. They do so in a variety of ways. The most successful mechanism of action for CaP is microtubule stabilization, which taxanes—most notably docetaxel—cause. Microtubule stabilization prevents the disassembly of these structures, consequently blocking cell division. Epothilones have emerged as a new class of microtubule-targeting drugs. Like taxanes, epothilones induce microtubule bundling, formation of multipolar spindles, and mitotic arrest (Lee FY, 2001). Also like taxanes, they induce microtubule polymerization by binding to microtubules and stabilizing microtubule activity. Epothilones are designed to overcome some of the problems associated with taxanes, most notably, susceptibility to resistance caused by the drug efßux protein P-glycoprotein. Other R&D strategies are focusing on such diverse mechanisms as inducing apoptosis via DNA binding (MGI Pharma’s irofulven [MGI-114]), DNA intercalation (Cell Therapeutics’ BBR-3576), and binding of platinum to DNA (GPC Biotech’s satraplatin [JM-216] and oxaliplatin [DebioPharm’s Eloxatin]). The last drug has elicited responses in heavily pretreated patients (Droz JP, 2003), but the sparse data preclude further discussion of it. Ixabepilone. Ixabepilone (Bristol-Myers Squibb’s BMS-247550, formerly NSC-710428) is an epothilone tubulin inhibitor in Phase II development for CaP and Phase III for breast cancer. An ongoing U.S. Phase II trial, which hopes to recruit 80 patients, is comparing the efÞcacy of ixabepilone versus mitoxantrone plus prednisone in patients with hormone-refractory CaP who have failed taxane therapy. A pilot study of ixabepilone plus estramustine (PÞzer’s Emcyt/Estracyt, Nihon Kayaku’s Estracyt, generics) involved 13 chemotherapy-naive patients with metastatic, hormone-refractory CaP (Smaletz O, 2003). Patients received estramustine 280 mg three times daily for Þve days plus ixabepilone at one of two dose levels: 35 mg/m2 or 40 mg/m2 . Three of six patients treated at the higher dose developed grade 4 neutropenia, making the lower dose the maximum tolerated dose. A greater than 50% PSA decline was observed in 11 of 12 evaluable patients (92%). Objective responses occurred in 57% of patients with soft tissue metastases and 40% with bone metastases. At ASCO 2004, investigators presented data from several Phase II trials. A multi-center, randomized Phase II trial involved 92 patients with chemotherapyn¨aive hormone-refractory CaP (Kelly WK, 2004). Patients received ixabepilone 35 mg/m2 by intravenous infusion over three hours every 21 days or the same dose of ixabepilone plus 280 mg estramustine three times daily for Þve days. Patients in the ixabepilone-only arm received a median of four cycles of treatment, while those in the combination arm received a median of Þve cycles. In the combination arm, 69% of patients achieved a greater than 50% decline in PSA and 44% had a partial regression of measurable disease. In the ixabepilone-only arm, 56% achieved a greater than 50% decline in PSA, while 23% had a partial response. Grade 3 or 4 febrile neutropenia occurred in 4% of patients in each arm; grade 1 or 2 neuropathy occurred in 42% of patients in the combination arm

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and 53% of patients in the ixabepilone-only arm. Grade 3 neuropathy occurred in 4% and 6% of combination-treated patients and ixabepilone-only-treated patients, respectively, and thrombosis in 5% and 2%, respectively. A retrospective evaluation presented at ASCO 2004 found no apparent crossresistance between ixabepilone and taxanes (Rosenberg JE, 2004). No difference was seen in response to second-line taxane therapy based on whether patients had received ixabepilone as part of their Þrst-line therapy. A second Phase II trial presented at ASCO 2004 investigated ixabepilone monotherapy in patients with chemotherapy-naive, hormone-refractory CaP and performance status 0–2 (Hussain M, 2004). Patients received 40 mg/m2 ixabepilone over three hours every three weeks. Forty-one patients (median age 74) received treatment. At the time of presentation, data were available for 22 patients. Forty-one percent of patients achieved a PSA response. Ten of the 22 patients had measurable disease, and 30% of these patients had an objective response (1 unconÞrmed complete response and 2 unconÞrmed partial responses). The estimated median progression-free survival was eight months, and the estimated one-year survival was 75%. The most frequent grade 3 toxicities were neutropenia (11 patients), fatigue (3 patients), and sensory neuropathy (3 patients). Grade 4 neutropenia occurred in one patient. Ixabepilone’s primary shortcomings are grade 1 and 2 neurotoxicity, which occur in about 50% of treated patients, and febrile neutropenia, which occurs infrequently but can be life-threatening. Patupilone. Patupilone (Novartis’ EPO-906), an intravenously administered formulation of epothilone B, is in Phase II trials for CaP in the United States. It is also in Phase II trials for non-small-cell lung cancer, ovarian cancer, and renal cancer. A microtubule stabilizer, patupilone is not a substrate for the multidrugresistance protein Pgp. The compound has demonstrated preclinical activity in taxane-sensitive and taxane-resistant tumors. At ASCO 2004, investigators presented Þndings of a Phase IIa trial involving 37 patients with hormone-refractory CaP with or without metastases and with PSA higher than 20 ng/mL (Hussain M, 2004). The patients received 2.5 mg patupilone weekly for three weeks followed by one week of rest for up to six cycles (with a median of three). Twenty-nine patients had received prior chemotherapy. Twenty-two percent of patients had a partial PSA response, 16% had stable disease, and 43% progressed. Four of 20 patients with measurable disease had a partial response. The most common adverse events were gastrointestinal: grade three or four diarrhea, fatigue, dehydration, abdominal pain, and vomiting were reported in 19%, 14%, 8%, 5%, and 5% of patients, respectively. Grade 2 diarrhea was reported in 19% of patients. Grade 1 or 2 peripheral neuropathy occurred in 14% and 5% of patients, respectively. Grade 2 anemia occurred in 19% of patients; no other myelosuppression was reported. Patupilone’s principal shortcoming is dose-limiting diarrhea. Neurotoxicity and myelosuppression are uncommon with this compound, unlike with ixabepilone.

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FIGURE 12. Structure of satraplatin.

Satraplatin. Satraplatin (GPC Biotech’s JM-216) (Figure 12) is a thirdgeneration oral platinum agent. Satraplatin reentered Phase III development for second-line treatment of hormone-refractory CaP in September 2003; a previous Phase III trial was halted early by the previous developer, Bristol-Myers Squibb. The ongoing Phase III trial, whose primary endpoint is time to disease progression, is known as SPARC (Satraplatin and Prednisone Against Refractory Cancer). In 2003, the FDA granted satraplatin fast-track status, and in early 2004, the Phase III trial was extended to Europe. A Phase I study protocol that will combine satraplatin with docetaxel to treat various tumor types is in preparation. Because satraplatin is orally administered and does not require hydration to prevent renal toxicity, home administration is possible. In animal models, satraplatin has demonstrated efÞcacy in tumors that are resistant to platinum agents, possibly as a result of increased lipophilicity, allowing for more passive uptake of the platinum complexes by cancer cells (Orr RM, 1994). At ASCO 2003, investigators presented Þndings from the Þrst 50 patients treated in the original Phase III trial in patients with hormone-refractory CaP (Sternberg CN, 2003). The trial was initiated by Bristol-Myers Squibb in 1998, and accrual was halted when Bristol-Myers Squibb discontinued development of the agent, although the patients were allowed to continue treatment on a compassionate-use basis. Patients received 100 mg/m2 satraplatin for Þve days plus 10 mg prednisone orally twice daily, or prednisone alone. A greater than 50% decline in PSA was seen in 8.7% of patients in the prednisone arm and in 33.3% in the satraplatin/prednisone arm (p = 0.046). Toxicity was minimal in both arms; one patient in each arm died from stomach perforation, probably related to prednisone. Median survival was 12 months in the prednisone arm and 15 months in the combination arm. The median time to disease progression was 5.2 months for the satraplatin arm versus 2.5 months for the control arm. At 6 months, 41% of patients in the satraplatin arm were progression-free versus 22% in the control arm; at 12 months, 70% of patients in the satraplatin arm were alive versus 48% in the control arm. A Phase II trial treated 39 patients with hormone-refractory CaP with 120 mg/m2 /day satraplatin for Þve of 28 days (Peereboom D, 1998). Of nine evaluable patients with measurable disease, one had a partial response and six had stable disease. Seven patients had PSA reductions of greater than 50% for a duration of 28 days; six had reductions of greater than 80%. The principle toxicities were myelosuppression, nausea, and fatigue.

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In Phase I trials, neutropenia was the dose-limiting toxicity. Other common toxicities include thrombocytopenia, severe vomiting, diarrhea, and fever. No nephro-, oto-, or neurotoxicities have been reported. Because of irregular pharmacokinetics in this trial, researchers introduced dose fractionation in subsequent trials. Irofulven. Irofulven (hydroxymethylacylfulvene, MGI Pharma’s MGI-114) is an acylfulvene in Phase II trials for CaP. MGI Pharma is investigating the agent’s activity in several tumor types as a monotherapy and in combination with other chemotherapeutic drugs. Although investigation of irofulven’s effect began in 1998, the company announced a new international, multicenter Phase II trial in April 2004. The trial will assess time to disease progression, PSA response, pain palliation, survival, and quality of life in 135 hormone-refractory CaP patients receiving one of three treatments: prednisone, prednisone/irofulven, or prednisone/irofulven/capecitabine. An ongoing Phase II program is evaluating irofulven’s activity in combination with capecitabine and a platinum agent in hormone-refractory CaP patients who have failed docetaxel-containing chemotherapy. A semisynthetic derivative of illudin S (a natural product), irofulvene is a DNA-binding agent that causes apoptosis in cancer cells. The compound inhibits DNA synthesis, inducing S-phase arrest. Its activity is independent of several common resistance mechanisms, including multidrug resistance and apoptosisinhibiting proteins. At ASCO 2004, investigators presented a summary of irofulven data from hormone-refractory CaP patients treated in two Phase II monotherapy trials and two Phase I combination trials (Cvitkovic E, 2004). In one Phase II monotherapy trial, in which irofulven was administered every other week, one of 56 evaluable patients had a complete response, eight had a partial response, and 17 achieved stable disease. One patient had a complete PSA response, eight had a partial PSA response, and 17 had stable PSA. The principal toxicities associated with irofulven were grade 3 in each case: thrombocytopenia (13% of patients), neutropenia (10%), visual disturbances (8%), vomiting (7%), and asthenia (3%). Grade 4 neutropenia occurred in 2% of patients, and grade 3 or 4 anemia in 7%. Discontinuation for toxicity was necessary in 13 of 56 patients. In a second monotherapy trial among chemotherapy-naive patients with hormone-refractory CaP, 32 patients received irofulven daily for Þve days every 28 days (Cvitkovic E, 2004). One of six patients with measurable disease achieved a partial response and another three patients achieved stable disease. Thirteen percent of patients showed a partial PSA response, and 84% had stable PSA. Of nine patients treated with irofulven/cisplatin in a Phase I study, one patient had a complete PSA response, two had partial PSA responses, and one achieved stable PSA (Cvitkovic E, 2004). In a Phase I irofulven/capecitabine combination study, one of seven patients had a complete PSA response and two had stable PSA (Cvitkovic E, 2004).

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Irofulven is limited by the fact that it must be delivered by intravenous infusion, rendering referral to a medical oncologist necessary in most markets, and by toxicities that are serious enough to require discontinuation of treatment in nearly 25% of patients. Endothelin-Receptor Antagonists Overview. Endothelin-receptor antagonists represent a new generation of oral, targeted, cytostatic agents. They have huge commercial potential if they can successfully negotiate the obstacles to development: demonstration of efÞcacy in terms of an end point that regulatory agencies deem is sufÞciently clinically relevant. Endothelins are part of a family of autocrine/paracrine peptides (ET-1, ET-2, and ET-3) that are potent vasoconstrictors. They are produced from precursor proteins upon cleavage by metalloproteinase endothelin-converting enzymes. An increase in ET-1 protein expression is found in both primary and metastatic CaP tumor sites. Atrasentan (Abbott Laboratories’ Xinlay), discussed further on, is the ETA-receptor antagonist that has progressed furthest in development for CaP. A second ET-A-receptor antagonist in development for CaP is AstraZeneca’s ZD4054, which is in Phase II trials, but the lack of published clinical data precludes further discussion of this agent. Mechanism of Action. ET-1 stimulates tumor-cell and osteoblast proliferation, inhibits apoptosis, and stimulates nociception. It can increase the mitogenic effect of other growth factors and directly stimulate growth of CaP cells. ET-1 has a synergistic effect on angiogenic factors (Pirtskhalaishvili G, 2000). It acts by binding to two receptors, ET type A (ET-A) and ET type B (ET-B). ET-A has the most afÞnity with ET-1 and is associated with the progression of CaP. ET-B mediates ET-1 clearance and may inhibit ET-1 secretion. The purpose of targeting the ET-A-receptor signaling pathway is to slow the progression of CaP. Atrasentan. Atrasentan (Abbott Laboratories’ Xinlay, formerly ABT-627) is an oral, small-molecule, selective ET-A-receptor antagonist that appears to slow the progression of CaP. It is in Phase III trials, and Abbott submitted a new drug application (NDA) for the drug with the FDA at the end of 2004. In June 2001, two 1,000-patient international Phase III trials began. Trial M00211 involved patients with metastatic, hormone-refractory CaP, and trial M00244 involved patients with nonmetastatic hormone-refractory CaP. In both trials, patients were randomized to receive 10 mg atrasentan or placebo once daily. An Independent Data Monitoring Committee found trial M00-211 failed to meet its primary endpoint of time to disease progression and stopped the trial early despite improvements in several surrogate markers. M00-244 has completed accrual and is ongoing. Meanwhile, a Phase II study (M01-366) is investigating the safety and efÞcacy of atrasentan among approximately 200 men with hormone-n¨aive CaP and rising PSA (0.4–5.0 ng/mL and a doubling time of less than one year). Abbott has been granted fast-track status for atrasentan, a designation that allows the company to Þle a rolling NDA.

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Analyses of the curtailed Phase III trial involving patients with metastatic CaP (M00-211) found that atrasentan-treated patients experienced improvements compared with those treated with placebo in time to disease progression based on bone alkaline phosphatase (BAP) and the development of symptoms (Lipton A, 2003; Yount S, 2004). In an analysis of all patients (Carducci MA, 2004), atrasentan achieved a nonsigniÞcant trend in delayed time to progression (composite measure of radiological and clinical measures) and a signiÞcant delayed time to BAP progression (505 versus 254 days), as well as signiÞcantly smaller increases than placebo-treated patients in PSA and BAP. A long-term, open-label safety evaluation of atrasentan in 220 patients with hormone-refractory CaP found that prolonged administration of atrasentan at higher doses (20 or 30 mg) was well tolerated (Eliopoulos HB, 2003). The most commonly reported adverse events for subjects receiving atrasentan versus placebo were similar: peripheral edema (40% versus 44%), bone pain (31% versus 35%), anemia (25% versus 27%), asthenia (24% versus 21%), and rhinitis (23% versus 19%). No signiÞcant hepatic or renal toxicity was associated with prolonged exposure to atrasentan at higher doses. Patients with baseline renal insufÞciency or liver function abnormalities experienced no clinically signiÞcant toxicities with prolonged exposure. A Phase II double-blind, randomized, placebo-controlled, multi-center trial involving 288 patients with asymptomatic, metastatic, hormone-refractory CaP found that 10 mg atrasentan signiÞcantly increased median time to PSA progression compared with placebo (155 days versus 71 days) (Carducci MA, 2003). The primary end point was time to disease progression. The study was powered to detect a 50% improvement in median time to progression. Treatment signiÞcantly delayed disease progression only in the evaluable patient analysis (196 days for 10 mg atrasentan versus 129 days for placebo, p = 0.029). A similar effect was found in a lower-dose atrasentan treatment group (2.5 mg atrasentan) versus placebo (184 versus 129 days, p = 0.035). In the intentto-treat analysis, the improvement in time to progression did not reach statistical signiÞcance, but time to PSA progression was statistically signiÞcant (155 days for the 10 mg atrasentan group versus 71 days for placebo). According to updated results presented at the Third Annual CaP conference in Bermuda, median survival in the evaluable patient analysis was 583 days in the atrasentan group versus 500 days in the placebo group (p < 0.05) (Carducci MA, 2003; Lee D, 2003). Only one patient discontinued therapy. An analysis of the effects of atrasentan on bone deposition and resorption markers found significant differences between atrasentan- and placebo-treated patients (Nelson JB, 2003). Patients receiving 10 mg atrasentan maintained stable mean total alkaline phosphatase and BAP values compared with baseline while subjects receiving placebo experienced a 58% elevation in the former parameter and a 99% elevation in the latter. N-telopeptide, C-telopeptide, and deoxypyridinoline elevation from baseline was consistently less in patients receiving 10 mg atrasentan compared with those receiving placebo. An analysis of pharmacokinetics and pharmacodynamics found that no dose adjustment for age or body weight is

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necessary (Carr RB, 2004). Atrasentan modestly increased plasma endothelin-1 levels. Vaccines Overview. The development of vaccines is one of the most active areas of CaP research. Each of the vaccines described in the following paragraphs has demonstrated it can induce the desired immune response. The Þrst two vaccines discussed here—Dendreon’s Provenge and Cell Genesys’s GVAX—have also shown preliminary evidence of a survival beneÞt. The third, Onyvax’s Onyvax P, has shown a beneÞt in time to disease progression in early trials. Note that Provenge and GVAX are being tested in combination with docetaxel, the most potent chemotherapeutic agent used to treat CaP. Although vaccines are being tested in late-stage disease, they may ultimately Þnd their place earlier in the course of disease in patients experiencing biochemical relapse (rising PSA), either in combination with hormone therapy or alone, thereby enabling hormone therapy to be kept in reserve. In addition, patients may potentially receive more than one vaccine, either concurrently or consecutively. The following sections discuss only vaccines that have demonstrated beneÞt in terms of PSA response. Other vaccines in development include Duke University’s RNA vaccine, which is in Phase I/II trials for CaP, and ImmunoDesigned Molecules’ IDD1, which completed Phase I/II trials in 2001. IDD1 achieved some PSA responses and caused circulating CaP cells to disappear in PCR-positive patients. Mechanism of Action. All cancer immunotherapeutic approaches use elements of the patient’s immune system to kill cancer cells, but their means of doing so varies. Approaches include isolating and modifying the patient’s own dendritic cells, creating cytokine gene-transduced vaccines (tumor cells modiÞed to secrete granulocyte-macrophage colony-stimulating factor [GM-CSF]), and employing tissue-speciÞc antigens such as PSA. Provenge. Provenge (formerly APC-8015) is a dendritic cell-based vaccine in Phase III development by Dendreon. The company prepares the vaccine by isolating dendritic cells (an important element of the immune system) from a patient’s blood and then incubating them with a fusion protein comprising prostatic acid phosphatase (PAP) and a dendritic cell-targeting element, which is structurally similar to GM-CSF. The vaccine is prepared over 48 hours. When readministered to the patient, these cells “teach” the immune system to search out and destroy PAP-producing cells. Investigations have conÞrmed that the key target for Provenge therapy—PAP—is expressed by 95% of CaP tumors (Small EJ, 2003). On the basis of Phase II trials demonstrating that Provenge is safe and well tolerated and induces antigen immunity, Dendreon initiated three Phase III trials: two trials (DD-901 and DD-902B) involve patients with hormone-refractory metastatic CaP; the third trial (P-11) involves patients with rising PSA but without signs of metastases. The ongoing DD-902B trial is designed to conÞrm DD-901’s

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Þndings that Provenge improves survival and time to onset of pain in patients with a Gleason score of 7 or less (less aggressive cancers). A National Cancer Institute (NCI)-sponsored Phase II trial (P-16) is investigating Provenge in combination with the angiogenesis inhibitor bevacizumab (Genentech/Roche’s Avastin) among patients with hormone-responsive CaP. Dendreon’s Þrst Phase III trial, DD-901, involved 127 men with metastatic, hormone-resistant CaP (Kylstra JL, 2004; Lee D, 2003). Patients were randomized in a 2:1 ratio to receive three vaccinations of Provenge or placebo every two weeks for three cycles. The primary end point was the time to objective disease progression. Median patient age was 73, median Gleason score 7, and median PSA 46. The vaccine was prepared over 48 hours. A trend toward improved time to progression was observed in the Provenge arm (p = 0.061), but researchers observed a statistically signiÞcant clinical beneÞt only after a subset analysis. Among patients with a Gleason score of seven or less (approximately 75% of hormone-resistant patients), the placebo group had a median time to progression of 9.0 weeks compared with 16.0 weeks in the Provenge-treated group (p = 0.001). Seventy-eight percent of these men responded to treatment. Patients with a Gleason score of 7 or less also had a signiÞcantly increased time to onset of pain (p = 0.016). Median time to onset of disease-related pain was 18.7 weeks for the placebo arm, while the median time was not yet reached for Provenge-treated patients. The vaccine’s effect on PSA levels was minor: only 2% of vaccine-treated patients had a 25–50% reduction in PSA levels and only 5% had a greater than 50% reduction. No placebo-treated patients experienced a PSA reduction. T-cell stimulation was seven times higher in patients with a low Gleason score versus those with a Gleason score of 8 or more. Additionally, according to a preliminary analysis of the same trial reported by Dendreon in August 2002, the subset of Provenge patients who had not progressed six months after randomization achieved a greater than eight-fold advantage in progression-free survival compared with those patients who received placebo (34.7% versus 4%). The vaccine’s common side effects were grade 1 and 2. They included rigors (60% of vaccine-treated patients versus 7% of placebo patients), pyrexia (29% versus 2%), dyspnea (17% versus 4%), and fever, which were most often infusion-related and short-lived. According to Dendreon, data from the Þnal three-year follow-up of this Phase III trial showed a statistically signiÞcant survival beneÞt in patients who were treated with Provenge, regardless of Gleason score (Dendreon, press release, October 28, 2004). Although Dendreon did not provide details describing the magnitude of the survival beneÞt, the company claims this beneÞt is greater than that observed with any type of treatment in any published Phase III study in latestage prostate cancer. At the 36-month Þnal follow up, the percentage of patients alive in the Provenge-treated group was substantially greater than the percentage of patients alive in the placebo-treated group. Preliminary survival results described in a previous press release show a beneÞt for vaccine-treated patients (Dendreon, press release, January 12, 2004). Patients

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with Gleason scores of seven or less receiving Provenge had, on average, an 89% overall increase in their survival time as compared with placebo-treated patients (log rank: p = 0.047, hazard ratio = 1.89). Median survival time for patients receiving Provenge increased by 8.4 months (30.7 months versus 22.3 months). Thirty months from randomization, the survival rate for Provenge-treated patients was 3.7 times higher than for patients receiving placebo (53% versus 14%, p = 0.001). On a crossover salvage protocol, patients on the placebo arm whose disease progressed received the vaccine for three cycles. Among patients in the placebo arm who received the vaccine as salvage therapy, mean survival time was 23.9 months versus 4.6 months for those who did not receive the salvage therapy. In a Phase II trial presented at ASCO 2003, 11 patients with nonmetastatic, hormone-responsive CaP and rising PSA (0.4–6.0 ng/mL) after radical prostatectomy or radiation therapy received Provenge and bevacizumab (Rini BI, 2003). Provenge was given intravenously on weeks 0, 2, and 4. Bevacizumab (10 mg/kg) was given intravenously on weeks 0, 2, and 4 and every two weeks thereafter until toxicity or progressive disease. The median patient age was 61, the Gleason score ranged from 5 to 8, and median baseline PSA was 1.88 ng/mL. Investigators measured T-cell proliferation and cytokine production in response to PAP and dendritic-cell costimulatory/activation marker expression. Of nine evaluable patients, only three experienced a reduction in PSA doubling time and one experienced a PSA decline approaching 50%. No patient had objective disease progression. Thirty-one patients with hormone-refractory CaP were treated with Provenge in a Phase I/II trial (Small EJ, 2000). In the Phase I portion, all patients had metastases, the median patient age was 69, the performance status was 0–1, and the median PSA was 41.3 ng/mL. Sixty-six percent of patients had received chemotherapy. In the Phase II portion, the patients did not have metastases. Twelve patients in the Phase I part of the trial received increasing doses of Provenge, and six of them received the maximum dose. Nineteen more patients subsequently received treatment at the maximum dose in the Phase II portion. No treatment-related hematologic, hepatic, or renal toxicity occurred. Fifteen percent of infusions were associated with febrile reactions—two of which were grade 3; the others were grade 1 or 2. Mild myalgias occurred in two patients and mild fatigue in one. Five patients experienced mild urinary complaints. All patients developed immune responses to the recombinant fusion protein used to prepare Provenge and 38% developed immune responses to PAP. Three patients experienced a greater than 50% reduction in PSA and three experienced PSA reductions of 25–49%. Time to disease progression correlated with the development of an immune response to PAP (34 weeks median time to progression for immune responders versus 13 weeks for nonresponders) and with the dose of dendritic cells received (median 31.7 weeks for patients receiving more than 100 × 106 cells versus 12.1 weeks for patients receiving fewer cells). Seven Phase II patients had not progressed by the end of one year of follow-up. Investigators noted that the correlation of immune response to PAP with time to progression should be

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treated with caution: disease burden or aggressiveness of disease may have been confounding factors. Provenge’s limitations include the need for intravenous administration, a requirement that is likely to restrict its administration to oncologists, and the fact that the vaccine must be prepared for each individual over a 48-hour period, a requirement that will make its manufacture expensive. GVAX. Cell Genesys’s GVAX vaccine for CaP consists of tumor cells that have been irradiated and genetically modiÞed to secrete GM-CSF, a hormone that plays a key role in stimulating the body’s immune response. Unlike Provenge, the vaccine is not patient-speciÞc and is administered by intradermal injection, making it signiÞcantly more convenient to deliver and less expensive to manufacture. GVAX has completed a Phase II trial, and the Þrst of two planned Phase III trials among patients with metastatic, hormone-refractory CaP is ongoing. The ongoing Phase III trial (Vaccine ImmunoTherapy with Allogeneic CaP Cell Lines [VITAL-1]) aims to enroll 600 chemotherapy-naive, asymptomatic patients and will compare GVAX vaccine with taxane chemotherapy to determine the survival beneÞt. Patients will receive intradermal injections of the vaccine every two weeks for up to six months. The second trial (VITAL-2) will enroll symptomatic patients and compare the effect of GVAX vaccine plus taxane chemotherapy with that of taxane monotherapy on palliation of bone pain. Both trials will enroll patients at all levels of the Gleason score. In May 2004, the FDA granted Cell Genesys a special protocol assessment (SPA) for VITAL-1. The SPA is an indication that the FDA agrees that the trial’s design is adequate to support a biologics license application (BLA) and that the study Þndings will allow a deÞnitive assessment of GVAX’s effectiveness. The trial aims to achieve a 33% improvement in overall survival. The company initiated the second trial in 2005. At ASCO 2004, investigators presented mature results from a Phase II trial of GVAX involving 80 patients with hormone-refractory CaP with evidence of metastasis to the bone and other sites (Small EJ, 2004). Patients received treatment for 24 weeks in a dose-escalation trial. Doses ranged from 100 million to 300 million cells every two or four weeks; patients in the highest-dose group also received an initial dose of 500 million cells. Vaccine-speciÞc antigenic protein antibodies and a marker of osteoclast activity (type I carboxy-terminal telopeptide [ICTP]) were measured at weeks 12 and 14. At a median of 12 months, no doselimiting toxicities had been observed and median survival had not been reached. Thirty-two percent of patients in the high-dose arm achieved PSA declines following repeat vaccinations. Of all patients, 62% had stable or declining ICTP levels, suggesting osteoclast inhibition. Bone scans showed stabilization of disease in 43% of patients; two patients had completely normal bone scans. Immunogenicity appears to be dose-dependent: 87% of the patients in the high-dose group achieved an immune response to at least one cell line, compared with 72% and 40% in the medium- and low-dose groups, respectively. Investigators presented Phase II trial data for GVAX at ASCO 2002: the data showed a dose-dependent trend toward prolonged survival and longer median

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time to disease progression (Simons J, 2002). Thirty-four patients with hormonerefractory CaP and metastatic bone disease received one of two dosing regimens of GVAX. Twenty-four patients received an initial dose of GVAX (500-millioncell prime dose) followed by 12 booster doses of 100 million cells each at two-week intervals; ten patients received the same prime dose followed by treatment with a higher dose of 300 million cells at the same intervals. At two-year follow-up, 9 of the 22 lower-dose-arm patients were alive (41%; 2 were lost to follow-up). By contrast, 7 of the 10 higher-dose-arm patients (70%) were alive at two-year follow-up. Investigators reported no dose-limiting or autoimmune toxicities. In September 2002, the company reported Þnal data from this trial: the combined median survival for both dose groups was 26.2 months. A potential survival beneÞt requires conÞrmation in a Phase III trial, but the available data compare favorably with the median survival of 18.9 months achieved by docetaxel/prednisone in the same setting. In 2002, at the Targeted Therapies First International Congress, Cell Genesys reported data from a small trial in the hormone-naive setting. Some patients experienced a reduction in PSA and bone lesions. Researchers also observed a dose-response trend in terms of overall survival. On the basis of the encouraging dose-related response data reported in the Phase II trial of GVAX, Cell Genesys initiated clinical evaluation (Phase I/II) of a second-generation, high-potency GVAX vaccine in patients with hormonerefractory CaP and metastatic bone disease. This second-generation agent secretes GM-CSF at Þve- to tenfold higher levels than its predecessor. Treatment involves one of two doses of the vaccine (100 or 200 million cells) injected monthly for six months. GVAX’s limitations include its failure to achieve an immune response in all patients and the need for intradermal or subcutaneous injection, which can cause local injection-site inßammation. Onyvax-P. Onyvax’s Onyvax-P is a whole-cell allogeneic vaccine in Phase II development in the United Kingdom for CaP. The vaccine consists of three replication-deÞcient allogeneic cell lines representing cell types from different stages of the disease—primary tumor, secondary disease, and bone metastases. Onyvax plans to start a Phase III trial in the Þrst half of 2005; manufacture of the vaccine for this trial is under way at Berna Biotech. In an open-label Phase II trial involving 26 men with hormone-refractory CaP that had not metastasized to the bone, 11 patients were vaccinated monthly for one year (Pandha HS, 2004). The proÞle of the patient group was favorable: median age was 67, median PSA at presentation was 11.1 ng/mL, and performance status was 0. Each month, patients received a mixture of the three cell lines (8 × 106 of each) supplemented with bacillus Calmette-Gu´erin (BCG) on weeks 0 and 2, followed by the three lines alone. All 11 patients experienced a prolonged reduction in PSA velocity. Median progression-free survival was 58 weeks. The most frequent side effects were fatigue and local injection-site reaction; no serious adverse effects occurred. Clinical delay in disease progression was correlated with evidence of immunological activation. Proliferation of patient T cells occurred

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after vaccination and cytokine responses were enhanced. Responding participants showed a mixed TH1- and TH2-type response rather than simply TH1 activation. Immunoconjugates Overview. Immunoconjugation is a means of delivering cytotoxic molecules to tumor cells. The effector molecules are attached to monoclonal antibodies (MAbs), which target the agent to speciÞc antigens expressed on the tumor cell. When in circulation, the linkage between the effector molecule and antibody is stable, but, once inside the cell, it breaks down and the cytotoxic molecule is released, resulting in cell death. This method allows a higher concentration of the drug to be safely administered to the patient. This section focuses on MLN-2704, a form of the anti-PSMA (prostate-speciÞc membrane antigen) MAb MLN-591 conjugated with DM1, and on MLN-591 RL, a second form of the same antibody conjugated with yttrium-90. Other MAbs in development include Medarex’s MDX-010 (formerly MDX101), a human MAb against cytotoxic T-lymphocyte antigen-4 (CTLA-4); Medarex’s MDX-214, a human MAb speciÞc to CD89; Medarex’s MDX-070 (HuRx-Prostate), a human MAb against PSMA; MedImmune’s Vitaxin, a humanized version of LM-609 that blocks the alpha-v/beta-3 integrin receptor (vitronectin) and thus inhibits angiogenesis by inducing apoptosis in newly formed blood vessels; and Cell Therapeutics/Micromet’s MT-201, a fully human IgG1 antibody that targets EpCAM. These agents are not discussed in detail here because of the lack of clinical data. Mechanism of Action. The identiÞcation of unique proteins found predominantly in prostate tissue has allowed the development of several immunotherapeutic approaches. PSA, PAP, and PSMA have all been explored as target antigens. PSMA has several potential advantages over PAP and PSA: it is a cell-surface glycoprotein rather than a secreted protein and the higher the PSMA level, the more aggressive the cancer. In other words, PSMA levels are higher in high-grade, metastatic, hormone-resistant CaP than in low-grade, localized, hormone-sensitive disease. MLN-2704. MLN-2704 (previously ATG-J591 DM) is a conjugated form of the anti-PSMA MAb MLN-591 in Phase II development for CaP by Millennium. The FDA granted MLN-2704 fast-track designation. The immunoconjugate is designed to deliver the maytansinoid antimicrotubule agent DM1 directly to CaP cells through the PSMA-targeted humanized MAb MLN-591. PSMA is a highly prostate-restricted transmembrane glycoprotein expressed on CaP epithelial cells in virtually all CaPs. PSMA expression increases as the cancer becomes more aggressive; higher Gleason score cancers and metastatic CaPs have more PSMA on their surface than do prostate-conÞned cancers. Hormone-refractory tumors have the highest expression levels. Cornell University, Millennium, and BZL Biologics are also developing MLN-591 RL, a deimmunized, radiolabeled version of the same antibody (discussed later).

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A Phase I study involving 23 men with hormone-refractory CaP demonstrated that MLN-2704 is active in the disease (Galsky MD, 2004). Patients received a single ascending dose (18–343 mg/m2 ) of MLN-2704 administered intravenously over 2.5 hours. Repeat dosing at four-week intervals was permitted once initial safety and pharmacokinetics were established. At the time of presentation, 17 patients had received more than three doses. Investigators were able to detect free DM1 in plasma and found no autoantibodies against MLN-2704, MLN591, or DM1. One patient treated at the highest dose demonstrated a sustained, greater than 50% reduction in PSA, and one patient treated at the 264 mg/m2 dose achieved a partial response, improvement in intradermal metastatic lesions, and a 70% decline in PSA lasting at least 24 weeks. Side effects included transient elevations in hepatic transaminases, infusion-related fever and chills, nausea, and fatigue. At the highest dose, a case of uncomplicated febrile neutropenia occurred. MLN-591 RL. In development by Cornell University, BZL Biologics, and Millennium, MLN-591 RL is a radiolabeled version of MLN-591, a PSMA-speciÞc MAb. It is in Phase II development. MLN-591 is engineered using a technique known as DeImmunisation. This process is designed to allow repeat dosing by preventing the induction of T-cell responses. Currently available antibodies trigger immunogenicity because they provoke T-cell responses. To date, no immune responses to MLN-591 have been reported in human subjects who have received the antibody. In a Phase I trial involving 29 patients with hormone-refractory CaP, patients received one dose of 111 indium-DOTA-J591 to measure biodistribution and pharmacokinetics, followed one week later by 90Y-J591 at one of Þve dose levels (5, 10, 15, 17.5, or 20 mCi/m2 ) (Galsky MD, 2004). Eleven patients had previously received chemotherapy. The principal toxicity was dose-related, reversible myelosuppression (predominantly thrombocytopenia). Based on imaging data, MLN-591 RL accurately targeted the tumor in 89% of the patients with bone lesions and 69% of the patients with soft tissue lesions. Uptake of 111 In-DOTAJ951 in bone marrow did not correlate with the degree of myelosuppression. At the highest-dose level, two patients developed thrombocytopenia requiring platelet transfusions, and the 17.5 mCi/m2 level was added and determined to be the maximum tolerated dose for this trial. One patient at the 15 mCi/m2 dose level with a history of venous thrombosis died after self-discontinuing coumadin. No human antihuman antibodies were seen. None of the four patients who received a second dose experienced irreversible myelosuppression (no dose-limiting toxicities). Two patients treated at the 20 mCi/m2 level experienced major (85% and 70%) reductions in PSA lasting 245 and 258 days, respectively, and both had objective responses with 90% and 40% declines, respectively, in measurable disease. Another six patients (21%) experienced a less than 50% decline in PSA or PSA stabilization. MLN-591 RL’s limitations include its expense and the inconvenience involved in radioimmunoconjugation. Indeed, the Þrst radioimmunotherapeutic approved for non-Hodgkin’s lymphomas (ibritumomab tiuxetan [Biogen Idec/Schering

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AG’s Zevalin]) suffered from very slow uptake. In addition, because both patients who experienced major reductions in PSA and had objective responses in measurable disease received a dose higher than the maximum-tolerated dose, it remains to be seen whether the therapeutic index is sufÞciently wide. If response rates in larger trials are lower than those achieved by taxane chemotherapy, then MLN-591 RL will most likely be used as a second-line therapy. If so, researchers will have to examine the effect of prior chemotherapy on the toxicities associated with this agent. Proteasome Inhibitors Overview. The proteasome is a novel target for cancer drugs, and clinical studies in myeloma have generated intense excitement in this Þeld. Because the proteasome interacts with many proteins (as described in the next section), research into the downstream consequences of drugs that inhibit this target is ongoing. Mechanism of Action. Proteasomes are enzyme complexes involved in the disposal of damaged cellular proteins and the degradation of short-lived proteins that regulate cell proliferation, apoptosis, adhesion, angiogenesis, and signal transduction. Accordingly, inhibition of proteasomes can stimulate apoptosis and suppress tumor growth and spread. Regulatory proteins degraded by proteasomes include p53, p21, p27, NF-κB, I-κB, and bcl-2. Down-regulation of the NF-kB survival pathway, activated by anthracyclines, can reverse drug resistance. In preclinical studies, cancer cells appear more susceptible than normal cells to the effects of proteasome inhibition. Bortezomib. Bortezomib (Millennium’s Velcade, formerly PS-341, MLN-341, and LDP-341) is a small-molecule proteasome inhibitor in Phase I/II trials for CaP. The agent has been approved in the United States and Europe for the treatment of multiple myeloma and is under investigation for non-Hodgkin’s lymphomas. Bortezomib has high selectivity for the proteasome over other proteases (e.g., thrombin) and has demonstrated in vitro cytotoxicity against a wide range of tumor cell lines. Bortezomib inhibits the 26S proteasome that works through multiple pathways, including pathways that inßuence apoptosis and angiogenesis. Because NF-kB transcriptionally activates bcl-2, its inhibition may induce tumor cell apoptosis or reduce bcl-2-associated drug resistance. Bortezomib also stimulates the proapoptosis gene p53 (Williams SA, 2003). A Phase I trial involving 53 patients with metastatic, hormone-refractory CaP was designed to determine the dose-limiting toxicity and maximum tolerated dose of bortezomib (Papandreou CN, 2004). The patients’ median age was 66, their median performance status was 1, and their median PSA was 42 ng/mL. Seventy-seven percent of the patients had previously received chemotherapy; the median number of prior chemotherapy regimens was two (range zero to Þve). Patients received 0.13–2.0 mg/m2 bortezomib once a week for four of every Þve weeks. Dose-limiting toxicity of diarrhea and hypotension occurred at 2.0 mg/m2 ,

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thus making 1.6 mg/m2 the maximum tolerated dose. Other side effects included fatigue, hypertension, constipation, nausea, and vomiting. Two patients achieved a PSA response and two experienced a partial response in lymph nodes. After two cycles of treatment at the 0.5 mg/m2 dose, the adenopathy in one of these patients almost completely resolved, a resolution that lasted eight months, and the patient’s PSA remained stable. Nine patients (19%) had stable PSA. Among the patients treated at close to the maximum tolerated dose, 8% experienced a PSA response and 11% a partial response in measurable lymph node disease. A Phase I/II trial involving treatment with docetaxel and bortezomib involved 31 patients with advanced, hormone-refractory CaP (Dreicer R, 2004). In 21-day cycles, patients received 40 mg/m2 docetaxel intravenously over 30 minutes on days 1 and 8 and bortezomib at 1.3 mg/m2 intravenous push on days two and nine. Median patient age was 67 and median pretreatment PSA was 270. Nineteen patients had previously received chemotherapy, of whom 14 had received taxane treatment. Grade 3 diarrhea, peripheral neuropathy, hyperglycemia, neutropenia, and fatigue were observed in less than 5% of patients. Among the evaluable patients, 36% experienced a greater than 50% decline in PSA, and most of this group had a greater than 90% decline. Seventeen percent of patients with measurable disease achieved a partial response. Researchers are evaluating a cohort of patients receiving a higher dose of bortezomib. Bortezomib’s disadvantages include limited single-agent activity as secondline therapy and signiÞcant toxicities. Several cases of vascular leak syndrome have been reported in bortezomib trials for other tumor types. Response rates in combination with docetaxel among a population that includes taxane-pretreated patients are intriguing because they suggest bortezomib may reverse taxane resistance when combined with docetaxel. Because the trial included chemotherapy-naive patients, it is not yet possible to determine the extent of any additional activity contributed by bortezomib. Angiogenesis Inhibitors Overview. Angiogenesis inhibition for the treatment of solid tumors has received a boost from the approval of bevacizumab (Genentech/Roche’s Avastin) for the treatment of metastatic colorectal cancer. Angiogenesis inhibitors in development for CaP span a wide range of classes, including MAbs, selective metalloproteinase inhibitors, and thalidomide and its derivatives. Some agents discussed here have already demonstrated modest single-agent activity; current combination trials are designed to exploit synergies with chemotherapy. Some agents have also been investigated for maintenance use. The next sections describe the most promising of the many angiogenesis inhibitors in development—bevacizumab (Genentech/Roche’s Avastin) and Celgene’s thalidomide and CC-4047 (Actimid). One agent in Phase II development that may hold promise is squalamine (Genaera’s MSI-1246), but the lack of published data precludes further discussion of it. Mechanism of Action. Most anticancer angiogenesis inhibitors under development inhibit a single antiangiogenic pathway; the most promising pathway for

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inhibition appears to be vascular endothelial growth factor (VEGF). VEGF, a multifunctional cytokine and potent permeability factor secreted in response to hypoxia, has a major angiogenesis-promoting effect. VEGF is one of the most attractive angiogenic molecule targets for inhibiting angiogenesis. However, inhibition of a single stimulatory signal is likely to have an effect only in a subset of patients and so cannot be universally applied to all cancers, especially end-stage cancers, when multiple proangiogenic factors are expressed. A more promising approach, therefore, may be to inhibit VEGF receptors—there are only two VEGF receptors versus four VEGF ligands. Also, because the VEGF receptor is expressed on normal endothelial cells, these cells have a much lower mutation rate than that of tumor cells and therefore are less likely to become resistant to a VEGF-receptor-targeted drug. In the case of some of the molecules proÞled in this section, antiangiogenesis is only one of several mechanisms of action that contributes to the antitumor effect. Bevacizumab. Bevacizumab (Genentech/Roche’s Avastin) is a recombinant humanized MAb to VEGF. The agent, which has been approved for colorectal cancer, is in Phase II development for CaP. Roche acquired worldwide rights to bevacizumab except in the United States, where Genentech retained exclusive rights. A Phase II study sought to examine whether the PSA reduction rate achieved by docetaxel plus estramustine (69% of patients achieving greater than 50% reduction) can be improved by the addition of bevacizumab (Picus J, 2003). Seventy-nine men with hormone-refractory CaP received treatment with estramustine, docetaxel, and bevacizumab as well as dexamethasone (Merck/Banyu’s Decadron, generics). Warfarin was encouraged but not required. The median PSA level was 12.8 ng/mL, the median Gleason score at diagnosis was 8, and 92% of the patients had a performance status of zero or one. At the time of presentation at the Chemotherapy Foundation Symposium XXI, 77% of the patients had achieved a PSA response and, of 34 patients with measurable disease, 44% achieved a response and 32% had stable disease. Median time to PSA progression was 10.5 months—considerably longer than the time to progression achieved by docetaxel/estramustine (six months). Adverse events included one death due to thrombosis and one perforation of the diverticulum in the sigmoid colon. Less than 3% of the men developed febrile neutropenia, and 53% of the patients developed grade 3 or 4 rapidly resolving neutropenia. Other grade 3 or 4 toxicities included thrombocytopenia (3%), fatigue (19%), constipation (6%), diarrhea (5%), arrhythmia (7%), and hypertension (4%). In a Phase II trial presented at ASCO 2003, 11 patients with nonmetastatic, hormone-responsive CaP and rising PSA (0.4–6.0 ng/mL) after radical prostatectomy or radiation therapy received Provenge and bevacizumab (Rini BI, 2003). Median patient age was 61, Gleason score ranged between Þve and eight, and median baseline PSA was 1.88 ng/mL. Provenge was given intravenously on weeks 0, 2, and 4. Bevacizumab (10 mg/kg) was given intravenously on weeks 0, 2, and 4, and every two weeks thereafter until toxicity or progressive disease. Investigators measured T-cell proliferation and cytokine production in response

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O N O

O

NH

O

FIGURE 13. Structure of thalidomide.

to PAP and dendritic-cell costimulatory/activation marker expression. Of nine evaluable patients, only 3 had a reduction in PSA doubling time and one had a PSA decline approaching 50%. No patient had objective disease progression. Thalidomide. The FDA licensed thalidomide (Celgene’s Thalidomid) (Figure 13) in 1998 for the acute treatment of erythema nodosum leprosum. It is in Phase II trials for CaP. Thalidomide has several mechanisms of action, and those mechanisms are important to its anticancer activity but they remain controversial because of the severe side effects they can trigger (discussed later). The agent possesses several immunomodulatory activities: it is potentially synergistic with GM-CSF, enhances interleukin-2 (IL-2) activity, induces and inhibits T-cell stimulation and proliferation, and suppresses TNF-alpha and downregulated IL-6. Thalidomide also inhibits angiogenesis induced by beta-Þbroblast growth factor (β-FGF) and VEGF, and it has antimetastatic properties. In a Phase II monotherapy study involving 63 patients with metastatic, hormone-refractory CaP, thalidomide demonstrated modest single-agent activity (Figg WD, 2001). Patients received 200 mg thalidomide (low-dose arm) or 200 mg escalating to 1,200 mg (high-dose arm). Twenty-seven percent of all men achieved a PSA response of at least 40%. No patient in the high-dose arm experienced a decline in PSA of 50% or more, although 18% of the patients in the low-dose arm did. Four men were maintained on thalidomide for more than 150 days. The most frequent complications were constipation, fatigue, and neurotoxicities. An open-label study of low-dose thalidomide (100 mg daily) administered for up to six months involved 20 men with hormone-refractory CaP (Drake MJ, 2003). The study found that 15% of the men achieved a greater than 50% decline in PSA sustained throughout treatment. Among the 16 men treated for at least two months, 37.5% achieved an absolute decline in PSA; median PSA fell 48%. Recent studies have combined thalidomide with other agents to exploit potential synergies and minimize the toxicities of chemotherapy and thalidomide. Table 7 presents the details of these combination trials. Thalidomide’s principal limitation is its toxicity. The most frequently reported toxicities include constipation, somnolence, thromboembolism, and neurotoxicity. In CaP patients, the most dangerous toxicity is its potential to cause venous thromboembolism (VTE). A retrospective analysis of a randomized Phase II trial

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involving 70 men with advanced, hormone-refractory CaP found that 19% of the patients receiving docetaxel and thalidomide developed VTE, compared with none in the docetaxel arm (Behrens RJ, 2003). A prospective evaluation of thalidomide-induced neuropathy found that six of 67 patients developed neuropathy (Molloy FM, 2001). Of the 67 patients initially enrolled in the study, 24 remained on treatment for three to nine months. CC-4047. CC-4047 (Celgene’s Actimid) is the lead compound in a series of thalidomide derivatives in Phase II development for CaP. A multi-center study aims to enroll 36 patients with hormone-refractory CaP. CC-4407 is also in Phase II trials for multiple myeloma and osteoarthritis. At ASCO 2004, investigators presented data from the Þrst 22 patients in the Phase II study of CC-4407 (Sison B, 2004). Patients had hormone-refractory CaP with or without radiographic evidence of disease, castrate levels of serum testosterone, performance status 0 or 1, and a maximum of one previous chemotherapy regimen. Patients received 1 mg/day CC-407 orally. Nine of 22 patients completed at least 12 weeks of therapy. PSA fell in six patients; seven were too early for evaluation. Toxicity was generally mild and included constipation, nausea, vomiting, and fatigue. Only one grade 3 toxicity occurred—nausea. One patient on therapeutic doses of coumadin died after a central nervous system bleed. The trial is ongoing. Investigators have increased the dose of CC-4407 to 2 mg per day. Antisense Therapies Overview. Despite its promise, no antisense therapy for the treatment of cancer has yet been approved. Genta (in collaboration with SanoÞ-Aventis) and Isis are the main players in antisense approaches. This section focuses on Genta’s agent, oblimersen (Genasense), because it has been the most extensively investigated for CaP. Another antisense oligonucleotide in development is Lorus Therapeutics’ GTI-2501, which is targeted to the R1 component of ribonucleotide reductase, a highly regulated enzyme in the cell cycle of mammalian cells that plays an essential role in DNA synthesis and cell proliferation. The lack of published data precludes further discussion of it. Antisense oligonucleotides are short sequences of single-stranded DNA that can bind to a speciÞc region of corresponding messenger RNA (mRNA) sequence, thereby blocking both the expression and translation of the mRNA itself and the generation of the corresponding protein encoded by the mRNA. In this way, antisense molecules can block the expression of undesirable genes and their proteins. Mechanism of Action. Oblimersen. Oblimersen (Genta’s Genasense, formerly G-3139) is an antisense oligonucleotide designed to block the production of the Bcl-2 protein from bcl-2 (a proto-oncogene). In November 2004, SanoÞ-Aventis terminated its agreement with Genta for the development of oblimersen. The drug entered Phase II

TABLE 7. Findings of Key Trials Investigating Thalidomide for Prostate Cancer Trial Reference Frank RC, 2004

Lilly M, 2004

Design

273

n = 12 Patient selection: HR CaP after HT and antiandrogen withdrawal, PS 0–2, no prior chemotherapy or angiogenesis inhibitor. Treatment: six cycles of combination treatment followed by six months thalidomide maintenance treatment. Docetaxel: 25 mg/m2 weekly every 21 days. Estramustine: 140 mg tid three days per week. Thalidomide: 100 mg or 200 mg as tolerated. n = 10 Patient selection: HR CaP Treatment: Sargramostim: (GM-CSF) 500–250 mcg daily for 14 days, then QOD for three months. Thalidomide: 200–100 mg orally daily, beginning on day 15 for three months. Note: The dose range of both drugs shown reflects protocol dose reductions due to toxicity.

Patient Characteristics

Efficacy

Toxicities (number of patients)

Comments

Median age: 76 Median PS:1 Bone metastases: 83%

67% experienced >50% PSA decline. 44% experienced >75% PSA decline. 22% experienced normalized PSA. Partial response among patients with measurable disease (five patients): 40%. Stable disease: 40%.

Grade 3 or 4: Asthenia: 2 Constipation: 1 DVT: 1

Only 2 patients entered the maintenance phase; one achieved nine months of stable disease.

N.A.

Rapid PSA response: 78%. Average PSA decrease: 50.4. Partial response among patients with measurable disease: one patient.

Grade 3 or 4: Fatigue: 2 DVT: 1 Acute MI: 1 Pleural/pericardial effusion: 1 Peripheral neuropathy: 1 Neurogenic bladder: 1

Time for PSA level to return to pretreatment levels was inversely correlated with PSA magnitude.

274

TABLE 7. (continued) Trial Reference Shevrin DH, 2003

Salimichokami M, 2003

Sarao H, 2003

Design n = 15 Patient selection: Metastatic HR CaP, normal cardiac function. Treatment: Mitoxantrone: 12 mg/m2 every three weeks. Prednisone: 10 mg/day. Thalidomide: 200 mg daily, escalating to 800 mg daily. n = 30 Patient selection: Chemotherapy-naive HR CaP Treatment: Weekly docetaxel at 35 mg/m2 for six weeks every eight weeks. Thalidomide 100 mg qd. Weekly docetaxel alone. n = 12 Patient selection: HR CaP Treatment: Paclitaxel: 80–100 mg weekly for three weeks of a 5-week cycle. Doxorubicin: 20 mg weekly for three weeks of a 5-week cycle. Thalidomide: 200–500 mg daily.

Patient Characteristics

Efficacy

Toxicities (number of patients)

N.A.

Partial response: 33%. >80% drop in PSA: three patients. >50% drop in PSA: one patient.

Median age: 65 PS: 0-1

>50% reduction in PSA Thalidomide/docetaxel arm: 66% Docetaxel arm: 32%

Median age: 65

Decrease in PSA: 90%. Grade 3 or 4: The decrease ranged from Neutropenia 0.5 to 39.5 ng/ml. (grade 3): 27% Neutropenia (grade 4): 54% Leukopenia: 63% Constipation: 12 Fatigue: 12 Nausea: 12

Comments

Grade 3 or 4: The response rate Neutropenia: 5 achieved by the Rash: 1 three-drug Venous combination is thrombosis: 1 similar to that Other: achieved by Constipation: 11 mitoxantrone/ Peripheral prednisone alone. neuropathy: 3 Somnolence: 11 Grade 3 or 4: DVT: 2 Other: Neutropenia: 2 Thrombocytopenia: 3

DVT = Deep vein thrombosis; GM-CSF = Granulocyte-macrophage colony-stimulating factor; HR = Hormone-refractory; HT = Hormone therapy; MI = Myocardial infarction; PS = Performance status; PSA = Prostate-specific antigen; qd = Once a day; QOD = Every other day; Tid = Three times daily.

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trials for CaP in 1997, but despite demonstration of activity, it has not yet entered Phase III trials. In fact, in April 2004, oblimersen received a negative recommendation for malignant melanoma from the FDA’s Oncologic Advisory Committee. The committee stated the data presented did not provide substantial evidence of oblimersen’s efÞcacy, as measured by response rate and progression-free survival, to outweigh the increased toxicity endured by the patients receiving it. Genta subsequently announced it was withdrawing the NDA for malignant melanoma. Bcl-2 is an apoptosis regulator that, when overexpressed, inhibits the process of natural cell death that should occur when cells are damaged—for example, by chemotherapy. Bcl-2 is located in the mitochondrial membrane and prevents the release of cytochrome c, protecting the cell from entering the intrinsic apoptotic pathway and promoting survival. Inhibition of Bcl-2 allows cells to progress through the cell death pathway. CaP patients with hormone-sensitive disease have low levels of bcl-2 expression, whereas patients with hormone-refractory disease have high levels of bcl-2 expression. Bcl-2 is overexpressed in virtually all hormone-refractory, metastatic CaPs. Bcl-2 overexpression also correlates with resistance to chemotherapy. Like taxanes, oblimersen interacts with Bcl-2 to overcome this mechanism of drug resistance. At ASCO 2003, investigators reported preliminary Þndings from a nonrandomized Phase II trial combining oblimersen with docetaxel to treat 29 patients with metastatic, hormone-refractory CaP (Chi KN, 2003). The median age was 66, and the median time from diagnosis to study entry was 5.8 years. Eight of the patients had undergone chemotherapy. Patients received 7 mg/kg/day oblimersen by continuous infusion over 8 days and 75 mg/m2 docetaxel IV on day 6 every 21 days until progression or toxicity. The median number of cycles of docetaxel/oblimersen was four (range one to ten); 20 patients were continuing treatment at the time of presentation. Twenty-seven percent of the patients achieved a partial response and 48% achieved a greater than 50% reduction in PSA. Grade 3–4 neutropenia occurred in 42% of patients (18% of cycles). The PSA response rate was the same as that achieved by docetaxel/prednisone (Eisenberger MA, 2004). Five patients had grade 3–4 febrile neutropenia. The most common grade 1–2 side effects were fatigue (35%) and non-neutropenic fever (31%). Because the trial was nonrandomized, it is difÞcult to distinguish the activity of oblimersen from that of docetaxel. A Phase I dose-Þnding trial investigating oblimersen in combination with mitoxantrone in 26 patients with hormone-refractory CaP found the combination was well tolerated (Chi KN, 2001). Patients were treated at seven dose levels ranging from 0.6 to 5.0 mg/kg/day by 14-day continuous infusion every 28 days and mitoxantrone from 4 mg/m2 to 12 mg/m2 as an IV bolus on day 8. Two patients experienced greater than 50% reductions in PSA, and one patient had symptomatic improvement in bone pain. The researchers did not observe any dose-limiting toxicities, and hematological toxicities (including neutropenia, thrombocytopenia, and lymphopenia) were transient. Nonhematological toxicities included fatigue, fever, nausea, arthralgias, myalgias, and transient elevations in serum creatinine, none of which were severe.

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Epidermal Growth Factor Receptor Inhibitors Overview. A vast amount of R&D has been committed to the study of epidermal growth factor receptor (EGFR) inhibition. Although several EGFR inhibitors have been launched for other solid tumor types, results for CaP have been very disappointing. The two main approaches researchers have investigated are the speciÞc inhibition of EGFR tyrosine kinase (e.g., geÞtinib [AstraZeneca’s Iressa]) and MAbs directed at the external domain of the EGFR (e.g., trastuzumab [Genentech/Roche’s Herceptin] and, initially, panitumumab). Both approaches have failed to deliver results in CaP. The following sections discuss only trastuzumab and geÞtinib because of their success in treating other tumor types. Please note that Panitumumab, has now been discontinued. One agent targeting the EGFR pathway that is in earlier-stage development is pertuzumab (Genentech/Roche/Chugai’s Omnitarg). This next-generation, humanized, anti-HER/neu MAb is in Phase II development. Enrollment for the trial, which recruited men with hormone-refractory CaP who had received a taxane or epothilone, has been completed. In a Phase I trial, pertuzumab achieved a partial response in a single patient with CaP (Agus DB, 2003). The lack of published clinical data precludes further discussion of this agent. Mechanism of Action. The precise role of EGFR inhibition is not fully understood because some EGFR inhibitors are active in EGFR-negative patients as well as in EGFR-positive patients. Conßicting results on EGFR family expression in CaP have been reported, likely because of differences in testing technologies, lack of standardization of immunohistochemical assays, or different scoring systems. Estimates of the prevalence of HER-2 (ErbB-2) range from 6% to 60% (Gray CR, 2001). Several recent studies have found evidence of a role of HER-2 in the progression of CaP to hormone-refractory status, but other studies have found very low levels of HER-2 expression even in metastatic disease (Lorenzo GD, 2003; Savinainen KJ, 2002). Estimates of the prevalence of overexpression of EGFR, also known as HER-1, are wide-ranging (40–80%). Its expression is often raised in metastases compared with its expression in the primary tumor, and expression is increased in hormonerefractory CaP. Overexpression of EGFR is associated with de-differentiation, so the expression of EGFR rises along with the Gleason score. Investigators have proposed that EGFR family receptors and androgen receptors function synergistically in the absence of androgen, suggesting cross-talk between the ErbB-2 and androgen-receptor pathways. According to recent studies, mitogen-activated protein kinase and phosphatidylinositol 3-kinase can be considered the transduction pathways (Lorenzo GD, 2003). Trastuzumab. Trastuzumab (Genentech/Roche/Chuga’s Herceptin) is already marketed in the United States and Europe for the treatment of advancedstage breast cancer. It has undergone Phase II trials in the United States for patients with recurrent or hormone-refractory CaP. Researchers hypothesize that trastuzumab blocks the tumor growth stimulus delivered via the EGFR HER-2.

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In general, studies of trastuzumab in CaP have been disappointing. Preliminary results from a Phase II trial published at ASCO 2000 found that trastuzumab alone was ineffective in HER-2-negative, androgen-independent tumors. In 2002, investigators reported results from a failed trial combining trastuzumab with the farnesyl transferase inhibitor tipifarnib (Janssen/Johnson & Johnson’s Zarnestra) (Macdonald JS, 2002). A U.S. multicenter Phase II trial set out to compare trastuzumab and docetaxel, followed by a combination of both agents, in 160 patients with hormone-refractory CaP (Lara PN, 2004). Investigators screened hormonerefractory patients for HER-2 positivity and treated HER-2-positive patients with trastuzumab (4 mg/kg IV week one, 2 mg/kg thereafter) or docetaxel (30 mg/m2 weekly for six weeks followed by a two-week break). After two eight-week cycles, nonresponding patients received the combination of trastuzumab and docetaxel. Of 100 patients screened, only seven patients had 3+ or 2+ HER-2 by immunohistochemistry (IHC), and no correlation between IHC, ßuorescent in situ hybridization (FISH), or enzyme-linked immunosorbent assay occurred. Of the seven eligible patients, only four agreed to participate, so the trial was closed early for nonfeasibility. No patient responded to trastuzumab alone. Gefitinib. GeÞtinib (AstraZeneca’s Iressa, previously ZD-1839) is a once-daily oral EGFR tyrosine kinase inhibitor that acts on a range of other kinases. It does not require high levels of EGFR (HER-1) expression to be active (Arteaga CL, 2001). GeÞtinib has been approved in Japan and the United States for non-smallcell lung cancer. It is in Phase II development for CaP. In Phase I trials, the compound showed signs of activity as a monotherapy for late-stage CaP, but the results of Phase II monotherapy trials and Phase II combination therapy trials have been disappointing. A Phase I/II trial in the United States will investigate geÞtinib in combination with the sirolimus analogue everolimus (Novartis’s Certican) to treat progressive, metastatic CaP. The Þnal analysis of a Phase II trial that randomized 58 men with hormonerefractory CaP to geÞtinib (500 mg daily) or placebo had disappointing results. Comparison of PSA slopes revealed doubling times of 3.9 months for the placebo arm and 5.0 months for the geÞtinib arm. No signiÞcant differences in progression rates, time to progression, and overall survival were observed. In 2002, at the 27th European Society of Medical Oncology Congress, the disappointing results of two Phase II trials investigating geÞtinib in combination with chemotherapy in patients with hormone-refractory CaP were presented. The Þrst, 21-patient trial evaluated geÞtinib in combination with mitoxantrone and prednisone. Patients received 12 mg/m2 mitoxantrone on day 1 of a 21-day cycle to a maximum cumulative dose of 140 mg/m2 , 10 mg/day prednisone, and 250 or 500 mg/day of geÞtinib. Five patients (two of eight patients receiving lowdose geÞtinib and three of 13 receiving high-dose geÞtinib) experienced a PSA response (a decline of at least 50% lasting for at least four weeks). The second trial evaluated geÞtinib in combination with estramustine and docetaxel in 30 chemotherapy-naive patients. Patients received up to six 21-day cycles of treatment comprising 840 mg/day estramustine on days one through

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Þve, docetaxel 60 mg/m2 on day 2 as a one-hour infusion, and geÞtinib as a oncedaily oral dose of 250 or 500 mg from day 3 onward. Four patients experienced a reduction in bone pain at each dose level, and 10 of the 30 patients demonstrated a PSA response. GeÞtinib is not associated with the side effects usually seen with chemotherapy (myelosuppression, alopecia). Its side effects in the CaP trials are consistent with those recorded in the large non-small-cell lung cancer Phase II trials (IDEAL I and II) (Fukuoka M, 2002; Kris MG, 2002). They were generally mild; the most common were grade 1–2 rash, diarrhea, pruritus, and dry skin. GeÞtinib’s principal shortcoming is its failure to demonstrate clinical beneÞt either alone or in combination with chemotherapy. Hormonal Therapies Overview. Several hormonal therapies are in development for CaP. Baxter Oncology’s D-63153 and teverelix (Ardana’s Antaralix) are decapeptide luteinizing hormone-releasing hormone (LHRH) antagonists in Phase II development. Lack of data precludes further discussion of them. Toremifene (GTx’s Acapodene) is a nonsteroidal selective estrogen-receptor modulator (SERM) in Phase IIb/III development. It is being tested for the prevention of CaP among patients with high-grade prostatic intraepithelial neoplasia and for the prevention and treatment of osteoporosis caused by adjuvant LHRH analogues. This agent falls outside the scope of this study. In late-stage development for CaP are Þnasteride. Mechanism of Action. Figure 2 illustrates the major sites of hormone blockade relevant to CaP treatment. Finasteride is a steroid analogue of testosterone that treats benign prostatic conditions by blocking the activity of the enzyme 5-alpha reductase and obstructing the conversion of testosterone to dihydrotestosterone, a hormone that plays a role in benign prostatic growth and is believed to contribute to the development of CaP. Histrelin hydrogel implant is an LHRH antagonist. Unlike LHRH analogues (e.g., goserelin, leuprorelin [leuprolide acetate]), which produce their effect by activating and then desensitizing androgen-producing cells to LHRH, histrelin hydrogel implant directly blocks the effect of the releasing hormone. Finasteride. Finasteride (Merck’s Proscar) (Figure 14) was approved in 1992 for the treatment of benign prostatic hypertrophy (BPH) and is being tested as both a preventive and a therapeutic agent for CaP. In patients with BPH, Þnasteride can reduce prostate volume by 25–30%; in preclinical experiments, it has been shown to inhibit CaP cell-line growth. The focus here is on Þnasteride as a therapy to treat CaP; discussion of it as a preventive agent falls outside the scope of this study. Finasteride is in Phase II development for the treatment of CaP. A Phase II trial involving 71 patients with biochemical recurrence of CaP after primary therapy received Þnasteride (5 mg twice daily) and ßutamide (125 mg twice daily) (Barquawi AB, 2003). At a mean of 44 months’ follow-up, 38% had

REFERENCES

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FIGURE 14. Structure of finasteride.

no evidence of PSA progression (and continued on treatment), 8% had greater than 50% reduction in PSA, and 29% had PSA progression. EfÞcacy was greatest in men who achieved a PSA nadir of 0.1 ng/mL or less after starting treatment (58% of patients). Major side effects were breast tenderness (90%), gynecomastia (72%), and gastrointestinal disturbances (22%). According to investigators, the side effects were well tolerated by most patients. Histrelin Hydrogel (now launched for Valera Pharmaceuticals). Histrelin hydrogel implant (Valera Pharmaceuticals’ Vantas, formerly SPD-424) is a 12-month LHRH analogue implant approved by the FDA for the palliative treatment of advanced prostate cancer in November 2004. A Phase III open-label trial was completed in September 2003, and in December of the same year, Valera submitted an NDA to the FDA. The original developer, Shire, retains a marketing option outside of the United States. Few data have been made publicly available regarding histrelin hydrogel implant, although company-reported early data show that its efÞcacy is comparable to that of other LHRH agonists (Valera Pharmaceuticals, September 4, 2003). Histrelin hydrogel implant presumably has the same side effects as other, marketed LHRH analogues (e.g., impotence, gynecomastia, hot ßashes). REFERENCES Abate-Shen C, Shen MM. Molecular genetics of prostate cancer. Genes and Development. 2000;14:2410–2434. Abuzallouf S, et al. Baseline staging of newly diagnosed prostate cancer: a summary of the literature. Journal of Urology. 2004;171:2122–2127. Agus DB, et al. Clinical activity in a Phase I trial of HER-2-targeted rhuMAb 2C4 (pertuzumab) in patients with advanced solid malignancies (AST). Proceedings of the American Association of Clinical Oncology Annual Meeting. 2003.Abstract 771. Ahlbom A. Cancer in twins: genetic and nongenetic familial risk factors. Journal of the National Cancer Institute. 1997;89:287–293. American Joint Committee on Cancer (AJCC). AJCC Comparison Guide: Fifth Versus Sixth Edition. www.cancerstaging.org/initiatives.html#guide. Accessed January 15, 2002.

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Arteaga CL, Johnson DH. Tyrosine kinase inhibitors-ZD1839 (Iressa). Current Opinion in Oncology. 2001;13(6):491–498. Bander NH, et al. Phase I radioimmunotherapy (RIT) trial of humanized monoclonal (MAb) antibody J591 to the extracellular domain of prostate-speciÞc membrane antigen (PSMAext) radiolabeled with 177leutetium (177Lu) in advanced prostate cancer (Pca). Proceedings of the American Society for Clinical Oncology. 2003. Abstract 1612. Barchielli A, et al. Has the PSA wave already crashed upon us? Changes in the epidemiology of prostate cancer from 1985 to 1994 in central Italy. Annals of Oncology. 1999;10:361–362. Barquawi AB, et al. Combination of low-dose ßutamide and Þnasteride for PSA-only recurrent prostate cancer after primary therapy. Urology. 2003;62(5):872–876. Barry MJ. Prostate-speciÞc-antigen testing for early diagnosis of prostate cancer. New England Journal of Medicine. 2001;344:1373–1377. Bauer JJ, et al. p53 nuclear protein expression is an independent prognostic marker in clinically localized PC patients undergoing radical prostatectomy. Clinical Cancer Research. 1995;1:1295–1300. Behrens RJ, et al. Pulmonary toxicity during prostate cancer treatment with docetaxel and thalidomide. American Journal of Therapeutics. 2003;10(3):228–232. Benson RC, Gill GM. Estramustine phosphate compared with diethylstilbestrol: a randomized, double-blind, crossover trial for stage D prostate cancer. American Journal of Clinical Oncology. 1986;9(4):341–351. Bolla M, et al. Improved survival in patients with locally advanced prostate cancer treated with radiotherapy and goserelin. New England Journal of Medicine. 1997;337(5): 295–300. Bollack C, et al. Hormonal therapy trials in prostatic cancer: an EORTC Genitourinary Group Study. American Journal of Clinical Oncology. 1988;(11 suppl 2):S156–S159. Bono AV, et al. REICAP: prospective epidemiologic study in prostate cancer in Italy. European Urology. 1996;30(suppl 1):2–6. Bookstein R, et al. Suppression of tumorigenicity of human prostate cancer cells by replacing a mutated RB gene. Science. 1990;247:712–715. Bray F, et al. Estimates of cancer incidence and mortality in Europe in 1995. European Journal of Cancer. 2002;38:99–166. Brewster DH, et al. Rising incidence of prostate cancer in Scotland: increased risk or increased detection? British Journal of Urology International . 2000;85:463–473. Carducci MA, et al. The endothelin-A receptor antagonist atrasentan (ABT-627) delays clinical progression in hormone-refractory prostate cancer: a multinational, randomized, double-blind, placebo-controlled trial. Proceedings of the American Society of Clinical Oncology Annual Meeting. 2001. Abstract 694. Carducci MA, et al. Effect of endothelin-A receptor blockade with atrasentan on tumor progression in men with hormone-refractory prostate cancer: a randomized, Phase II, placebo-controlled trial. Journal of Clinical Oncology. 2003;21(4):679–689. Carducci MA, et al. Effects of atrasentan on disease progression and biological markers in men with metastatic, hormone-refractory prostate cancer: Phase III study. Proceedings of the American Society of Clinical Oncology. 2004. Abstract 4508. Carr RB, et al. Atrasentan (ABT-627, ATN) pharmacokinetics (PK) and endothelin-1 (ET-1) pharmacodynamics (PD) in a Phase II study of patients with asymptomatic

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Weinzimer SA, et al. Transferrin is an insulin-like growth factor-binding protein-3 binding protein. Journal of Clinical Endocrinology and Metabolism. 2001;86(4):1806–1813. Whittemore AS, et al. Prostate cancer in relation to diet, physical activity, and body size in blacks, whites, and Asians in the United States and Canada. Journal of the National Cancer Institute. 1995;87:652–661. Williams SA, McConkey DJ. The proteasome inhibitor bortezomib stabilizes a novel active form of p53 in human LNCAP-PRO5 prostate cancer cells. Cancer Research. 2003;63:7338–7344. Wingo PA, et al. Patterns of inpatient surgeries for the top four cancers in the United States: National Hospital Discharge Survey, 1988–1995. Cancer Causes and Control. 2000;11(6):497–512. Witte JS, et al. Genome-wide scan for prostate cancer-aggressiveness loci. American Journal of Human Genetics. 2000;67(1):92–99. Xu J, et al. Germline mutations and sequence variants of the macrophage scavenger receptor 1 gene are associated with prostate cancer risk. Nature Genetics. 2002;32:321–325. Xue W, et al. Susceptibility to prostate cancer: interaction between genotypes at the androgen receptor and prostate-speciÞc-antigen loci. Cancer Research. 2000;60(4):839–841. Yount S, et al. Impact of atrasentan on prostate-speciÞc outcomes with hormone-refractory prostate cancer patients. Proceedings of the American Society of Clinical Oncology Annual Meeting. 2003. Abstract 4582.

Small-Cell Lung Cancer

ETIOLOGY AND PATHOPHYSIOLOGY Introduction Lung cancer is the leading cause of cancer death in men and women in the United States, and its incidence is growing throughout the world. Recent Þndings suggest that lung cancer is increasing more rapidly in women than in men (Khuder SA, 2001). Lung cancer is divided into two major types: small-cell lung cancer (SCLC) and non-small-cell lung cancer (NSCLC). SCLC is the more aggressive form of the disease and accounts for approximately 20% of all lung cancer cases. The link between SCLC and smoking is well established. Before World War I, when cigarette smoking rates were low, lung cancer was relatively rare. The genetic pathogenesis of SCLC has yet to be fully elucidated. Researchers estimate that more than 15 genetic events occur during the development of SCLC (Wistuba II, 2001). Determining the prognostic signiÞcance of each genetic alteration and designing new therapies that directly target these alterations is a signiÞcant challenge for researchers who are working to prevent SCLC and improve treatment outcomes. In general, researchers accept the hypothesis that lung cancer develops over an extended period ranging from 20 to 30 years. Pathophysiology Pathogenesis and Natural History. The 1999 World Health Organization (WHO) classiÞcation categorizes SCLC as a neuroendocrine tumor of highly Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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aggressive nature. The neuroendocrine origin of SCLC was Þrst hypothesized because of the disease’s association with paraneoplastic syndromes, a group of endocrine disorders and neuromyopathies mediated by compounds produced by or in response to a cancer. These disorders are typically caused by inappropriate secretion of peptides (e.g., hormones, growth factors, cytokines, prostaglandins) by the tumor and by normal tissues in response to antitumor antibodies. The most common paraneoplastic syndromes observed in SCLC patients are described in Table 1. Figure 1 gives a theoretical overview of the pathogenesis of SCLC. SCLC tumors express different regulatory peptides and hormones (Table 2). SCLC-released peptide growth factors play a role in regulating cell motility and the invasiveness of cancer cells; several serve as growth factors (e.g., gastrin-releasing peptide [GRP], stem cell factor [SCF], neurotensin, vasopressin, cholecystokinin, granulocyte-macrophage colony-stimulating factor [GM-CSF], insulin-like growth factor I, and transferrin). When these growth factors interact with receptors on the surface of SCLC cells, they serve an autostimulatory role for the SCLC tumors that produced them, thus supporting continued expansion of the tumor. For example, GRP binds to GRP receptors on the SCLC cell surface very soon after being released by SCLC cells. The GRP-receptor interaction causes intracellular calcium levels to rise, in turn stimulating SCLC proliferation. Preventing the action of these autostimulatory peptides may provide a possible therapeutic target. When SCLC arises, its primary location is most commonly a central bronchial site. Typically, mediastinal lymph nodes are involved with the disease at the time of diagnosis; these lymph nodes are found in the area between the left and right lungs, where the heart, trachea, and esophagus are located. Metastasis is a common and early event in the natural course of SCLC; approximately 65% of SCLC patients have metastatic disease when they are diagnosed. The most common sites of metastases are other pulmonary locations, the liver, bones, and brain. More than 50% of people with SCLC develop brain metastases at some point in their illness. Classification. ClassiÞcation of SCLC has changed over time. The WHO, for example, classiÞed the subtypes of SCLC in 1981 as oat cell type, intermediate cell type, and combined type. Because of similarities in the clinical behavior and growth characteristics of the subtypes, the International Association for the Study of Lung Cancer (IASLC) reclassiÞed SCLC in 1988 into the following subtypes: • • •

Small-cell carcinoma (oat cell cancer). Mixed small-cell/large-cell carcinoma. Combined small-cell carcinoma (SCLC combined with neoplastic squamous and/or glandular components).

The small-cell carcinoma subtype accounts for more than 90% of SCLC cases. In this classiÞcation system, the small-cell carcinoma subtype represents cancers that do not have a non-small-cell component. Both small-cell carcinoma and mixed small-cell/large-cell subtypes respond to treatment; the combined

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TABLE 1. Common Paraneoplastic Syndromes Observed in Small-Cell Lung Cancer Patients Paraneoplastic Syndrome Cancer cachexia

Causes Complex metabolic responses to a tumor

Clinical Description

Cachexia (wasting) may be the most common paraneoplastic disorder. One database of SCLC patients indicates that weight loss of more than 10% at presentation was observed in 56% of cases. Syndrome of Overproduction of SIADH develops in approximately 10% of inappropriate hormones such as SCLC patients. The syndrome is antidiuretic arginine, vasopressin, characterized by disturbances in the fluid hormone secretion and neurophysin and electrolyte balance: hyponatremia (low (SIADH) sodium levels in the blood), increased circulating plasma volume, and high sodium content and osmolarity (concentration of dissolved substances) in the urine. Hyponatremia Overproduction of atrial Hyponatremia has been observed in 13% of natriuretic peptide SCLC patients. ANP, which is known to (ANP) and vasopressin cause natriuresis (increased secretion of sodium in the urine) and hypotension, is often found at elevated levels in hyponatremic SCLC patients. Hyponatremia is also often associated with SIADH. Lambert-Eaton Autoimmune response This syndrome, arising in approximately 6% myasthenic involving of SCLC cases, can cause muscle syndrome immunoglobulin weakness (especially in the pelvis and antibodies thighs) and autonomic dysfunction (e.g., dry mouth, impotence in males). Cushing’s syndrome Ectopic secretion of Ectopic ACTH secretion occurs in up to 50% adrenocorticotropic of SCLC patients. Cushing’s syndrome, a hormone (ACTH) condition that develops secondarily to ACTH secretion, develops in approximately 5% of patients. The syndrome is characterized by low potassium levels, high sugar levels, peripheral edema (swelling of the extremities), and muscle weakness. Paraneoplastic Autoimmune response Although more often associated with SCLC cerebellar involving antineuronal than other cancers, PCD is nevertheless degeneration (PCD) antibodies rare even in SCLC cases. PCD involves pronounced neurological symptoms, including an ataxic (uncoordinated) gait, loss of coordination in the trunk and extremities, dysarthria (imperfect articulation of speech due to loss of muscular control), and nystagmus (a spasmodic, involuntary movement of the eyeball). When it does develop, PCD is usually seen before SCLC is diagnosed.

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TABLE 1. (continued) Paraneoplastic Syndrome

Causes

Clinical Description

Paraneoplastic Autoimmune response SCLC is the most common tumor associated encephalomyelitis/ involving antineuronal with these rare disorders. Symptoms can paraneoplastic antibodies include sensory deficiencies or pain, limbic sensory symptoms (e.g., confusion, anxiety, neuropathy depression), motor weakness, cerebellar dysfunction, and other neurological abnormalities. Cancer-associated Autoimmune response This retinopathy is observed in association retinopathy involving with several cancers but is linked primarily photoreceptors in the to SCLC. Photoreceptor degeneration eye leads to rapid vision loss, night blindness, and loss of color vision. Onset of retinopathy often occurs before cancer is diagnosed.

small-cell carcinoma subtype has a poorer prognosis. This simpliÞed classiÞcation of SCLC will promote consistency in diagnosis and advance researchers’ understanding of the clinical signiÞcance of the rarer SCLC subtypes. Signs and Symptoms. Only approximately 10% of SCLC patients are diagnosed before symptoms emerge. Typically, diagnosis of asymptomatic SCLC is a fortuitous occurrence associated with a routine chest X-ray or treatment for an unrelated condition. The remaining 90% of SCLC patients are symptomatic at diagnosis. Coughing, the most common symptom, is often ignored because most smokers are accustomed to coughing frequently. Most smokers visit their doctors only when they notice more severe symptoms, such as dyspnea (difÞculty breathing), wheezing, shortness of breath, hemoptysis (coughing up blood or bloody sputum), chest pain, weakness, fever, or unexplained weight loss. Disease that encroaches on mediastinal structures can lead to dysphagia (difÞculty swallowing) and superior vena cava syndrome (enlargement of the neck with venous distension caused by compression or invasion of the superior vena cava). Superior vena cava syndrome is present in approximately 10% of SCLC patients at diagnosis, but it does not inßuence a patient’s prognosis. In some patients, symptoms associated with paraneoplastic syndromes precede pulmonary symptoms and prompt patients to see a doctor. The three paraneoplastic syndromes most commonly reported in association with SCLC are syndrome of inappropriate antidiuretic hormone secretion (SIADH), Lambert-Eaton myasthenic syndrome, and Cushing’s syndrome; these syndromes occur in just 10%, 6%, and 5% of SCLC patients, respectively. Pleural effusion (accumulation of ßuid in the pleural sac surrounding each lobe of the lung) is seen in approximately 12% of lung cancer patients—more often in NSCLC than in SCLC. The presence of pleural effusion is associated with a poor prognosis.

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FIGURE 1. Theoretical overview of the pathogenesis of small-cell lung cancer.

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TABLE 2. Regulatory Peptides and Hormones Released by Small-Cell Lung Cancer Cells Adrenocorticotropic hormonea Atrial natriuretic peptidea Calcitonin Cholecystokininb Corticotrophina ß-endorphin Erythropoietina Gastrin-releasing peptideb,d Glucagon Granulocyte-macrophage colony-stimulating factorb Growth hormone ß-human chorionic gonadotropina Hepatocyte growth factorc Insulin-like growth factor Ib

Interleukin-10 Melanocyte-stimulating hormonea Neurone-specific enolased Neurophysina Neurotensinb Parathyroid hormonea Physalaemin Parathyroid hormone-related peptidea Somatostatin Stem cell factorb,c Transferrinb Transforming growth factors O and ß Vasoactive intestinal polypeptidea Vasopressina,b

a Hormone that produces documented paraneoplastic syndromes. b Growth factors that promote tumor formation. c Assists in metastatic activities, such as migration, invasion, and angiogenesis. d Serves as a potential tumor marker.

Prognostic Factors. SCLC has a poor prognosis regardless of good initial response to chemotherapy. Extensive-stage disease (ED-SCLC) has a Þve-year survival rate of 1–2% (Argiris A, 2001). In contrast, limited-stage disease (LDSCLC) generally has a Þve-year survival rate of 10–26% (Turrisi A, 1999; De Ruysscher D, 2000; Takada M, 2002; Sundstrom S, 2002). In search of potential new therapeutic targets, researchers are trying to identify genetic factors that inßuence disease survival (Table 3). Although stage of disease at diagnosis is the single most important determinant of SCLC patients’ outcome, prognosis may be modiÞed by several additional factors (Table 4). After stage of disease, performance status is probably the most signiÞcant prognostic indicator for SCLC patients. The most commonly used performance status assessment tools, worldwide, are the Karnofsky Scale (KS) and the Eastern Cooperative Oncology Group (ECOG) Scale (Table 5). One prognostic index, the Manchester Score, is a numerical expression that incorporates patient performance status and several other health performance measures (e.g., liver function, biochemical parameters). According to the Manchester Score scale, patients are considered to have a good prognosis if they have none or just one of the following features: extensive disease (ED-SCLC), poor performance status (KS), elevated alkaline phosphate, elevated lactate dehydrogenase (LDH), decreased sodium, or decreased bicarbonate. Staging. As in treatment of other types of cancers, the course of treatment for SCLC patients depends on the extent of the disease at the time of diagnosis. The Veterans Administration Lung Group (VALG) staging system classiÞes SCLC cases into two categories: limited-stage disease (LD-SCLC), which encompasses patients who can beneÞt from thoracic radiotherapy, and extensive-stage disease

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TABLE 3. Selected Genetic Mutations in Small-Cell Lung Cancer Gene/ Protein

Function

Tumor suppressor genes p53 The p53 protein normally prevents the propagation of DNA damage by causing cell arrest at the G1/S checkpoint and inducing apoptosis. Mutations in p53 remove this regulatory control, allowing mutation-carrying cells to progress through the cell cycle. An intact p53 regulatory mechanism is thought to be necessary for the activity of DNA-damaging cytotoxic agents. Rb The retinoblastoma tumor suppressor gene (Rb) encodes a protein (pRb) that acts downstream of p53, also causing cell arrest at the G1/S checkpoint. The p16 gene regulates pRb function by inhibiting CDK4 enzyme activity. Inactivation of this gene therefore has the downstream effect of disrupting pRb-mediated cell-cycle control. FHIT The fragile histidine triad (FHIT) gene is a candidate tumor suppressor gene located at chromosome 3p14.2. FHIT encodes the enzyme dinucleotide hydrolase. Decreased function or loss of this enzyme is thought to result in accumulation of diadenosine tetraphosphate, which promotes DNA replication. RASSFIA RASSFIA is a putative tumor suppressor gene on 3p21.3 and exhibits loss of heterozygosity in most lung cancers.

BAX

A protein that antagonizes the action of bcl-2 and promotes apoptosis.

Comments p53 gene is mutated in 78% of SCLC patients (Matthay RA, 1993).

The rb gene is inactivated in 90% of SCLC (Wistuba II, 2001). In contrast, the p16 gene, which is the other component of the retinoblastoma/p16 pathway, is almost never abnormal in SCLC.

Early studies of chromosomal analyses have shown that some portion of the short arm of chromosome 3 is deleted in almost 100% of SCLC patients (Matthay RA, 1993). FHIT is inactivated in 50–70% of lung cancer and is associated with smoking (Wistuba II, 2001).

There is strong evidence in SCLC: researchers found hypermethylation of RASSFIA in 20/26 (77%) SCLCs (Agathangelou A, 2001). Some, however, reported it to be hypermethylated in more than 90% of SCLC (Wistuba II, 2001). —

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TABLE 3. (continued) Gene/ Protein PTEN/MMAC1

Oncogenes Myc Family

c-kit

c-met

HER-2/neu (C-erbB-2)

Function

Comments

Phosphatase and tensin homolog deleted in chromosome 10 (PTEN), also called mutated in multiple advanced cancers (MMAC1), is a new TSG that has been located on chromosome 10q23.3 (Li J, 1997). PTEN phosphatase negatively regulates cell interactions with the extracellular matrix (Tamura M, 1998).

Loss of heterozygosity has been found in 91% of SCLC (Virmani AK, 1998). Mutations of the PTEN/MMAC1 gene were found in 11% of all lung cancers, indicating that the gene may play a role in the pathogenesis of a small subset of lung cancers (Forgacs E, 1998).

The myc family of genes (c-myc, Myc genes are amplified in 18–31% of L-myc, and N-myc) are nuclear SCLC tumors. Patients with myc transcription factors that activate amplification often have unfavorable genes that drive cell growth. prognosis; it is more frequently seen in metastatic SCLC cell lines than in primary tumors. It is also observed more frequently in chemotherapytreated patients (Wistuba II, 2001). c-kit is a receptor tyrosine kinase SCLC tumor cells are frequently found that encodes the stem cell factor to express c-kit. Mutational (SCF) receptor. The ligand of the activation of c-kit is evident in many SCF receptor, SCF, is a peptide malignancies and is associated with growth factor that stimulates cell 70% of SCLC (Dy GK, 2000). proliferation; many types of cancers, including SCLCs, produce SCF. c-kit indirectly plays an important role in modulating SCLC metastasis by virtue of its role in producing SCF receptors. c-met indirectly plays an important c-met, which codes for hepatocyte role in modulating SCLC growth factor (HGF) receptor, is metastasis because of HGF’s overexpressed in 87% of SCLC role in assisting cellular cases. Just as SCLC tumor cells migration and invasion as well as self-regulate their activities by the initiation of angiogenesis (the producing both SCF and its formation of blood vessels). receptor (c-kit), these tumor cells promote their reproduction by producing both HGF and its receptor (c-met). Receptor tyrosine kinase that is C-erbB-2 is expressed in more than involved with epithelial cell 10% of SCLC, and the expression growth and division. becomes more important in Overexpression of the growth advanced stages of the disease receptor on the surface of cells (Micke P, 2001). enhances metastatic potential.

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TABLE 3. (continued) Gene/ Protein

Function

Comments

bcl-2

The bcl-2 (B-cell lymphoma/ leukemia-2) gene encodes a protein that blocks apoptosis. Overproduction of bcl-2 allows cells to escape apoptosis.

Other Oncogenes

c-Jun is a transcription factor that Based on a limited body of evidence, belongs to the activator protein 1 cancer researchers suspect that (AP-1) family; the c-src gene c-jun and c-src genes are encodes a protein tyrosine kinase. inappropriately expressed in SCLC The c-Raf-1 protein kinase is a tumors. The c-raf-1 and c-myb major element of several signal genes might be implicated in the transduction pathways. The c-myb pathophysiology of SCLC as well. gene encodes a specific transcription factor that is expressed at high levels in immature hematopoietic cells.

Bcl-2 protein is overexpressed in 75–95% of SCLC cases (Kaiser U, 1996). Overexpression of bcl-2 is hypothesized to be a source of treatment resistance in SCLC tumors and is also suspected of facilitating the metastasis of SCLC cells. Studies show that cells with activated myc and bcl-2 oncogenes develop tumors rapidly and aggressively, while cells with activated myc oncogenes alone often die.

Other relevant events in SCLC Retinoic acid Retinoids are essential in the growth RAR-beta methylation occurs in 70% receptor beta and differentiation of normal lung of SCLC (Wistuba II, 2001). (RAR-beta) epithelial tissues. They interact with nuclear retinoid receptors (the retinoic acid receptors [RARs] and retinoid X receptors [RXRs]), both of which are involved in regulating transcription of specific genes, which, in turn, regulate cell differentiation, proliferation, and loss. Telomerase An enzyme that extends the ends of The RNA component of telomerase is telomeres that have been up-regulated in most lung cancers shortened with successive cell (98% of SCLC). This may therefore divisions. Telomerase activity is provide a target for future therapies suppressed in most normal cells (Sarvesvaran, 1999). Recent but is expressed in many tumor evidence suggests that telomerase cells. When activated, telomerase activity occurs frequently in prevents or reverses telomere smokers and appears at a very early shortening, an action that stage in lung cancer caused by probably contributes to the smoking, and that its expression immortalization of cancerous cells can be modulated by treatment with (Soria JC, 2001). a retinoid (Soria JC, 2001).

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TABLE 4. Prognostic Indicators of Small-Cell Lung Cancer Extent of disease

Pleural effusions Performance status

Gender Age Lactate dehydrogenase (LDH) concentration Neuron-specific enolase (NSE) Alkaline phosphatase concentration Albumin concentration Albumincimia concentration Sodium concentration ACTH secretion

Weight loss Myelosuppression

Patients with limited-stage disease have a better prognosis than those with extensive-stage disease. Of patients with limited-stage disease, those with very limited disease (i.e., without mediastinal lymph node involvement) have the best prognosis. Of patients with extensive stage disease, those with a single metastatic site have a better prognosis than those with multiple sites. Pleural effusions, with or without malignant cells in the effusion, indicate poor prognosis. Performance status is one of the most significant prognostic indicators for patients with limited-stage and extensive-stage disease. Patients with poor performance status have a worse prognosis than those with good performance status. For example, compared with patients with good performance status, bedridden patients do not tolerate aggressive chemotherapy well, they have increased morbidity, and they rarely survive two years after diagnosis. Women with small-cell lung cancer have a better prognosis than men with the disease. People younger than age 70 have a better prognosis than patients who are older (elderly patients often have a poor performance status). High concentrations of LDH correlate with poor prognosis.

Some studies suggest that NSE serves as a stronger prognostic indicator than LDH. Levels of NSE are higher in extensive-stage patients than in limited-stage patients. High concentrations of alkaline phosphatase correlate with poor prognosis. Low concentrations of albumin correlate with poor prognosis. High concentrations of albumincimia correlate with poor prognosis. Low concentrations of sodium correlate with poor prognosis. Patients with ectopic ACTH secretion generally have a poorer prognosis, show poor response to chemotherapy, and experience more complications during therapy. Patients who lose more than 2 kg (4.4 lb) of weight have a poor prognosis. Low hemoglobin, platelet, and white blood cell counts correlate with poor prognosis.

(ED-SCLC), which encompasses patients who cannot beneÞt from such radiotherapy. The designation LD generally means that the tumor is conÞned to the area where the tumor started; ED means that the tumor has spread to other parts of the body. The criteria for these two categories remain controversial: the VALG deÞnition of LD includes patients whose primary tumor nodal involvement is limited to one hemithorax; the IASLC recommends that LD include all patients

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TABLE 5. Performance Status Scales Karnofsky Scale: Description

Karnofsky Scale (%)

No complaints; no evidence of disease. Able to carry on normal activity; minor signs or symptoms of disease. Some signs or symptoms of disease with effort.

Cares for self; unable to do active work. Requires occasional assistance but is able to care for most personal needs. Requires considerable assistance and frequent medical care. Disabled; requires special care and assistance. Severely disabled; hospitalization indicated although death not imminent. Very sick; hospitalization necessary; requires active support treatment. Moribund; fatal processes progressing rapidly. Expired.

100

ECOG Scale: Description

ECOG Scale

Normal activity.

0

Has symptoms of disease but ambulatory and able to carry out activities of daily living.

1

Out of bed more than 50% of time; occasionally needs assistance.

2

In bed more than 50% of time; needs nursing care.

3

Bedridden; may need hospitalization.

4

90

80

70 60

50 40

30

20

10 0

ECOG = Eastern Cooperative Oncology Group.

without distant metastasis (Micke P, 2002). P. Micke and colleagues reported that the IASLC staging system for SCLC has a higher prognostic impact and is therefore preferable in clinical practice (Micke P, 2002). Most studies indicate a 90% correlation in SCLC staging when clinicians use similar diagnostic protocols. Disagreement about staging a SCLC case is most common when patients have pleural effusions. According to one interpretation of the VALG staging system, the presence of pleural effusions is a deÞning condition of ED-SCLC. Alternatively, the presence of pleural effusions is qualiÞed by its malignancy status and extent; according to this interpretation of the VALG staging system, benign pleural effusions involving only one pleural sac are consistent with LD-SCLC. When surgery is a possibility, most physicians use the American Joint Committee on Cancer (AJCC) and Union Internationale Contre le Cancer’s (UICC) agreed TNM (tumor/regional lymph nodes/distant metastasis) staging system to better characterize the cancer (Table 6). For speciÞc deÞnitions of the SCLC

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TABLE 6. AJCC/UICC TNM Staging Classification System for Small-Cell Lung Cancer AJCC/UICC Stage Occulta Stage 0 Stage I Stage II Stage III Stage IV

TNM Criteria TX, N0, M0 Tis, N0, M0 T1, N0, M0 T2, N0, M0 T3, N0, M0 T4, N0, M0 Any T, N1, M0 Any T, N2, M0 Any T, Any N, M1

a Bronchopulmonary secretions contain malignant cells on multiple samples,

but no other evidence of the primary tumor or evidence of metastasis to the regional lymph nodes or distant metastasis is found. AJCC = American Joint Committee on Cancer; UICC = Union Internationale Contra le Cancer; TNM = Tumor, node, metastasis.

TABLE 7. Definitions for the TNM Staging Classification of Small-Cell Lung Cancer TNM Categories TX T0 Tis T1 T2 T3 T4 N0 N1 N2 M0 M1

Description Tumor not assessed, or assessed but not visualized No evidence of primary tumor Carcinoma in situ Tumor invades submucosa Tumor invades muscularis propria Tumor invades through muscularis propria Tumor invades serosa, nodes, and adjacent organs No lymph node involvement One to three positive nodes More than three positive nodes No distant metastasis Distant metastasis

TNM = Tumor, node, metastasis. T = Tumor (size, extent, or depth of penetration of primary tumor); N = Node (absence or presence and extent of regional lymph node involvement); M = Metastasis.

TNM staging classiÞcation system, see Table 7. In clinical practice, the TNM system is rarely needed because most SCLC patients are clearly beyond surgical intervention when they present for diagnosis. The TNM system is used more frequently in research settings than in community practice. Etiology Risk Factors. Most known cancer risk factors can be described as processes that increase a person’s exposure to mutagens, interfere with or prevent normal DNA repair, and/or increase the proliferation rate of cells (e.g., as a result of

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FIGURE 2. Contribution of independent risk factors for small-cell lung cancer in the seven major markets.

repetitive injury and tissue repair). The principal known risk factors for SCLC are related to mutagen (or carcinogen) exposure (Figure 2). Cigarette Smoking. Cigarette smoking is by far the most important risk factor for SCLC; at least 85% of cases are attributable to this habit, and some researchers attribute 95% of all cases to this risk factor. Approximately 20% of smokers develop lung cancer, and smokers are 13 times more likely to develop lung cancer than nonsmokers (Fauci AS, 1998). Environmental tobacco smoke in the home has been found to be a major source of passive smoking; large numbers of nonsmokers are being exposed to a signiÞcantly high risk of lung cancer by family members who smoke in their homes. Of all the major subtypes of lung cancer, small-cell lung and squamous cell carcinomas are most closely linked to smoking. As the number of cigarettes smoked and the time spent smoking per day increases, lung cancer risk rises sharply. Risk of lung cancer is also related to the type of cigarette smoked; low-tar or Þltered cigarettes are associated with lower risk than high-tar or unÞltered cigarettes (Blot WJ, 1996). The greatest reduction in risk, however, comes from cessation of smoking, which results in signiÞcant decrease in all lung cancer types. A study of the effect of smoking cessation found that risk reduction was highest for small-cell lung carcinoma, followed by squamous cell carcinoma, and lowest for large-cell cancer and adenocarcinoma (Khuder SA, 2001). Heavy smokers, particularly women, beneÞt most from quitting. The percentage reduction in risk after quitting depends on the duration of time smoked; shorter-term and younger smokers experience larger relative decreases in risk. The chance that risk of lung cancers in former smokers will revert to the level of nonsmokers is small (Blot WJ, 1996). Many people who quit smoking still develop lung cancer as a result of irreparable damage that occurred during their years of smoking. In a 1995 study

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conducted by researchers at the Dana-Farber Cancer Institute and Brigham & Women’s Hospital in Boston, 51% of the 685 lung cancer patients who received treatment at these institutions between 1988 and 1994 were former smokers; 22% of these former smokers had quit smoking 20 or more years before their diagnoses. Smoking is thought to contribute to cancer development by impairing mucociliary clearance in the lungs, by decreasing immunologic response, or by causing mutations in tumor suppressor genes (TSGs). Tobacco smoke is composed of two elements: the vapor phase and the particulate phase. The particulate phase, known as “tar,” contains most of the 55 known cigarette smoke carcinogens. These carcinogens require activation by carcinogenmetabolizing enzymes, principally cytochrome P450 enzymes. In recent years, researchers have made signiÞcant progress in elucidating the mechanisms by which certain tobacco carcinogens cause genetic mutations, but the relationship between speciÞc carcinogens and speciÞc TSG or oncogene mutations remains speculative. Once metabolized, carcinogens interact with DNA. If the by-products of this interaction escape the efforts of cellular repair mechanisms, they may cause a permanent mutation. Errors that occur in certain regions of an oncogene or TSG may ultimately result in cancer. Other Sources of Exposure. Compared with smoking, other risk factors—inhalation of radon or air pollution—play a minor role in SCLC development. Nevertheless, exposure to these agents can potentiate a smoker’s risk of developing SCLC. •

•

Prior Lung Disease and Carcinogen Exposure. Lung cancer risk is also high in people with a history of prior nonmalignant lung disease—most notably, asthma, pneumonia, emphysema, and tuberculosis (Alavanja MC, 1992). Benzopyrene, a known carcinogen found in tobacco smoke, has been the subject of numerous studies. Benzopyrene is thought to cause mutations to the p53 gene, a key TSG that controls tumor growth by triggering apoptosis in abnormal cells. NNK, a tobacco-speciÞc nitrosamine has been linked with mutations in the p53 gene. More recently, researchers have suggested that aßatoxins cause lung cancer. Aßatoxins are produced by fungi that invade agricultural commodities (such as tobacco) when these plants are stored in warm, damp conditions after harvesting. These products subsequently cause mutations in the p53 gene (Lane KS, 1999). The presence of aßatoxins in foodstuffs such as peanuts, corn, and grains has been monitored by the FDA since 1966, but their contamination of tobacco is not currently regulated. Diet. Researchers have found that risk of lung cancer is high in people with high dietary intake of food rich in fat and cholesterol, including whole milk and eggs (Jain M, 1990; Shekelle RB, 1991). Evidence obtained in North and South America also suggests that consumption of alcoholic beverages, mostly beer, may be associated with increased risk of developing lung cancer

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•

•

•

SMALL-CELL LUNG CANCER

(Potter JD, 1992; Bandera EV, 1992). However, in Bandera’s study, the association was limited to heavy smokers. Radon-222 . An inert gas produced by the decay of radium-226, radon-222 is naturally present in rock and soil. As radon-222 decays, it emits highenergy alpha particles. If inhaled, these particles can disrupt DNA in the cells of the bronchial lining. Most evidence that correlates radon with lung cancer pertains to mining environments, where levels of the gas are relatively high. In a 1995 meta-analysis of 11 studies, the National Cancer Institute (NCI) determined that 40% of lung cancer deaths in uranium miners were caused by radon exposure. The NCI estimates that residential radon exposure could account for as much as 2–4% of lung cancer deaths (Lubin JH, 1995). The combination of smoking and radon increases the risk of lung cancer in a manner that is between additive and multiplicative; U.S. and U.K. research indicates that 50–75% of radon-linked lung cancer mortality is attributable to the excess risk posed by smoking (Lubin JH, 1995). Pollution. Because lung cancer incidence and mortality rates are highest in urban areas, researchers suspect that high levels of air pollution are a risk factor for lung cancer. The relationship between air pollution and the increased incidence of SCLC is unclear because of the difÞculty of removing confounding variables (i.e., other sources of carcinogen exposure) from the analysis. For instance, Cancer Incidence on Five Continents reports a 20–50% excess risk of lung cancer in urban areas of the world, while an Italian study conducted in Trieste between 1979 and 1986 concludes that air pollution increases the risk of SCLC only moderately when confounding factors (e.g., occupational exposure, smoking) are taken into account. Genetic Predisposition. Researchers remain uncertain of the role played by genetic factors in the pathogenesis of lung cancer. Glutathione S-transferases, including GSTM1, GSTP1, and GSTT1, play a role in detoxifying metabolites of carcinogens in tobacco smoke. The data available are often conßicting. While some studies suggest that some people with common polymorphisms of these genes may be more susceptible to lung cancer from exposure to environmental tobacco smoke, others found no association between GSTM1, GSTT1, or GSTP1-polymorphisms and lung cancer risk (Lewis SJ, 2002; Schneider J, 2004). Additionally, CYP3A is a P450 gene involved in tobacco carcinogen and steroid metabolism; CYP3A4*1B allele carriers were found to have signiÞcantly increased SCLC risk (Dally H, 2003). Further studies would be required to elucidate these connections.

Other Risk Factors. The following paragraphs discuss other risk factors for SCLC. •

Gender and Age. The incidence of lung cancer in men exceeds the incidence in women, usually by two-fold or more (Blot WJ, 1996). The difference is primarily due to a lower prevalence of smoking in women. This difference is

ETIOLOGY AND PATHOPHYSIOLOGY

•

•

•

307

shrinking, however, because the percentage of women who smoke is rising. Lung cancer incidence also rises progressively with age. Arsenic. Mortality and survival studies have shown increased lung cancer mortality in male workers who are heavily exposed to inorganic arsenic for a limited time; researchers found only small-cell lung carcinoma in exposed workers. A recent study found that exposure to arsenic for Þve years is associated with an increased risk of developing lung cancer (Nakadaira H, 2002). The mechanisms by which arsenic causes cancer are uncertain, but data suggest that arsenic probably causes chromosomal abnormalities that lead to cancer. Scientists agree that chronic exposure to inorganic arsenic induces a wide range of adverse health effects, including cancers of various organs. Previous studies have also found a synergistic effect between arsenic and smoking in the development of lung cancer (Buchet JP, 1998; HertzPicciotto I, 1992; Tsuda T, 1995). Asbestos. The group of Þbrous minerals known as asbestos has a long history of industrial and commercial use. When inhaled, some asbestos Þbers are phagocytized (engulfed) by cells in the lining of the airways. This phagocytosis can release free radicals capable of damaging DNA. In addition, the asbestos Þbers themselves can cause damage by physically shearing chromosomes or disrupting mitosis. In combination with smoking, exposure to asbestos is believed to increase the risk of lung cancer as much as 50-fold. Radiation. Lung cancer is one of the major effects of exposure to high doses of ionizing radiation. Because studies have found signiÞcant incidence of lung cancer in patients receiving radiation therapy, especially smokers, it is clear that smoking and radiation have a combined effect on lung cancer. However, the precise manner in which these two factors interact is less clear.

Risk Reduction Factors. A diet that is rich in fruits and vegetables may decrease the risk of developing lung cancer by imparting high levels of antioxidants, such as vitamins C and E, carotenoids, and selenium. Such antioxidants are potent scavengers of DNA-damaging free radicals and may impede the effect of carcinogens. Researchers have found that patients who exhibit a speciÞc polymorphism in the promoter region of myeloperoxidase (MPO; an enzyme that generates reactive oxygen species and free radicals that damage DNA) have lower levels of MPO expression and a reduced risk of lung cancer (Kantarci OH, 2002). Genetic Mutations. Researchers are uncertain of the role played by genetic factors in the pathogenesis of lung cancer, but studies have shown that mutations in proto-oncogenes and TSGs are critical to the development and progression of lung tumors. The inactivation of TSGs is by far the most common mutational event observed during the development of lung cancer. The molecular changes include activation of dominant oncogenes (myc family and HER-2/neu genes) as well as loss of anti-oncogenes (p53, retinoblastoma [Rb], and an unidentiÞed gene or genes on chromosome 3). However, possible loss of DNA in multiple

308

SMALL-CELL LUNG CANCER

other speciÞc sites may be present in lung cancers. Abnormalities in two particular TSGs—the Rb gene on chromosome 13 and the p53 gene on chromosome 17—are pervasive in many types of cancer. Nearly all SCLC cells have mutations in these two TSGs and/or produce abnormal Rb and p53 proteins. When either p53 or Rb gene product is missing or altered, negative checks on cell growth are deactivated; the “switch” is turned on, and uncontrolled cell proliferation ensues. The pervasiveness of these genetic abnormalities has led researchers to suspect that these genes act as “master switches” that control tumor formation. Virtually all SCLC cells have deletions in the short arm of chromosome 3 (chromosome 3p). Researchers suspect that genes in the 3p14, 3p21, and 3p24–25 regions serve as TSGs or are linked to TSGs; according to this hypothesis, deletions in these regions would permit SCLC cells to grow unchecked. Similarly, aberrations in chromosome 5 lead to nonfunctional MCC and APC genes in approximately 80% of SCLC cells (D’Amico D, 1992). These possible TSGs become inactivated through loss of heterozygosity, the usual state of having two different genetic alleles in the same location on the chromosome. Defects in chromosome 9 have been found near the site of the gene complex that encodes interferon-alpha and interferon-beta, growth factors that might play a role in SCLC pathogenesis. Table 3 brießy describes the oncogenes and TSGs implicated in SCLC and their clinical signiÞcance. The identiÞcation of genetic factors that can predict response to speciÞc therapies for SCLC and exploration of new treatment strategies could lead to individualized treatment and better results for patients with these genetic mutations. Patterns of Recurrent Disease. Although most patients diagnosed with SCLC initially beneÞt from treatment, almost all relapse with increasingly unresponsive disease. A few fortunate patients are successfully treated and live for years without recurrent disease. Even those who have “beaten” SCLC, however, are not out of the woods: SCLC patients are at increased risk of developing second cancers of the aerodigestive tract (e.g., NSCLC, esophageal cancers, head and neck cancers). For example, even SCLC patients who survive for two years after treatment have an astounding 50% chance of eventually developing a second primary tumor; overall, the risk of developing a second primary lung cancer is approximately 5% per patient per year. CURRENT THERAPIES At the time of diagnosis, small-cell lung cancer (SCLC) usually manifests as a central tumor with spread to the regional lymph nodes. The vast majority of patients also show signs of spread to distant sites. Unfortunately, despite any apparent success of local treatment (i.e., radiotherapy or surgery) in select patients, the short- and long-term prognosis for SCLC patients is very poor. Therefore, because SCLC is primarily a systemic disease, chemotherapy is the cornerstone of treatment, although radiotherapy and (to a lesser extent) surgery also play a role.

CURRENT THERAPIES

309

The majority of SCLC patients die of their tumor despite the best available treatment. Treatment is, therefore, aimed primarily at extending survival and alleviating suffering. Most improvements in survival are attributable to clinical trials that have attempted to improve on the best available accepted therapy. Patient entry into clinical studies is a common pathway to disease treatment. The Þrst small advance in the treatment of SCLC was realized in the 1960s, when cyclophosphamide (Bristol-Myers Squibb’s Cytoxan, Baxter’s Endoxan/ Endoxana, generics), one of the cytotoxic drugs then available, demonstrated a therapeutic effect. Chemotherapeutic drugs available in the 1970s for treating SCLC—such as cyclophosphamide; doxorubicin (Pharmacia’s Adriamycin/Adriblastine, generics); and epirubicin (Pharmacia’s Ellence/Farmorubicin, generics) —are near the point of replacement with more-active and/or less toxic drugs, such as the platinum-containing agents and topoisomerase inhibitors that were introduced in the 1980s. Given all the drugs now available for the treatment of SCLC, it is possible to form many different combination chemotherapy regimens. Combinations of two to four agents produce better clinical outcomes than single agents (Schuette W, 2001). Hundreds of publications have focused on many of these regimens without demonstrating any major differences in activity. However, the regimens discussed in this section are routinely used and considered standard combination chemotherapies for SCLC (Postmus PE, 1998). Typically, four to six cycles of chemotherapy are administered over a period of four to six months. If concurrent chemoradiotherapy is administered, physicians typically use four cycles because the toxic effects of concurrent thoracic radiotherapy (TRT) are greater than those of sequential TRT. Also, because tumor burden is higher in extensive-stage disease (ED-SCLC) than in limited-stage disease (LD-SCLC), some physicians administer more cycles to ED-SCLC patients. Many clinical trials have attempted to improve survival by extending chemotherapy beyond the standard maximum of six cycles (as maintenance chemotherapy), but investigators have unanimously concluded that extended chemotherapy offers no signiÞcant advantage (Postmus PE, 1998; Giaccone G, 1993). Researchers have been experimenting with higher doses of the standard dosing regimens in hopes of Þnding a signiÞcant survival beneÞt. A landmark study compared the standard EP regimen (cisplatin [Bristol-Myers Squibb’s Platinol-AQ, Nippon Kayaku’s Randa, generics] 80 mg/m2 IV on day 1; etoposide [BristolMyers Squibb’s VePesid/Etopophos, PÞzer’s Lastet, generics] 80 mg/m2 /d IV on days 1 to 3) with the same regimen at a higher dose (cisplatin 27 mg/m2 /d IV on days 1 to 5; etoposide 80 mg/m2 /d IV on days 1 to 5). Investigators found no signiÞcant differences in efÞcacy between the two regimens, but the high-dose regimen produced a signiÞcant increase in toxicity (Ihde DC, 1994). In a two-arm trial of the PCDE regimen (cisplatin, cyclophosphamide, doxorubicin, etoposide), 105 patients received the same doses of doxorubicin and etoposide throughout the six cycles of chemotherapy, but during the Þrst cycle, patients in one arm received a higher dose of cyclophosphamide and cisplatin. Investigators reported signiÞcantly higher response rates (67% versus 54%) and

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survival (43% two-year survival versus 26%) in the high-dose group (Arriagada R, 1993). Table 8 describes the regimens most commonly used for the treatment of SCLC. Etoposide/Cisplatin Regimen Overview. The etoposide (Bristol-Myers Squibb’s VePesid/Etopophos, PÞzer’s Lastet, generics) plus cisplatin (Bristol-Myers Squibb’s Platinol-AQ, Nippon Kayaku’s Randa, generics) (EP) regimen is considered the standard treatment for LD-SCLC patients who have an Eastern Cooperative Oncology Group (ECOG) performance status (PS) of 0 to 2 (Postmus P, 1998). See Table 5 for details of performance status. In most major markets, EP is used interchangeably with the carboplatin (Bristol-Myers Squibb’s Paraplatin, generics) plus etoposide (EC) regimen (discussed in the following section), but historically, EP has been the standard regimen. Both regimens are equally effective. Because EP induces nephrotoxicity and neurotoxicity, patients must be hydrated. EP causes more nausea and vomiting, but EC produces more myelosuppression. Neutropenia can be controlled with granulocyte-macrophage colony-stimulating factor (GM-CSF); thrombocytopenia is dose-limiting, and no agent is available to control this toxicity. Mechanism of Action •

•

Etoposide (Figure 3) is an epipodophyllotoxin. Epipodophyllotoxins cause single-strand breaks in DNA. Epipodophyllotoxins also inhibit DNA topoisomerase II to cause metaphase arrest, thereby inducing cytotoxic activity. Etoposide is cell-cycle-phase speciÞc, with predominant activity occurring in the late S phase and G2 phase. Cisplatin (Figure 4) is a platinum agent. Platinum agents inhibit tumor-cell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that cisplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle.

R


O O HO

O O OH O O O O CH3O

OCH3 RO

FIGURE 3. Structure of etoposide (R = H, R1 = CH3 ).

TABLE 8. Current Regimens Used for Treating Small-Cell Lung Cancer Regimen Components Regimen Etoposide/cisplatin (EP) regimen

Etoposide/carboplatin (EC) regimen

Carboplatin/etoposide/ vincristine (CEV) regimen

Agent

Availability

Grade III/IV Dose

Toxicities mg/m2 /d

Etoposide (Bristol-Myers Squibb’s VePesid/Etopophos, Pfizer’s Lastet, generics)

US, F, G, I, S, UK, J

Etoposide, 80–100 on days 1–3; cisplatin, 80–100 mg/m2 on day 1. Both IV.

Cisplatin (Bristol-Myers Squibb’s Platinol-AQ, Nippon Kayaku’s Randa, generics) Etoposide (Bristol-Myers Squibb’s VePesid/Etopophos, Pfizer’s Lastet, generics)

US, F, G, I, S, UK, J

Cycle repeated every 21 days.

US, F, G, I, S, UK, J

Etoposide, 100 mg/m2 /d days 1–3; carboplatin, 300–450 mg/m2 on day 1. Both IV.

US, F, G, I, S, UK, J

Cycle repeated every 21–28 days.

US, F, G, I, S, UK, J

Carboplatin, 300 mg/m2 on day 1; etoposide, 140 mg/m2 on days 1–3; vincristine, 1.4 mg/m2 on days 1, 8, and 15. All IV. Cycle repeated every 28 days.

Carboplatin (Bristol-Myers Squibb’s Paraplatin, generics) Carboplatin (Bristol-Myers Squibb’s Paraplatin, generics)

311

Etoposide (Bristol-Myers Squibb’s VePesid/Etopophos, Pfizer’s Lastet, generics)

US, F, G, I, S, UK, J

• Leukopenia • Anemia • Nausea/vomiting • Alopecia

• Leukopenia • Thrombocytopenia • Alopecia

• Leukopenia • Thrombocytopenia • Alopecia • Nausea/vomiting

312

TABLE 8. (continued) Regimen Components Regimen

Cyclophosphamide/ doxorubicin/vincristine (CAV) regimen or cyclophosphamide/ epirubicin/ vincristine (CEpiV) regimen

Agent

Availability

Vincristine (Eli Lilly/EG Labo/Shionogi’s Oncovin, generics) Cyclophosphamide (Bristol-Myers Squibb’s Cytoxan, Baxter’s Endoxan/Endoxana, generics)

US, F, G, I, S, UK, J

Doxorubicin (Pfizer’s Adriamycin/Adriblastine, Kyowa’s Adriacin, generics) Vincristine (Eli Lilly/EG Labo/Shionogi’s Oncovin, generics) Epirubicin (Pfizer’s Ellence/Farmorubicin, Pfizer/Kyowa’s Farmarubicin, Kenfarma’s Farmorubicina, generics)

Dose

US, F, G, I, S, UK, J

Cyclophosphamide, 800–1,000 mg/m2 ; doxorubicin, 50 mg/m2 ; vincristine, 1.4–2 mg/m2 . All IV on day 1.

US, F, G, I, S, UK, J

Substitute doxorubicin for epirubicin: 50 mg/m2 IV on day 1 for CEpiV.

US, F, G, I, S, UK, J

Cycle repeated every 21–28 days.

US, F, G, I, S, UK, J

Grade III/IV Toxicities

• Leukopenia • Thrombocytopenia • Gastrointestinal • Alopecia

TABLE 8. (continued) Regimen Components Regimen Ifosfamide/carboplatin/ etoposide (ICE) regimen or Vincristine/ ifosfamide/carboplatin/etoposide (VICE) regimen

Agent Ifosfamide (Bristol-Myers Squibb’s Ifex, Baxter’s Mitoxana/Holoxan, Shionogi’s Ifomide, generics)

Availability

Dose

US, F, G, I, S, UK, J

Ifosfamide, 5 g/m2 on day 1; carboplatin, 300 mg/m2 on day 1; etoposide, 120 mg/m2 /d on days 1 and 2. All IV. Etoposide, 240 mg/m2 orally on day 3. Plus vincristine, 0.5 mg/m2 on day 14 for VICE regimen. Cycle repeated every 28 days.

Carboplatin (Bristol-Myers Squibb’s Paraplatin, generics) Etoposide (Bristol-Myers Squibb’s VePesid/Etopophos, Pfizer’s Lastet, generics) Vincristine (Eli Lilly/EG Labo/Shionogi’s Oncovin, generics)

US, F, G, I, S, UK, J

US, F, G, I, S, UK, J

US, F, G, I, S, UK, J

Grade III/IV Toxicities • Leukopenia • Thrombocytopenia • Alopecia • Vomiting

313

314

TABLE 8. (continued) Regimen Components Regimen Cyclophosphamide/ doxorubicin/etoposide (CDE) regimen or Cisplatin/cyclophosphamide/ doxorubicin/ etoposide (PCDE) regimen

Agent

Availability

Dose

Cyclophosphamide (Bristol-Myers Squibb’s Cytoxan, Baxter’s Endoxan/Endoxana, generics)

US, F, G, I, S, UK, J

Cyclophosphamide, 1 g/m2 ; doxorubicin, 45 mg/m2 . Both IV on day 1. Etoposide, 150 mg/m2 IV on days 1 and 2. Plus cisplatin, 100 mg/m2 IV on day 1 for PCDE regimen. Cycle repeated every 21 days.

Doxorubicin (Pfizer’s Adriamycin/Adriblastine, Kyowa’s Adriacin, generics) Etoposide (Bristol-Myers Squibb’s VePesid/Etopophos, Pfizer’s Lastet, generics) Cisplatin (Bristol-Myers Squibb’s Platinol-AQ, Nippon Kayaku’s Randa, generics)

US, F, G, I, S, UK, J

US, F, G, I, S, UK, J

US, F, G, I, S, UK, J

Grade III/IV Toxicities • Leukopenia • Thrombocytopenia • Alopecia • Nausea/vomiting • Febrile neutropenia induced death, 5% in PCDE

TABLE 8. (continued) Regimen Components Regimen Cisplatin/cyclophosphamide/epirubicin/ etoposide (PCEE)

Agent

Availability

Dose

Cisplatin (Bristol-Myers Squibb’s Platinol-AQ, Nippon Kayaku’s Randa, generics)

US, F, G, I, S, UK, J

Cisplatin, 100 mg/m2 on day 2; cyclophosphamide, 400 mg/m2 /d on days 1–3; epirubicin, 40 mg/m2 on day 1; etoposide, 100 mg/m2 /d on days 1–3. All IV. Cycle repeated every 28 days.

Cyclophosphamide (Bristol-Myers Squibb’s Cytoxan, Baxter’s Endoxan/Endoxana, generics) Epirubicin (Pfizer’s Ellence/Farmorubicin, Pfizer/Kyowa’s Farmarubicin, Kenfarma’s Farmorubicina, generics)

US, F, G, I, S, UK, J

US, F, G, I, S, UK, J

Grade III/IV Toxicities • Neutropenia • Thrombocytopenia • Nausea/vomiting

315

316

TABLE 8. (continued) Regimen Components Regimen

Dose

Grade III/IV Toxicities

Agent

Availability

Cisplatin/cyclophosphamide/epirubicin/ etoposide (PCEE) (cont.) Topotecan, single agent

Etoposide (Bristol-Myers Squibb’s VePesid/Etopophos, Pfizer’s Lastet, generics) Topotecan (GlaxoSmithKline/Merck/Nihon Kayaku’s Hycamtin)

US, F, G, I, S, UK, J

US, F, G, I, S, UK, J

1.5 mg/m2 /d for 5 days per week every 21 days.

• Leukopenia • Neutropenia • Thrombocytopenia • Anemia

Etoposide, single agent

Etoposide (Bristol-Myers Squibb’s VePesid/Etopophos, Pharmacia’s Lastet, Novartis Vepeside Sandoz)

US, F, G, I, S, UK, J

100 mg orally, twice daily for five days. Cycle repeated every five days.

• Neutropenia • Infection • Nausea/vomiting

CURRENT THERAPIES

317

FIGURE 4. Structure of cisplatin.

Clinical Performance. Many trials have evaluated the EP regimen at varying dose intensities (Ihde DC, 1994; Schiller JH, 2001[a]). In an ECOG Phase III study of 402 ED-SCLC patients, EP Þrst-line treatment provided an overall response rate and an overall survival of 35% and 9.6 months, respectively. Of the patients receiving EP, 50% experienced grade 4 neutropenia and thrombocytopenia, and 5% experienced grade 4/5 infection (Schiller JH, 2001[a]). In other randomized trials, EP has demonstrated survival rates similar to those of the ECOG trial and to survival rates of the traditional CAV regimen (cyclophosphamide plus doxorubicin [PÞzer’s Adriamycin/Adriblastine, Kyowa’s Adriacin, generics] plus vincristine [Eli Lilly/EG Labo/Shionogi’s Oncovin, generics], discussed in a later section) with much higher overall response rates—more than 80% (Fukuoka M, 1991; Noda K, 2002; Sundstrom S, 2002). A Phase III trial randomized 143 SCLC patients to receive EP or EC; 82 patients had LD-SCLC and were therefore eligible for additional thoracic irradiation. For LD patients, overall response was 73% for EP and 86% for EC; for ED-SCLC patients, overall response was 50% for EP and 64% for EC. Median overall survival was 12.5 months for patients receiving EP and 11.8 months for patients receiving EC. The differences in response rates and survival were not statistically signiÞcant, but toxicity was signiÞcantly less in the EC arm, particularly gastrointestinal toxicity, nephrotoxicity, and neurotoxicity (Kosmidis PA, 1994). Compared with the CAV regimen, the EP regimen is associated with less leukopenia, neutropenia, thrombocytopenia, peripheral neuropathy, and nonhematologic toxicities. Hepatic and renal toxicities occur equally in CAV and EP regimens (Fukuoka M, 1991; Ihde DC, 1994; Schiller JH, 2001[b]). Etoposide/Carboplatin Regimen Overview. The etoposide/carboplatin (EC) regimen is generally as popular as EP in the seven major markets (United States, France, Germany, Italy, Spain, United Kingdom, and Japan). Randomized studies have shown that compared with cisplatin, carboplatin is associated with less nephrotoxicity, neurotoxicity, ototoxicity, and gastroenteric toxicity (Rozencweig M, 1990). Because its toxicity proÞle is generally considered advantageous compared with that of EP, the EC regimen is used more often than EP for poor-PS patients, the elderly, and patients with renal insufÞciencies. Unlike cisplatin, carboplatin does not require pre- or post-treatment hydration, so administration takes no longer than one hour. Therefore, EC has the additional advantage of outpatient administration; EP can also be given on an outpatient basis, but because administration takes eight hours, EP is usually given in a hospital overnight.

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SMALL-CELL LUNG CANCER

Mechanism of Action. •

•

Etoposide (Figure 3) is an epipodophyllotoxin. Epipodophyllotoxins cause single-strand breaks in DNA. Epipodophyllotoxins also inhibit DNA topoisomerase II to cause metaphase arrest, thereby inducing cytotoxic activity. Etoposide is cell-cycle-phase speciÞc, with predominant activity occurring in the late S phase and G2 phase. Carboplatin (Figure 5) is a platinum agent. Platinum agents inhibit tumorcell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that carboplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle.

Clinical Performance. Carboplatin has been investigated in combinations and compared with cisplatin combinations. A Phase III trial randomized 143 SCLC patients to receive EP or EC; 82 patients had LD-SCLC and were therefore eligible for additional thoracic irradiation. Overall response to EP was 73% for LD patients and 50% for ED patients; overall response to EC was 86% for LD patients and 64% for ED patients. Median overall survival was 12.5 months for patients receiving EP and 11.8 months for patients receiving EC. The differences in response rates and survival were not statistically signiÞcant, but toxicity was signiÞcantly less in the EC arm, particularly gastrointestinal toxicity, nephrotoxicity, and neurotoxicity (Kosmidis PA, 1994). Carboplatin/Etoposide/Vincristine Regimen Overview. The carboplatin/etoposide/vincristine (CEV) regimen is used more in Germany than anywhere else. Outside of Germany, the general consensus is that CEV is more toxic than EC, with an insigniÞcant improvement in efÞcacy. However, it is associated with less life-threatening leukopenia than CAV. Mechanism of Action. •

Carboplatin (Figure 5) is a platinum agent. Platinum agents inhibit tumorcell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that carboplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle.

FIGURE 5. Structure of carboplatin.

CURRENT THERAPIES

N

OH CH3

NH H3CO H3CO

319

N

O

H N R

H HO

CH3 OAc O

H3CO FIGURE 6. Structure of vincristine (R = CHO). •

•

Etoposide (Figure 3) is an epipodophyllotoxin. Epipodophyllotoxins cause single-strand breaks in DNA. Epipodophyllotoxins also inhibit DNA topoisomerase II to cause metaphase arrest, thereby inducing cytotoxic activity. Etoposide is cell-cycle-phase speciÞc, with predominant activity occurring in the late S phase and G2 phase. Vincristine (Figure 6) is a vinca alkaloid. Vinca alkaloids bind with microtubular proteins of the mitotic spindle, thus leading to mitotic arrest or cell death. The drug achieves its selective toxicity against cancer cells by acting on proliferating cells.

Clinical Performance. A 316-patient ED-SCLC Phase III study showed that CEV produces an overall response rate and median survival of 83% and ten months, respectively, compared with 65% and nine months, respectively, for the etoposide/vincristine (EV) regimen alone. However, the three-drug combination was associated with higher toxicities, and survival beneÞt was more evident in patients with a better PS. Grade 3/4 hematologic toxicities of the CEV regimen include leukopenia (35%), thrombocytopenia (21%), and anemia (7%); nonhematologic toxicities include alopecia (82%) and polyneuropathy (21%) (Gatzemeier U, 1994). Cyclophosphamide/Doxorubicin (Epirubicin)/Vincristine Regimen Overview. The cyclophosphamide/doxorubicin/vincristine (CAV) regimen and the cyclophosphamide/epirubicin (PÞzer’s Ellence/Farmorubicin, PÞzer/Kyowa’s Farmarubicin, Kenfarma’s Farmorubicina, generics) plus vincristine (CEpiV) regimen were some of the Þrst multidrug regimens for treatment of SCLC; they were widely used throughout the 1970s. Because doxorubicin and epirubicin are of the same class, they are sometimes used interchangeably. However, epirubicin is more expensive and still patent-protected in some territories, so it is not as commonly employed as doxorubicin. CAV was widely used throughout the

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SMALL-CELL LUNG CANCER

1970s and set the standard against which emerging combinations were measured. CAV remains in use in most countries, but its popularity has declined since the emergence of more-active, less toxic regimens in the 1980s and 1990s, such as the platinum-containing regimens. In most countries, CAV is used primarily for patients whose disease recurs after Þrst-line therapy with platinum-based regimens because CAV is non-cross-resistant with platinum. Mechanism of Action. •

•

Cyclophosphamide (Figure 7) is an alkylating agent. These agents alkylate DNA bases, thereby producing cross-links that covalently link the two DNA strands and prevent cell replication. Doxorubicin (Figure 8) and epirubicin (Figure 9) are anthracyclines. Anthracyclines interact with several different cellular targets, most importantly topoisomerase II. By inhibiting this DNA religation enzyme, anthracyclines exert their cytotoxic effect. Another mechanism leading to cell death, known as DNA intercalation, involves insertion of the anthracycline molecule between base pairs. This phenomenon causes single- and double-stranded

FIGURE 7. Structure of cyclophosphamide.

FIGURE 8. Structure of doxorubicin (R = OCH3 , R1 = OH, R2 = H, R3 = H, R4 = OH).

CURRENT THERAPIES

O

OH

321

O OH OH

H3CO

O H3C HO

OH

O

O NH2

FIGURE 9. Structure of epirubicin.

•

breaks in DNA that inhibit cell proliferation. The free radicals generated by the reductive metabolism of anthracyclines may also damage cellular structures. Vincristine (Figure 6) is a vinca alkaloid. Vinca alkaloids bind with microtubular proteins of the mitotic spindle, thus leading to mitotic arrest or cell death. The drug achieves its selective toxicity against cancer cells by acting on proliferating cells.

Clinical Performance. CAV and, to a lesser extent, CEpiV were the standard Þrst-line treatments in the 1970s and 1980s. More recently, randomized clinical trial data have led to their replacement by newer-generation, platinumbased regimens and relegation to second-line treatment. A 288-patient, three-arm study compared CAV with EP and with CAV/EP as Þrst-line treatments. CAV yielded overall response rates of 51% in LD-SCLC patients and 59% in ED-SCLC patients, compared with 77% and 78% for EP and 88% and 63% for CAV/EP, respectively (Fukuoka M, 1991). The overall response rates were signiÞcantly higher in the EP and CAV/EP arms than in the CAV arm. The CAV, EP, and CAV/EP regimens demonstrated median survival of 12.4, 11.7, and 16.8 months, respectively, in patients with LD-SCLC and 8.7, 8.3, and 9.0 months, respectively, in patients with ED-SCLC. Median overall survival was signiÞcantly greater for LD-SCLC patients treated with CAV/EP than for the other two arms, but no signiÞcant difference was evident in ED-SCLC patients. Grade 3/4 leukopenia occurred more frequently in the CAV (78%) and CAV/EP (72%) arms than in the EP arm (46%), but no signiÞcant differences were observed in thrombocytopenia. A Phase III clinical study randomized 211 patients whose disease recurred at least 60 days after Þrst-line treatment to receive either CAV or single-agent topotecan (GlaxoSmithKline/Merck/Nihon Kayaku’s Hycamtin) (von Pawel J, 1999). Both regimens are popular second-line therapies. Overall response rates in the CAV and topotecan arms were 18.3% and 24.3%, respectively. Overall survival was not statistically different between patients on the CAV regimen (24.7 weeks) and patients on the topotecan regimen (25 weeks). Symptom improvement was reportedly greater in the topotecan group than in the CAV group for four

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out of eight symptoms evaluated, including dyspnea, anorexia, hoarseness, and fatigue. Grade 4 neutropenia occurred in 37.8% of patients in the topotecan arm and 51.4% of patients in the CAV arm. Grade 4 thrombocytopenia and grade 3/4 anemia occurred more frequently with topotecan than with CAV (9.8% and 17.7% versus 1.4% and 7.2%, respectively). Nonhematologic toxicities were mild and similar in both arms. Comparable results were seen in a 436-patient, randomized trial of CEpiV versus EP (Sundstrom S, 2002). The two- and Þve-year survival rates in the EP arm (14% and 5%) were signiÞcantly higher compared with those in the CEpiV arm (6% and 2%). For LD-SCLC patients, the median survival time of 14.5 months in the EP arm was signiÞcantly greater than the median survival time of 9.7 months in the CEpiV arm. The two- and Þve-year survival rates of 25% and 10% in LD-SCLC patients in the EP arm were also signiÞcantly greater than the rates of 8% and 3% in the CEpiV arm. For ED-SCLC patients, no signiÞcant survival difference between the treatment arms was evident. Median overall survival for both ED- and LD-SCLC patients combined was 10.2 months in the EP arm, which was signiÞcantly greater than the 7.8 months observed in the CEpiV arm. Hematologic toxicities are considered a major concern with the CAV regimen; characteristic toxicities are grade 3/4 neutropenia (approximately 85%), leukopenia (approximately 80%), thrombocytopenia (approximately 15%), and anemia (approximately 20%). Nonhematologic toxicities such as alopecia, fatigue, and gastrointestinal disturbances are also common side effects of the CAV regimen (Fukuoka M, 1991). (Vincristine)/Ifosfamide/Carboplatin/Etoposide Regimen Overview. The ifosfamide (Bristol-Myers Squibb’s Ifex, Baxter’s Mitoxana/ Holoxan, Shionogi’s Ifomide, generics) plus carboplatin plus etoposide (ICE) regimen and the vincristine plus ICE (VICE) regimen are used infrequently in clinical practice. ICE and (to a lesser extent) VICE are still occasionally used in the United Kingdom, however, where these regimens were pioneered. The dosing schedules for ICE and VICE are identical, except that the latter includes a midcycle dose of vincristine. Theoretically, VICE should be more effective than ICE; researchers argue that adding vincristine in midcycle prevents relapses between cycles. No randomized trials have been conducted to test this supposition. Some evidence suggests that these combinations offer better survival than other regimens (Lorigan P, 1995), but ICE and VICE produce signiÞcant toxicity, are more difÞcult to administer than other regimens (ifosfamide must be coadministered with an inert compound called mesna to prevent hemorrhagic cystitis), and are costly. Both regimens are extremely myelosuppressive; efforts to reduce myelosuppression by using stem cell support have not been successful (Steward WP, 1998). Because of their toxicity, ifosfamide-based regimens are used principally in patients with strong PS scores and good Þtness.

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FIGURE 10. Structure of ifosfamide.

Mechanism of Action •

•

•

•

Ifosfamide (Figure 10) is an alkylating agent. These agents alkylate DNA bases, thereby producing cross-links that covalently link the two DNA strands and prevent cell replication. Carboplatin (Figure 5) is a platinum agent. Platinum agents inhibit tumorcell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that carboplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle. Etoposide (Figure 3) is an epipodophyllotoxin. Epipodophyllotoxins act by causing single-strand breaks in DNA. Epipodophyllotoxins also inhibit DNA topoisomerase II to cause metaphase arrest, thereby inducing cytotoxic activity. Etoposide is cell-cycle-phase speciÞc, with predominant activity occurring in the late S phase and G2 phase. Vincristine (Figure 6) is a vinca alkaloid. Vinca alkaloids act by binding with microtubular proteins of the mitotic spindle, thus leading to mitotic arrest or cell death. The drug achieves its selective toxicity against cancer cells by acting on proliferating cells.

Clinical Performance. The exact dose intensity of VICE was the subject of a European trial that randomized 300 ED-SCLC patients to receive VICE in three-weekly cycles (dose-intense regimen) versus four-weekly cycles (standardintensity regimen) (Steward WP, 1998). The two-by-two factorial design also allowed evaluation of the effect of granulocyte-macrophage colony-stimulating factor (GM-CSF) on reduction of the myelosuppression associated with VICE. The overall response rate was 83% (51% complete response) for all treated patients, with no signiÞcant difference between treatment groups. Median overall survival and two-year survival rates were signiÞcantly increased in the intensiÞeddose arm (443 days and 33%) versus the standard-dose arm (351 days and 18%). GM-CSF did not reduce the incidence of myelosuppression. Neutropenia occurred in 54% of all patients, with no signiÞcant difference between treatment groups. Thirty treatment-related deaths (10%) occurred; the most frequent cause of death was infection, which accounted for ten deaths. A similar but smaller trial evaluated ICE in 50 ED-SCLC patients; patients were randomized to receive a cycle of ICE either every two weeks (dose-intense)

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or every four weeks (standard dose). IntensiÞed ICE was given with GM-CSF (Woll PJ, 2001); this study did support the use of GM-CSF to reduce myelosuppression. Febrile neutropenia was more common in the standard-dose arm (84%) than in the intensiÞed-dose arm (56%). However, overall response rate and median overall survival were similar in the dose-intense and standard-dose arms (80% versus 76% and 327 days versus 272, respectively), although the study was not powered enough to detect any potential differences. (Cisplatin)/Cyclophosphamide/Doxorubicin/Etoposide Regimen Overview. The cyclophosphamide/doxorubicin/etoposide (CDE) regimen is widely used internationally to treat SCLC, although associated neutropenia prevents it from being given at full doses. However, as with all nonplatinum-based regimens, its use has declined since the emergence of data demonstrating the superiority of platinum-containing regimens. Nevertheless, CDE’s non-cross-resistant proÞle with platinum-based regimens makes it a useful second-line option in patients whose disease progresses after treatment with these other regimens. Cisplatin is sometimes incorporated into the CDE regimen, making the four-drug combination regimen, PCDE, which is used only in France. Mechanism of Action. •

•

•

•

Cyclophosphamide (Figure 7) is an alkylating agent. These agents alkylate DNA bases, thereby producing cross-links that covalently link the two DNA strands and prevent cell replication. Doxorubicin (Figure 8) is an anthracycline. Anthracyclines interact with several different cellular targets, most importantly topoisomerase II. By inhibiting this DNA religation enzyme, anthracyclines exert their cytotoxic effect. Another mechanism leading to cell death, known as DNA intercalation, involves insertion of the anthracycline molecule between base pairs. This phenomenon causes single- and double-stranded breaks in DNA that inhibit cell proliferation. The free radicals generated by the reductive metabolism of anthracyclines may also damage cellular structures. Etoposide (Figure 3) is an epipodophyllotoxin. Epipodophyllotoxins act by causing single-strand breaks in DNA. Epipodophyllotoxins also inhibit DNA topoisomerase II to cause metaphase arrest, thereby inducing cytotoxic activity. Etoposide is cell-cycle-phase speciÞc, with predominant activity occurring in the late S phase and G2 phase. Cisplatin (Figure 4) is a platinum agent. Platinum agents inhibit tumor-cell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that cisplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle.

Clinical Performance. Because of the dose-limiting neutropenia associated with CDE, studies have attempted to evaluate the effect of GM-CSF. One study randomized 413 patients to receive CDE either every three weeks (control group)

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or every two weeks with GM-CSF (dose-intense group) (Thatcher N, 2000). The dose intensity of CDE was increased by 50% in the dose-intense group compared with the control group. Complete response rates were 40% for the dose-intense group and 28% for the control group; overall response rates were 78% for the dose-intense group and 79% for the control group. Survival was longer in the dose-intense group: survival rates for the dose-intense and control groups were, respectively, 47% and 39% at 12 months and 13% and 8% at 24 months. In the dose-intense group, less neutropenia was observed, but more thrombocytopenia and more-frequent blood and platelet transfusions. In a multicenter clinical trial, 457 patients (79% with ED-SCLC) were randomized to receive either CDE or PCDE (Urban T, 1999). Although the objective response rate was higher in the PCDE group (72%) than in the CDE group (53%), median overall survival was similar for the groups that received CDE (266 days) and PCDE (271 days). A higher fatal neutropenia rate was observed in the PCDE group (n = 23) than in the CDE group (n = 4), mainly in patients with ED-SCLC. Researchers concluded that the addition of cisplatin to CDE is toxic to patients with ED-SCLC and does not improve overall survival. Cisplatin/Cyclophosphamide/Epirubicin/Etoposide Regimen Overview. The cisplatin/cyclophosphamide/epirubicin/etoposide (PCEE) regimen is popular in France. Because it is more toxic than double- or triple-drug regimens, its use is limited to good-PS LD-SCLC patients. In addition to replacing doxorubicin with epirubicin, the PCEE regimen is given at a slightly higher dose intensity than the previously discussed PCDE regimen. Compared with the PCDE regimen, PCEE is more hematotoxic. It is uncertain which regimen is more active because no randomized trials exist that compare PCDE with PCEE. However, the use of PCEE is limited because of the high cost of epirubicin compared with the cost of doxorubicin. Mechanism of Action. •

•

•

Cisplatin (Figure 4) is a platinum agent. Platinum agents inhibit tumor-cell replication by creating intra- and interstrand DNA cross-links. Early studies suggested that cisplatin was cell-cycle-phase nonspeciÞc; more recent studies have shown complex and variable effects on the cell cycle. Cyclophosphamide (Figure 7) is an alkylating agent. These agents alkylate DNA bases, thereby producing cross-links that covalently link the two DNA strands and prevent cell replication. Epirubicin (Figure 9) is an anthracycline. Anthracyclines interact with several different cellular targets, most importantly topoisomerase II. By inhibiting this DNA re-ligation enzyme, anthracyclines exert their cytotoxic effect. Another mechanism leading to cell death, known as DNA intercalation, involves insertion of the anthracycline molecule between base pairs. This phenomenon causes single- and double-stranded breaks in DNA that inhibit

326

•

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cell proliferation. The free radicals generated by the reductive metabolism of anthracyclines may also damage cellular structures. Etoposide (Figure 3) is an epipodophyllotoxin. Epipodophyllotoxins act by causing single-strand breaks in DNA. Epipodophyllotoxins also inhibit DNA topoisomerase II to cause metaphase arrest, thereby inducing cytotoxic activity. Etoposide is cell-cycle-phase speciÞc, with predominant activity occurring in the late S phase and G2 phase.

Clinical Performance. One randomized French study evaluated the PCEE regimen in 117 ED-SCLC patients compared with the EP regimen in 109 ED-SCLC patients. Investigators found signiÞcant response rate and survival differences between the two arms: in the PCEE arm, the overall response rate was 76% and median survival was 10.5 months; in the EP arm, overall response was 61% and median survival was 9.3 months. Hematologic toxicity was signiÞcantly higher in the PCEE arm—22% with infections, compared with 8% in the EP arm—but the rate of toxicity-related deaths was not signiÞcantly different (Pujol JL, 2001). Topotecan, Single Agent Overview. Topotecan (GlaxoSmithKline/Merck/Nihon Kayaku’s Hycamtin) is approved in the United States and Japan as an intravenous (IV) monotherapy for patients with recurrent SCLC. Results of an international Phase III trial of the IV formulation for treatment of recurrent SCLC led to topotecan’s U.S. and Japanese launches. Results of another international Phase III trial comparing IV with oral topotecan in the second-line setting show that both forms of topotecan have similar efÞcacy. Another Phase III trial, ongoing in the United States, is comparing oral topotecan plus cisplatin with EP as Þrst-line therapy for patients with ED-SCLC. An application for European marketing as an IV monotherapy for patients with recurrent SCLC was Þled with the European Medicines Evaluation Agency (EMEA), but an EMEA panel voted against approval until more robust data are available. Mechanism of Action. Topotecan (Figure 11) is a topoisomerase I inhibitor. Topotecan binds to the topoisomerase I DNA complex and prevents religation of DNA single-strand breaks that occur naturally during DNA synthesis. Researchers believe the cytotoxicity of topotecan results from double-strand DNA damage produced during DNA synthesis, when replication enzymes interact with the ternary complex formed by topotecan, topoisomerase I, and DNA. Mammalian cells cannot efÞciently repair these double-strand breaks. Clinical Performance. Japanese and U.S. approval for second-line, singleagent use of IV topotecan was based on a Phase III international trial that compared single-agent topotecan with CAV in 211 SCLC patients with recurrent disease (von Pawel J, 1999). Overall response rates of 18.3% and 24.3% occurred in the CAV and topotecan arms, respectively. Overall survival was not statistically different between patients in the CAV arm (24.7 weeks) and the

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CH3 N CH3 HO

O N N O HO H3C

O

FIGURE 11. Structure of topotecan.

topotecan arm (25 weeks). Symptom improvement was reported to be greater in the topotecan group than in the CAV group for four out of eight symptoms evaluated, including dyspnea, anorexia, hoarseness, and fatigue. Grade 4 neutropenia occurred in 37.8% of patients in the topotecan arm and 51.4% in the CAV arm. Grade 4 thrombocytopenia and grade 3/4 anemia occurred more frequently with topotecan than CAV (9.8% and 17.7% versus 1.4% and 7.2%, respectively). Nonhematologic toxicities were mild and similar in both arms. Clinical evaluations in the Þrst-line setting have so far yielded disappointing results (Ormrod D, 1999). Phase III data failed to show any signiÞcant survival beneÞt for EP followed by IV topotecan, compared with EP alone, in previously untreated ED-SCLC patients. Toxicity proÞles were similar in both groups (Schiller JH, 2001[b]; Johnson D, 2000). Results of a U.S. and European Phase III study showed that oral topotecan has activity and tolerability similar to that of IV topotecan in chemosensitive SCLC patients (Eckardt JR, 2003). Patients with LD- or ED-SCLC who had a documented response to Þrst-line therapy, ECOG PS < 2, and measurable, recurrent disease with a treatment-free interval > 90 days were randomized to oral (n = 153) or IV (n = 151) topotecan. Response rates for oral and IV topotecan were 18.3% versus 21.9%, respectively. Median survival and one-year survival were 33 weeks and 33% for oral topotecan and 35 weeks and 29% for IV topotecan. Quality of life results showed no signiÞcant differences between oral and IV topotecan. Oral and IV topotecan were generally well tolerated; incidence of grade 4 toxicity by patient (oral and IV) was neutropenia (47% and 64%); thrombocytopenia (29% and 18%); grade 3/4 anemia (23% and 31%); and sepsis (3% and 3%). A larger, ongoing Phase III trial has randomized ED-SCLC patients to receive oral topotecan plus cisplatin or EP as Þrst-line therapy; encouraging preliminary results were disclosed at the American Society of Clinical Oncology (ASCO) meeting in 2004 (Eckardt JR, 2004). Tolerability data for 732 patients are available so far, but no efÞcacy data have been disclosed. Principal adverse experiences per patient for topotecan/cisplatin versus EP were the following: grade 4 neutropenia

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(32% versus 58%); platelets < 25,000 (10% versus 6%); grade 3/4 anemia (35% versus 20%); febrile neutropenia (1% versus 4%); infection (9% versus 12%); all grades of nausea/vomiting (58% and 39% versus 62% and 42%). Etoposide, Single Agent Overview. Etoposide monotherapy (oral or IV) is used mainly as palliative treatment for elderly and low-PS patients who have ED-SCLC or recurrent disease. Oral etoposide is used only rarely; because oral absorption is poor, oral etoposide must be given at double the dose required for IV etoposide to achieve the same effect, but this dosing produces more toxic effects. However, IV etoposide is less convenient than the oral form, which can be administered at home. Mechanism of Action. Etoposide (Figure 3) is an epipodophyllotoxin. Epipodophyllotoxins act by causing single-strand breaks in DNA. Epipodophyllotoxins also inhibit DNA topoisomerase II to cause metaphase arrest, thereby inducing cytotoxic activity. Etoposide is cell-cycle-phase speciÞc, with predominant activity occurring in the late S phase and G2 phase. Clinical Performance. In a randomized trial of palliative treatment in advanced SCLC, oral etoposide was compared with IV chemotherapy consisting of alternating cycles of cisplatin and etoposide (PE) and cyclophosphamide, doxorubicin, and vincristine (CAV) (Souhami RL, 1997). After 155 patients had been randomly assigned from a projected intake of 365 patients, an independent data monitoring committee examined the interim results. Survival was inferior at one year in the oral etoposide group compared with IV therapy (9.8% for oral versus 19.3% for IV), and there was a trend toward inferior overall survival. Median survival was 4.8 months for oral treatment and 5.9 months for IV therapy. Progression-free survival was worse in the oral etoposide arm (median = 3.6 months versus 5.6 months); overall response rate was also worse (32.9% versus 46.3%). With the exception of acute nausea and vomiting associated with IV chemotherapy, all aspects of symptom control and quality of life were either the same or worse in the oral etoposide group. Nonpharmacological Approaches Radiotherapy is the main nonpharmacological approach used in the treatment of SCLC and is administered in the thoracic region (thoracic radiotherapy [TRT]) or the cranial region (prophylactic cranial irradiation [PCI] or as whole-brain radiotherapy [WBRT]). TRT is used mainly in LD-SCLC and in combination with chemotherapy. No standard consensus exists on the exact timing of TRT delivery during the course of combined modality treatment, although it is becoming more evident that concurrent TRT (during chemotherapy) is more efÞcacious than sequential TRT (before or after TRT). Brain metastases are detected in approximately 10% of SCLC patients at the time of presentation and are subsequently diagnosed in 20–25% of patients during

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their lifetimes, with the likelihood of development increasing as length of survival time increases (Komaki R, 1981). In the absence of PCI, patients have a 50–80% chance of developing brain metastases during their Þrst two years of survival. Postmortem examination shows brain metastasis in 65% of SCLC patients (Rosen ST, 1983). A review of nine studies that examined the beneÞt of PCI showed that PCI reduced the frequency of brain metastases from 22% to 6% in LD-SCLC patients who had achieved a complete response. Therefore, PCI is given to all LD-SCLC patients who achieve a complete response to conventional treatment and, occasionally, to patients who achieve a very good partial response—that is, more than 75% decrease in tumor size (Armstrong J, 2002). The standard treatment for SCLC patients with brain metastases is WBRT plus chemotherapy. WBRT is sequentially administered, usually prior to chemotherapy or during chemotherapy. Because of the highly metastatic nature of most SCLC cases, surgery plays only a minor role. Some studies show that in early-stage SCLC (stages I to IIIA), surgery can achieve a high local control rate that may result in favorable longterm results (Passlick B, 2001). However, in more than 90% of SCLC patients, the disease is too systemic to be controlled by local resection; therefore, surgery is inappropriate (Beat-Ris H, 2002). EMERGING THERAPIES Commercial investment in R&D of small-cell lung cancer (SCLC) therapies is modest compared with such investment in many other tumor types, probably because of the relatively low incidence of SCLC. R&D activity in SCLC focuses chießy on investigating the activity of agents approved for other tumor types. During the past decade, median overall survival for SCLC patients has improved only by weeks. Only irinotecan (PÞzer’s Camptosar, Aventis/Prodesfarma’s Campto, Yakult/Daiichi’s Topotecin) has achieved any greater survival beneÞt over established agents. Myelosuppression—the primary toxicity of standard treatments—remains a problem with many of the cytotoxic therapies under investigation for SCLC. In common with most other cancer treatments, agents in development for SCLC are generally approved Þrst for recurrent or extensive-stage disease (EDSCLC) and later for limited-stage disease (LD-SCLC). A small number of agents with novel mechanisms of action are in Phase II trials, but currently, no clinical data are available for them. One of these novel agents is dehydrodidemnin B (PharmaMar’s Aplidin), a cyclodepsipeptide derived from the tunicate Aplidium albicans. This agent blocks cell division, induces apoptosis, and inhibits both the secretion of vascular endothelial growth factor and expression of its receptor. Phase II trials in SCLC began in May 2004 (PharmaMar, press release, May 2004). SanoÞ-Aventis’ reminertant (SR-48692) is an oral, nonpeptide, neurotensin NT1 receptor antagonist with neurotensin NT2 agonist properties. This agent entered Phase IIb trials for SCLC in February 2004. Introgen Therapeutics’ INGN-225 is an immunotherapeutic treatment in earlystage development; no clinical data are available at this time. INGN-225 entered a

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Phase I/II study in patients with SCLC in June 2003 (Introgen Pharmaceuticals, press release, June 2003); this agent stimulates patients’ own dendritic cells, which are then used as a therapeutic vaccine. Table 9 summarizes the drug therapies in development for SCLC. Conventional Cytotoxic Agents Overview. Conventional cytotoxic drugs are the mainstay of therapy for most cancers. Recent generations have been adapted to minimize toxicity proÞles and enhance therapeutic potential. Clinical trials for SCLC focus heavily on analyzing combinations of established cytotoxic drugs, aiming to improve response rate and survival time. Companies have also shown interest in developing new formulations and analogues in an attempt to reduce toxic side effects and increase the maximum tolerated dose. These agents may also have improved pharmacokinetic proÞles and different mechanisms of intracellular uptake. For SCLC, this section discusses new formulations of the vinca alkaloid vincristine (see “Onco TCS”) and the topoisomerase I inhibitor lurtotecan (see “OSI-211”). Mechanism of Action. Conventional cytotoxic agents interrupt the DNA replication and repair processes required for functional cell division. They act in several ways, including alkylation of DNA, resulting in strand breakage; inhibition of crucial enzymes required for DNA strand formation; and interference with spindle formation. Irinotecan. Irinotecan (PÞzer’s Camptosar, Aventis/Prodesfarma’s Campto, Yakult/Daiichi’s Topotecin) (Figure 12) is an injectable topoisomerase I inhibitor. Topoisomerase I inhibitors bind to the topoisomerase I-DNA complex, a nuclear enzyme that causes reversible single-strand breaks in DNA and prevents religation of the DNA strand, resulting in cell death. The drug is believed to exert its cytotoxic effects during the S phase of the cell cycle. Irinotecan is marketed worldwide for colorectal cancer and non-small-cell lung cancer (NSCLC). In Japan, irinotecan is registered for the treatment of ED-SCLC in combination with cisplatin (Bristol-Myers Squibb’s Platinol-AQ, Nippon Kayaku’s Randa, generics) and is under development for LD-SCLC. In the United States and Europe, the irinotecan/cisplatin combination (IP regimen) is undergoing Phase III trials for ED-SCLC. Japanese approval was based on the results of a 231-patient ED-SCLC Phase III Japanese trial that compared IP with the EP regimen (etoposide [Bristol-Myers Squibb’s VePesid/Etopophos, PÞzer’s Lastet, generics]) plus cisplatin (Noda K, 2002). Participation in the trial was limited to patients younger than age 70. Response rates for the IP regimen (84%) and median overall survival (12.8 months) were signiÞcantly higher than response rates and survival for the EP regimen (68%, 8.4 months). Grade 3/4 hematologic toxicities were signiÞcantly lower in the IP arm (neutropenia 65%, leukopenia 27%) than in the EP arm (neutropenia 92%, leukopenia 52%); nonhematologic toxicities were only slightly higher in the IP arm than in the EP arm (diarrhea 16% versus 0%).

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TABLE 9. Emerging Therapies in Development for Small-Cell Lung Cancer Compound

Development Phase

Marketing Company

Irinotecan (Camptosar, Campto, Topotecin) United States III Europe III Japan R

Pfizer Aventis Yakult/Daiichi

Paclitaxel (Taxol) United States Europe Japan

III III II

Bristol-Myers Squibb Bristol-Myers Squibb Bristol-Myers Squibb

Docetaxel (Taxotere) United States Europe Japan

II II I

Aventis Aventis Aventis

Vinorelbine (Navelbine) United States Europe Japan

II II —

Pierre Fabre/GlaxoSmithKline Pierre Fabre/GlaxoSmithKline —

Gemcitabine (Gemzar) United States Europe Japan

II III I

Eli Lilly Eli Lilly Eli Lilly

Pemetrexed (Alimta) United States Europe Japan

II — —

Eli Lilly — —

Oral topotecan (Hycamtin) United States Europe Japan

III III —

GlaxoSmithKline/Merck/Nihon Kayaku GlaxoSmithKline/Merck/Nihon Kayaku —

OSI-211 United States Europe Japan

II II —

OSI Pharmaceuticals OSI Pharmaceuticals —

Onco TCS United States Europe Japan

II — —

Inex Pharmaceuticals — —

Immunotherapies IMC-BEC2 United States Europe Japan

III III —

ImClone Merck KGaA —

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TABLE 9. (continued) Compound

Development Phase

Marketing Company

SRL-172 United States Europe Japan

— II —

— SR Pharma —

Tumor-activated pro-drugs huN901-DM1 United States Europe Japan

II I —

ImmunoGen Vernalis —

Angiogenesis inhibitors ZD-6474 United States Europe Japan

— II —

— AstraZeneca —

Bevacizumab (Avastin) United States Europe Japan

II — —

Genentech/Roche — —

Thalidomide (Thalomid) United States Europe Japan

II III —

Celgene Celgene —

Bioreductive agents Tirapazamine (Tirazone) United States Europe Japan

II — —

Sanofi-Aventis — —

Imatinib (Gleevec/Glivec) United States Europe Japan

II — —

Novartis — —

Synthetic retinoids Fenretinide United States

II — —

McNeil Pharmaceuticals/ National Cancer Institute — —

II — —

Millennium — —

Europe Japan Proteasome inhibitors Bortezomib (Velcade) United States Europe Japan R = Registered.

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H3C O

O

O N

N N O N

HO H3C

O

FIGURE 12. Structure of irinotecan.

A randomized Phase III trial was initiated in North America to conÞrm the results of the Japanese trial (Sandler A, 2003). Based on U.S. pilot data, doses of IP were modiÞed so that patients received 30 mg/m2 cisplatin and 65 mg/m2 irinotecan on days 1 and 8, every 21 days, rather than 60 mg/m2 of both agents on days 1, 8, and 15 every 28 days. No response data are yet available, but safety data from 122 patients demonstrate that the dose-adjusted IP regimen is better tolerated than the Japanese schedule. Grade 3/4 neutropenia and anemia were reduced from 65.3% and 26.7% in the Japanese study to 32% and 5%, respectively, in the North American study. In addition, 20% of patients experienced grade 3 diarrhea in the North American study compared with 11% grade 3 and 5% grade 4 diarrhea in the Japanese trial. Another Phase III study investigating IP, using the same regimen employed by the Japanese Clinical Oncology Group (JCOG) group, is recruiting patients. This study is sponsored by the Southwest Oncology Group (SWOG), National Cancer Institute (NCI), North Central Cancer Treatment Group (NCCTG), and Cancer and Leukemia Group B (CALGB). In Phase II trials of Þrst-line therapy in patients with ED-SCLC, irinotecan is being combined with gemcitabine (Eli Lilly’s Gemzar; see “Gemcitabine” section) in 81 patients (Akerley WL, 2004), with paclitaxel (Bristol-Myers Squibb’s Taxol, generics; see “Paclitaxel” section) plus carboplatin (BristolMyers Squibb’s Paraplatin, generics) in 25 patients (Glisson SD, 2003), and with etoposide in 23 patients (Farhat FS, 2004). Poor response rates were observed in the trial of irinotecan combined with gemcitabine: the overall response rate was 29%, but survival time was eight months, which was not signiÞcantly different from results reported with the EP regimen. The triple regimen of paclitaxel, irinotecan, and carboplatin (PIC regimen) demonstrated promising results. An overall response rate of 88% included 36% complete responses (CR) and 52% partial responses (PR). Mean survival time was 453 days (approximately 15 months), and the most signiÞcant toxicities included 40% neutropenia, 24% thrombocytopenia, and 28% diarrhea. The combination of irinotecan and etoposide resulted in 24% CR and 40% PR.

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Other Þrst-line regimens being evaluated in ED-SCLC patients include alternating schedules of double-and triple-drug combinations. Phase II data presented at the American Society of Clinical Oncology (ASCO) meeting in 2004 analyzed 27 patients who received IP (60 mg/m2 irinotecan on days 1, 8, and 15, and 60 mg/m2 cisplatin on day 1) in the Þrst, third, and Þfth cycles and received doxorubicin (PÞzer’s Adriamycin)/cyclophosphamide/etoposide (ACE regimen) (50 mg/m2 doxorubicin on day 1; 750 mg/m2 cyclophosphamide on day 1; 80 mg/m2 etoposide on days 1–3) in the second, fourth, and sixth cycles of chemotherapy (Yonei T, 2004). Both regimens were repeated every three to four weeks, toxicity permitting, and granulocyte colony-stimulating factor (GCSF) was administered when patients experienced grade 4 neutropenia or grade 3 febrile neutropenia. The overall response rate was 92.6% (14.8% CR; 77.8% PR), with a median survival of 13.8 months and one- and two-year survival rates of 51.6% and 9.0%, respectively. This alternating regimen induced signiÞcant toxicities, including 96.3% neutropenia, 37% febrile neutropenia, and 25.9% thrombocytopenia. Phase II Japanese trials for irinotecan in the LD-SCLC setting have yielded promising results. A randomized trial evaluated IP alone versus IP plus etoposide (IPE regimen) after radiotherapy in 23 patients (Saito H, 2002). Trial results showed that the triple combination is too toxic; this arm was discontinued. However, based on an acceptable toxicity proÞle and response rates of 91% in the IP arm, this regimen is still being evaluated in a single-arm study. In a follow-up study to the IP versus IPE trial, the West Japan Thoracic Oncology Group evaluated the safety proÞle and response rates of EP with concurrent radiation followed by IP in 49 patients with LD-SCLC (Mitsuoka S, 2004). Median survival had not yet been reached, and the overall and CR rates were 87.8% and 40.8%, respectively. These rates are comparable to those achieved for EP with concurrent thoracic radiotherapy (TRT) (96% overall response rate; 40% CR response rate) in previous studies (Takada M, 2002). Although the oneyear survival rate of 86.4% showed improvement on PE/concurrent TRT (75%), two-year survival is equivalent (54% for EP/TRT; 51% for EP/TRT followed by IP). Grade 4 toxicities were high in the EP/TRT/IP regimen and included 84% neutropenia and 10% anemia. A Korean study evaluated the same drug regimens in a different schedule of administration (Lee DH, 2003). Twenty-three LD-SCLC patients received two cycles of IP (80 mg/m2 irinotecan, 40 mg/m2 cisplatin on days 1 and 8, every three weeks) followed by concurrent radiation and EP (60 mg/m2 cisplatin on day 1, 100 mg/m2 etoposide on days 1–3, every three weeks). After completion of this regimen, 100% of patients responded with 26% CR and 74% PR. SigniÞcant toxicities occurred throughout this regimen but particularly during the concurrent radiation/chemotherapy cycles. Grade 3/4 toxicities included 27% neutropenia and 20% febrile neutropenia during the IP chemotherapy, followed by 100% neutropenia, 60% febrile neutropenia, 60% anemia, and 52% esophagitis. One patient died of radiation pneumonitis four months after completing therapy, and 18 patients required hospitalization for esophagitis and/or febrile neutropenia.

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A carboplatin/irinotecan combination achieved a response rate and median overall survival that is approximately comparable to outcomes of currently marketed agents, such as the EP regimen, in a trial of 27 LD-SCLC patients (89%, 18.2 months) and 34 ED-SCLC patients (84%, 9.7 months) (Kinoshita A, 2002). Grade 3/4 toxicities included leukopenia (35%), neutropenia (76%), thrombocytopenia (42%), and diarrhea (13%). Irinotecan is also under Phase II investigation in combination with other chemotherapeutic agents in the second-line setting. Response rates of 68.2% were achieved in 24 patients receiving carboplatin/irinotecan, and the median survival time was 194 days (approximately 6.5 months) (Hirose T, 2003). Grade 3/4 toxicities included 58% leukopenia, 63% neutropenia, 58% thrombocytopenia, and 67% anemia. When irinotecan was combined with ifosfamide (Bristol-Myers Squibb’s Ifex, Baxter’s Mitoxana/Holoxan, Shionogi’s Ifomide, generics), an overall response rate of 52.9%, including 5.8% CR, was achieved in 34 patients (Ichiki M, 2003). This study showed a favorable toxicity proÞle: grade 3/4 neutropenia in 53% of patients and no other reported hematologic toxicities. A trial of IPE in 40 patients with recurrent disease yielded response rates and overall survival of 78% and 11.4 months, respectively (Koichi G, 2002). Patients had received chemotherapy treatment when SCLC was Þrst diagnosed. These response and survival rates are far greater than expected for recurrent disease (median survival with currently available therapies is only six months). Two abstracts presented at the ASCO meeting in 2003 described the combination of gemcitabine and irinotecan as a second-line treatment in SCLC patients. In one study, 29 patients received 1,500 mg/m2 gemcitabine and 150 mg/m2 irinotecan on day 1 every two weeks (Domine M, 2003). The overall response rate was 27.6%, including one CR in a patient with chemotherapy-sensitive disease. When the data were analyzed according to relapsed disease (chemotherapy-sensitive) versus refractory disease (chemotherapy-insensitive), 25% of patients with refractory disease responded versus 31% of patients with relapsed disease. Median survival was 9.3 months, median time to progression (TTP) was 5.3 months, and no grade 3/4 hematologic toxicity was observed. In a slightly different schedule, 26 SCLC patients received 1,000 mg/m2 gemcitabine on days 1 and 8 and 300 mg/m2 irinotecan on day 8, every 21 days (Agelaki S, 2003). Only three patients (10%) achieved a PR; two of these were refractory to previous chemotherapy and one was sensitive. Median survival was six months. Grade 3/4 neutropenia, thrombocytopenia, and diarrhea were observed in 29%, 13%, and 10% of patients, respectively; three patients experienced febrile neutropenia. Paclitaxel. Paclitaxel (Figure 13), an extract from the bark of the yew (Taxus brevifolia), was the Þrst taxane to enter the clinic as an anticancer agent. Taxanes inhibit the dynamic reorganization of microtubule networks that is essential for vital interphase and mitotic cellular functions. Paclitaxel achieves selective toxicity against rapidly proliferating cells and is active mainly in the G2/M phase of the cell cycle. This intravenously administered compound is used to treat a wide variety of cancers in major markets worldwide, but despite some Þrst-line,

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FIGURE 13. Structure of paclitaxel.

off-label use in research institutes, it is still in development for SCLC treatment. Researchers have carried out several clinical trials, but no existing data demonstrate any increase in response rates resulting from paclitaxel-containing regimens compared with standard treatments. Paclitaxel is being investigated in Phase III trials in the United States and Europe and in Phase II trials in Japan. A Phase III trial investigated whether a paclitaxel/etoposide/carboplatin (TEC) regimen could improve the outcome for previously untreated patients with SCLC compared with the standard CEV regimen of carboplatin/etoposide/vincristine (Eli Lilly/EG Labo/Shionogi’s Oncovin, generics) (Reck M, 2003). Six hundred and eight patients with stages I–IV SCLC received a maximum of six cycles of either TEC or CEV. No statistically signiÞcant differences in response rate were observed when data were analyzed for all stages or stratiÞed for stage. However, median survival was improved in the TEC arm (12.7 months) compared with the CEV arm (11.7 months), and mean one-year, two-year, and three-year survival rates were 51%, 20%, and 17% in the TEC arm compared with 48%, 16%, and 9% in the CEV arm. When survival was stratiÞed by stage, the data showed a survival advantage in the TEC arm for patients with LD-SCLC (stages I–IIIa, 18.7 months versus 16.9 months; stage IIIb, 15.7 months versus 13.2 months) but no difference for patients with ED-SCLC (stage IV). Although the rates of leukopenia, neutropenia, and febrile neutropenia were similar in both arms of the study, a signiÞcantly higher frequency and severity of anemia and thrombocytopenia in patients receiving CEV resulted in a higher incidence of platelet and red blood cell transfusions. This trial is the Þrst randomized study to demonstrate an increase in survival and a reduction in drug-related toxicities for a paclitaxel-containing regimen compared with standard therapy. The TEC regimen was compared with paclitaxel plus topotecan (GlaxoSmithKline/Merck/Nihon Kayaku’s Hycamtin) in patients with previously untreated ED-SCLC (Thompson DS, 2004). The overall response rate for TEC was 77% (13% CR) compared with 49% (6.5% CR) for the paclitaxel/topotecan arm. In addition, median survival, median progression-free survival, and the toxicity proÞle were superior in the TEC arm compared with paclitaxel/topotecan. Phase III trials that compared paclitaxel combined with the etoposide/cisplatin regimen (TEP) with the EP regimen alone yielded disappointing clinical results. In a 587-patient trial, investigators observed no signiÞcant differences in median

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survival between TEP and EP, but TEP resulted in more toxic deaths (Niell HB, 2002). Toxicities and treatment-related deaths led to early closure of a 133-patient Phase III trial (Mavroudis D, 2001). These results suggest that paclitaxel- and cisplatin-containing, three-drug combinations are too toxic in SCLC. Other noteworthy paclitaxel regimens that have been evaluated for Þrst-line therapy include paclitaxel and carboplatin in combination with either ifosfamide or topotecan. Trials have failed to demonstrate any improvement in survival or toxicity proÞle when these combinations are compared with standard treatments. The ifosfamide-containing regimen yielded a response rate of 69% and median overall survival of 9.5 months in 35 ED-SCLC patients; neutropenia was the principal grade 3/4 toxicity (Neubauer MA, 2002). The topotecan-containing regimen effected a CR rate of 51% and median overall survival of 20 months in 68 LD-SCLC patients who completed the therapy; grade 3/4 toxicity included leukopenia (60%), thrombocytopenia (42%), and fatigue (14%) (Gray JR, 2002). The paclitaxel/topotecan regimen produced efÞcacy rates similar to those of standard chemotherapy, but it was associated with a high incidence of myelosuppression and febrile neutropenia in previously untreated patients with ED-SCLC (Ramalingam S, 2004). Paclitaxel is also under investigation as a single-agent, Þrst-line therapy for SCLC. A study in NSCLC investigated the use of single-agent paclitaxel in a “dose-dense” regimen of 150 mg/m2 /week for six weeks of an eight-week cycle. Unprecedented results for a single agent in this disease were obtained, prompting its evaluation for Þrst-line therapy in ED-SCLC (Graziano SL, 2003). In 29 evaluable patients, an overall response rate of 28% was achieved (3% CR), with a median TTP and overall survival of 3.8 and 9.7 months, respectively, and a one-year survival rate of 43%. This agent induced grade 3/4 neutropenia in 24% of patients, febrile neutropenia in 6%, anemia in 9%, and thrombocytopenia in 3%. Grade 3/4 nonhematologic toxicities included sensory and motor neuropathy (27% and 12%, respectively) and fatigue and dyspnea (both occurred in 18% of patients). This dose-dense regimen achieved response rates similar to those gained in the more convenient three-weekly schedule of paclitaxel, but survival rates were comparable to those observed in patients receiving platinum-based combinations. SCLC is primarily a disease of older people, a population that can be difÞcult to treat with the same dose and schedule of chemotherapeutic agents used to treat younger, Þtter patients. Previous studies have demonstrated that administering a lower dose of paclitaxel on a weekly basis can provide greater dose intensity and less toxicity (Akerley W, 1998). A Phase II trial of weekly, low-dose paclitaxel/carboplatin attempted to improve survival and minimize toxicities in patients with ED-SCLC who were aged 70 or older or had a PS of two (Neubauer M, 2004). Sixty-six evaluable patients received 80 mg/m2 paclitaxel and area under the curve (AUC) = 2 carboplatin on days 1, 8, and 15 of a four-week cycle for up to six cycles. Response rates were 1.5% CR, 36.4% PR, 12.1% stable disease (SD), and 50% progressive disease (PD), with a median survival of 7.2 months, TTP of 3.5 months, and estimated one- and two-year survival rates

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of 30% and 4%, respectively. Grade 3/4 toxicities included neutropenia and anemia in 22% and 5.2% of patients, respectively. The response and survival rates observed in this trial are comparable to results achieved in trials that included patients with a better PS. In addition, the level of neutropenia was signiÞcantly less than reported in previous trials, indicating this regimen may be a preferred choice for elderly or poor-PS patients. Paclitaxel is also under investigation as a second-line therapy, in combination or as a monotherapy, for patients with relapsed or refractory SCLC. Patients who did not respond to Þrst-line therapy with etoposide/carboplatin (EC regimen) or who relapsed within three months of a conÞrmed prior response were eligible for therapy with single-agent paclitaxel (Joos G, 2004). Forty-four patients received 200 mg/m2 paclitaxel every three weeks for up to six cycles. Patients received a median number of three cycles; an overall response rate of 20% was achieved, with a median overall survival of four months and a TTP of three months. The most signiÞcant toxicity was febrile neutropenia, which occurred in 23% of patients. In a paclitaxel combination study, patients who had progressed after Þrst-line therapy with a platinum agent plus etoposide were treated with 80 mg/m2 paclitaxel on days 1, 8, and 15 and 1,000 mg/m2 gemcitabine on days 1 and 8, every three weeks (Dongiovanni V, 2004). Of the 31 patients enrolled in the study, Þve had LD-SCLC and 26 had ED-SCLC; their median age was 64; and they had a performance status of 0–2. Analysis of response revealed PRs in eight patients (26%), SD in Þve patients (16%), and progression in 18 patients (58%), with a median survival time of 7.4 months and a median TTP of 9.6 weeks. Toxicities included 26% grade 3 neutropenia, 22% grade 3 thrombocytopenia, 3% grade 4 thrombocytopenia, 13% grade 3–4 asthenia, and 32% grade 1–2 neuropathy. Docetaxel. An analogue of paclitaxel and a member of the taxane drug class, docetaxel (Aventis’ Taxotere) (Figure 14) is partially synthesized from an extract of the European yew (Taxus baccata). Taxanes inhibit the dynamic reorganization of microtubule networks that is essential for vital interphase and mitotic cellular functions. Docetaxel achieves selective toxicity against rapidly proliferating cells and is active mainly in the S phase of the cell cycle. It is available in all major markets for the treatment of breast cancer and NSCLC and is under investigation for a variety of other cancers. For SCLC, no randomized data are available that compare docetaxel with paclitaxel or with conventional regimens. Docetaxel is in Phase II trials in Europe and the United States for Þrst-line treatment of ED-SCLC and in Phase I trials in Japan for recurrent disease. Phase II docetaxel trials had disappointing results in terms of survival. One trial that combined docetaxel with cisplatin in 33 ED-SCLC patients showed an overall survival rate of only seven months; grade 3/4 neutropenia was low (20%) (Lianes P, 2001). Another study combined docetaxel with gemcitabine in untreated patients with ED-SCLC (Skarlos DV, 2003). Only six of 20 patients achieved a PR; the trial was closed prematurely because of the poor response rate. Median TTP was eight months, and median survival was 9.6 months; both

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FIGURE 14. Structure of docetaxel.

hematologic and nonhematologic toxicity was acceptable. The same combination was investigated in elderly ED-SCLC patients (>65 years), and the study found a median overall survival of only four months (Thompson DS, 2004). The combination of docetaxel/gemcitabine was also investigated in the secondline setting in 22 pretreated patients with LD-SCLC or ED-SCLC (Agelaki S, 2004). No complete or partial responses were observed, disease stabilization occurred in only one patient, and median survival was 14 weeks with a sixmonth survival rate of only 28%. These results show this combination is inactive as a salvage or second-line therapy in these poor-prognosis patients. Vinorelbine. Vinorelbine (Pierre Fabre/GlaxoSmithKline/Kyowa Hakko’s Navelbine) (Figure 15), an intravenous (IV) vinca alkaloid, was Þrst launched in 1989 as a treatment for NSCLC. Vinorelbine binds to tubulin and prevents its polymerization, thus hindering formation of the mitotic spindle. The agent’s role as a component of multidrug regimens in SCLC treatment is being evaluated in Phase II trials in the United States and Europe. Vinorelbine achieved disappointing response rates in clinical trials. A Phase II trial of a vinorelbine/carboplatin regimen in 58 SCLC patients with poor prognosis reported a 55% overall response rate, including 10% CR, TTP of 18 weeks, and a 26-week median survival; 76% of the patients experienced grade 3/4 hematologic toxicity, far greater than the toxicity observed in trials of carboplatin combined with etoposide (Mackay HJ, 2003). Investigators suggest their study

N

CH3 N

N H CH3O2C CH3O

CH3 N HO CH3

OCOCH3 CO2CH3

FIGURE 15. Structure of vinorelbine.

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demonstrates the vinorelbine/carboplatin regimen was active, but because of the associated high toxicity, it did not warrant further study in this setting. A similar conclusion was drawn in a study of vinorelbine/carboplatin in a population of elderly patients with ED-SCLC. Therapy was highly toxic: three patients (11%) died while on treatment (Gridelli C, 2002). In the second-line setting, vinorelbine has been combined with gemcitabine in patients with relapsed or refractory SCLC (Hainsworth JD, 2003). This study determined that the activity of vinorelbine/gemcitabine is modest and limited to patients with relapsed, rather than refractory, disease. Gemcitabine. First marketed as an anticancer therapy in the 1990s, the antimetabolite gemcitabine is available in the United States and Europe for the treatment of pancreatic cancer and NSCLC. Antimetabolites block normal DNA synthesis, thereby stopping cell replication. Gemcitabine (Figure 16) exhibits cellphase speciÞcity, primarily killing cells that are undergoing DNA synthesis (S phase) and blocking the progression of cells through the G1/S-phase boundary. It is being investigated for SCLC and has reached Phase III trials in Europe, Phase II trials in the United States, and Phase I trials in Japan. In the Þrst-line setting, gemcitabine is being investigated in combination regimens. In the second-line setting, it is being investigated both as a single agent and in combination with other agents. Data from Phase II studies investigating Þrst-line gemcitabine double combinations have reported poor response rates, compared with standard chemotherapy. Eighty-one patients with ED-SCLC received 1,000 mg/m2 gemcitabine and 100 mg/m2 irinotecan on days 1 and 8 of a 21-day cycle for a maximum of six cycles or until disease progression (Akerley WL, 2004). The overall response rate was 29%, with a median survival of eight months. Grade 3/4 toxicities were less than toxicities reported in other chemotherapeutic regimens: 23% neutropenia, 9% anemia, 7% thrombocytopenia, and 5% febrile neutropenia. In a randomized Phase III study involving 241 patients of unreported stage, the gemcitabine/carboplatin (GC) regimen resulted in a median survival (8.1 months) similar to that of the EP regimen (8.2 months), with more hematologic toxicities but fewer nonhematologic toxicities; GC patients required less frequent

NH2 N HO

O O

OH

N F F

FIGURE 16. Structure of gemcitabine.

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administration-related hospitalization than EP patients (James LE, 2002). Combination of gemcitabine with carboplatin in 69 ED-SCLC patients produced a response rate of 43%, median survival of 9.2 months, and an estimated median TTP of four months (Sirridge C, 2003). Grade 3/4 toxicities included 39% neutropenia, 33% thrombocytopenia, and 13% anemia. In a Phase II study, gemcitabine was combined with docetaxel, but the trial was closed early due to poor response rates (Skarlos DV, 2003). Gemcitabine is also under investigation as part of the triple regimen cisplatin/ etoposide/gemcitabine (PEG) in chemotherapy-naive LD-SCLC and ED-SCLC patients. A total of 54 patients received 100 mg/m2 gemcitabine on days 1 and 8, 70 mg/m2 cisplatin on day 2, and 50 mg/m2 etoposide on days 3, 4, and 5 every three weeks (De Marinis F, 2003). An overall response rate of 72.2% was achieved (including 18.5% CR and 53.7% PR), with a median survival of ten months and a one-year survival probability of 37.5%. Grade 3/4 hematologic toxicity was higher than toxicity reported for gemcitabine double regimens, with 66.7% of patients suffering from neutropenia and 53.7% suffering from thrombocytopenia. When the PEG regimen was compared with cisplatin/gemcitabine (PG regimen) in a randomized Phase II trial, both regimens were active in LD-SCLC and ED-SCLC patients, but the highest overall response rates were achieved by PG in LD-SCLC (80%) and by PEG in ED-SCLC (68%) (De Marinis F, 2002). The PEG combination showed a trend toward more grade 3/4 toxicity. Nevertheless, both combinations seem to be less toxic than standard regimens. Trials evaluating gemcitabine in previously treated SCLC patients show some activity in those who are sensitive to chemotherapy. Three of 12 relapsed SCLC patients (25%) receiving 100 mg/m2 gemcitabine and 20 mg/m2 vinorelbine on days 1, 8, and 15 of a 28-day cycle achieved a partial remission, but none of 17 patients with refractory disease responded (Hainsworth JD, 2003). Hematologic toxicities were fewer with this regimen, providing this subset of patients with an additional treatment option. In contrast, a study that analyzed gemcitabine in combination with docetaxel reported no responses, despite including patients who were sensitive to chemotherapy (Agelaki S, 2004). Two abstracts presented at ASCO 2003 described the combination of gemcitabine and irinotecan as a second-line treatment in SCLC patients. In one study, 29 patients received 1,500 mg/m2 gemcitabine and 150 mg/m2 irinotecan on day 1 every two weeks (Domine M, 2003). The overall response rate was 27.6%, including one CR in a patient with chemotherapy-sensitive disease. When the data were analyzed according to relapsed (chemotherapy-sensitive) versus refractory (chemotherapy-insensitive) disease, 25% of patients with refractory disease responded versus 31% of patients with relapsed disease. Median survival was 9.3 months, median TTP was 5.3 months, and no grade 3/4 hematologic toxicity was observed. In a slightly different schedule, 26 SCLC patients received 1,000 mg/m2 gemcitabine on days 1 and 8 and 300 mg/m2 irinotecan on day 8, every 21 days (Agelaki S, 2003). Only three patients (10%) achieved a PR; two of them were refractory to previous chemotherapy and one was sensitive. Median

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survival was six months. Grade 3/4 neutropenia, thrombocytopenia, and diarrhea were observed in 29%, 13%, and 10% of patients, respectively, and three patients experienced febrile neutropenia. Phase II data presented at ASCO 2004 described the administration of paclitaxel and gemcitabine in SCLC patients (Dongiovanni V, 2004). This study reported response rates of 20% in chemotherapy-refractory patients and 29% in chemotherapy-sensitive patients, with an overall median TTP of 9.6 weeks and overall survival of 7.4 months. Toxicities included grade 3/4 neutropenia, thrombocytopenia, and asthenia in 26%, 25%, and 13% of patients, respectively. Single-agent gemcitabine has shown modest responses as a second-line therapy in SCLC patients. Phase II data from the Eastern Cooperative Oncology Group reported an overall response rate of 11.9% in 44 patients receiving gemcitabine 1,000 mg/m2 on days 1, 8, and 15 of a 29-day cycle (Masters G, 2003); the responding patients included four who were sensitive to chemotherapy and one with refractory disease. The overall median survival was 7.1 months and was not signiÞcantly different in refractory and sensitive patients. Both grade 3/4 neutropenia and thrombocytopenia were observed in 27% of patients, and grade 3/4 pulmonary and neurological toxicities occurred in 9% and 14% of patients, respectively. Pemetrexed. Pemetrexed (Eli Lilly’s Alimta) (Figure 17) is a new-generation, multitargeted antifolate that inhibits thymidylate synthase as well as dihydrofolate reductase and glycinamideribonucleotide (GAR) formyl-transferase. Lilly hopes that the multitargeted approach will overcome resistance acquired through overexpression of any single enzyme. This agent has shown efÞcacy in NSCLC and is in Phase II trials for SCLC in the United States. Phase II data presented at ASCO 2004 reported results from 17 ED-SCLC patients randomized to receive either 500 mg/m2 pemetrexed plus AUC = Þve carboplatin or 500 mg/m2 pemetrexed plus 75 mg/m2 cisplatin every 21 days for a maximum of six cycles (Socinski MA, 2004). Nine patients achieved a PR (overall response rate of 55%), six had SD, and two had PD. These regimens were well tolerated: only two patients (12%) experienced grade 3/4 adverse events, including neutropenia, nausea, vomiting, pneumonia, dehydration, and hyponatremia. At the time of composing this reference, the trial has been complete, but no survival data seem yet to be available from this study. Current response rates do not show an improvement over the EP standard chemotherapy regimen.

FIGURE 17. Structure of pemetrexed.

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Oral Topotecan. Topotecan (GlaxoSmithKline/Merck/Nihon Kayaku’s Hycamtin) (Figure 11) is a water-soluble, semisynthetic analogue of camptothecin that is approved in the United States and Japan as an IV monotherapy for patients with recurrent SCLC. This topoisomerase I inhibitor binds to the topoisomerase I-DNA complex and prevents religation of DNA single-strand breaks that occur naturally during DNA synthesis. The oral formulation of topotecan is under Phase III investigation for SCLC in both the Þrst- and second-line settings in the United States and Europe. Results of a U.S. and European Phase III study showed that oral topotecan has activity and tolerability similar to that of IV topotecan in chemotherapy-sensitive SCLC patients (Eckhardt JR, 2003). Patients with LD-SCLC or ED-SCLC who had a documented response to Þrst-line therapy, Eastern Cooperative Oncology Group PS < 2, and measurable, recurrent disease with a treatment-free interval > 90 days were randomized to oral (n = 153) or IV (n = 151) topotecan. Response rates for oral and IV topotecan were 18.3% and 21.9%, respectively. Median survival and one-year survival were 33 weeks and 33% for oral topotecan and 35 weeks and 29% for IV topotecan. Quality-of-life results showed no signiÞcant differences between oral and IV topotecan, and both were generally well tolerated; incidence of grade 4 toxicity by patient (oral/IV) was neutropenia (47%/64%), thrombocytopenia (29%/18%), grade 3/4 anemia (23%/31%), and sepsis (3%/3%). Oral topotecan was also investigated in patients with ED-SCLC in the secondline setting in combination with carboplatin (Jett J, 2004). Twenty-six patients received 1.75 mg/m2 /daily for Þve days plus AUC = 5 carboplatin on day 5, with granulocyte colony-stimulating factor for support, every 21 days for a maximum of six cycles. The trial was closed early due to unacceptable toxicities, which included three deaths due to infection caused by low absolute neutrophil count and one death due to a cardiovascular event. Other grade 3+ toxicities included 85% hematologic events and 54% nonhematologic events. PRs occurred in 62% of patients; median survival was 11.8 months with a 46% one-year survival rate. Although this regimen achieved good response rates, the levels of toxicity were too high. In the Þrst-line setting, a large, ongoing Phase III trial has randomized EDSCLC patients to receive oral topotecan plus cisplatin or EP as Þrst-line therapy. Encouraging preliminary results were disclosed at ASCO 2004 (Eckardt JR, 2004). Tolerability data for 732 patients are available so far, but no efÞcacy data have been disclosed, although the blinded study has now been complete. Principal adverse experiences for topotecan/cisplatin versus EP were grade 4 neutropenia (32% versus 58%); platelets <25,000 (10% versus 6%); grade 3/4 anemia (35% versus 20%); febrile neutropenia (1% versus 4%); infection (9% versus 12%); and all grades of nausea/vomiting (58%/39% versus 62%/42%). Clinical evaluations of efÞcacy in the Þrst-line setting have so far yielded disappointing results in several trials (Ormrod D, 1999; Jett J, 2002). In previously untreated ED-SCLC patients, Phase III data failed to show any signiÞcant survival beneÞt for EP followed by IV topotecan, compared with EP alone. Toxicity

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proÞles were similar in both treatment groups (Schiller JH, 2001[b]; Johnson D, 2000). OSI-211. OSI Pharmaceuticals is developing OSI-211, a liposomal formulation of the topoisomerase-1 inhibitor lurtotecan. This agent is in Phase II trials for both relapsed epithelial ovarian cancer and recurrent SCLC in the United Kingdom and the United States. In late 2002, OSI Pharmaceuticals initiated an open-label, single-arm Phase II trial in patients with recurrent SCLC. The study planned to enroll 72 patients and analyze the safety and efÞcacy of OSI-211 when administered on three consecutive days every three weeks. A previous study involving 80 patients with relapsed ovarian cancer compared two dosing schedules: 1.8 mg/m2 /day on days 1, 2, and 3 versus 2.4 mg/m2 on days 1 to 8 (OSI Pharmaceuticals, Web site). Response rates were clearly superior in the three-day schedule, which was adopted for subsequent trials. OSI Pharmaceuticals has reported that to develop this agent further, the company requires that studies prove a clear difference in activity, safety, and convenience compared with topotecan (GlaxoSmithKline’s Hycamtin). An ongoing Phase II trial involving relapsed epithelial ovarian cancer patients is comparing OSI-211 with topotecan. OSI Pharmaceuticals is unlikely to proceed with development of OSI-211 if results are unfavorable in this setting. Onco TCS. Onco TCS (Inex Pharmaceuticals’ vincristine sulfate liposomal injection) contains the cytotoxic vincristine encapsulated in sphingomyelin liposomes. This drug is in Phase II clinical trials for SCLC in the United States and has been preregistered for non-Hodgkin’s lymphomas. The drug delivery system is described by Inex as a “transmembrane carrier system” (TCS). Encapsulation of cytotoxics in liposomes is intended to extend the active agent’s circulation in the bloodstream and promote accumulation of the drug in tumors. Free-form (unencapsulated) drugs circulate throughout the body, diluting their effectiveness and damaging healthy tissues. Therefore, because of its neurotoxicity, optimal doses of vincristine are often not administered to elderly patients or to patients who require repeated treatment. An ongoing Phase II trial investigating Onco TCS in SCLC was initiated in August 2000. Eligible patients include patients who relapsed following standard Þrst-line chemotherapy, chemotherapy-naive patients whose clinical status precludes the use of combination chemotherapy, and patients whose disease is refractory after standard second-line therapy. In May 2002, Inex disclosed interim data showing that of 24 evaluable patients, two achieved a PR with a reduction in tumor volume greater than 50% and four patients had SD (Inex, press release, May 2002). Immunotherapies Overview. Immunotherapeutic approaches for cancer involve both speciÞc and nonspeciÞc stimulation of the immune system. SpeciÞc stimulation occurs when

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tumor antigens are used in a vaccine formulation to direct the immune system against the cancer cells, avoiding healthy cells. NonspeciÞc stimulation occurs when an adjuvant (a substance such as bacille Calmette-Gu´erin [BCG] that evokes a generalized immune response) is administered. The idea of using vaccines to stimulate therapeutic tumor-antigen-speciÞc immune responses holds promise as a complementary approach to current treatment options in most cancers. Cancer vaccines often incorporate both tumor antigens and an immunostimulatory adjuvant. Another approach is to use the immunostimulatory adjuvant alone. Mechanism of Action. Cancer vaccines are designed to stimulate the immune system to launch a response against the speciÞc epitope contained in the vaccine. In general, research has shown the most effective antitumor immune responses are achieved by stimulating T lymphocytes, which can recognize and kill tumor cells directly. Immunostimulatory adjuvants also activate other cells of the immune system, including macrophages, natural killer (NK) cells, and dendritic cells; these cells are capable of recognizing and destroying tumor cells. IMC-BEC2. NOTE: the most recent data available at the time of composing this reference points to the decision to discontinue BEC2. SRL-172. SRL-172 (SR Pharma) is a heat-killed suspension of Mycobacterium vaccae that achieved a potent immune response in preclinical studies when used as an adjuvant with autologous stem cells (Assersohn L, 2002). A Phase III, 418patient NSCLC trial found that adding SRL-172 to chemotherapy failed to add any survival advantage over chemotherapy alone (SR Pharma, press release, April 2002). This Þnding did not deter the company, which conducted a U.K. Phase II trial in SCLC patients that produced positive results (Assersohn L, 2002). The Phase II trial randomized 28 patients to receive chemotherapy (EP or CDE) with or without SRL-172 (Assersohn L, 2002). Toxicity was similar in both groups, but investigators observed a trend toward improved overall survival in the vaccine group. Median overall survival increased from 8.6 months in the chemotherapy alone group to 12.9 months in the chemotherapy plus SRL-172 group (p = 0.1). The trend was the same regardless of disease extent or the chemotherapy used. Although SRL-172 is still under evaluation in investigator-led studies, SR Pharma has discontinued its development and is seeking to outlicense this agent so as to concentrate on other molecules in the pipeline. Tumor-Activated Pro-Drugs Overview. ImmunoGen is pioneering a novel technology known as tumoractivated pro-drug (TAP), a method of delivering potent, cytotoxic effector molecules to tumor cells. The effector molecules are attached to MAbs that target the agent to speciÞc antigens expressed on the tumor cell. While they are in circulation, the linkage between the effector molecule and antibody is stable, but

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once inside the tumor cell, the linkage breaks down and the cytotoxic molecule is released, resulting in cell death. This method allows a higher concentration of drug to be administered safely to the patient. Mechanism of Action. The effector molecule used in ImmunoGen’s lead TAP products is DM1. DM1 is a derivative of maytansine, a potent inhibitor of cell division that can kill cancer cells at low concentrations. In vitro screening of several human cancer cell lines showed maytansines to be 100-fold to 1,000-fold more toxic than other cytotoxic drugs. huN901-DM1. The TAP huN901-DM1 (BB-10901) consists of an antibody directed against the neural cell adhesion molecule (NCAM), also known as CD56, that is linked to the cytotoxic agent DM1. This agent is in development by both ImmunoGen and Vernalis (formerly British Biotech). However, in January 2004, ImmunoGen announced it would take over future development of huN901-DM1 from Vernalis. huN901-DM1 is in Phase II clinical trials in the United States and in Phase I trials in the United Kingdom. Vernalis will continue the clinical trials that are under way but has relinquished all rights to the product (ImmunoGen, press release, January 2004). CD56 is a member of a family of cell-surface glycoproteins that plays a role in embryogenesis and contact-mediated interactions between neural cells. The molecule is expressed on a variety of normal and abnormal cells and tissues, including NK cells and lymphocyte subsets, neurons, astrocytes, Schwann cells, skin, neuroblastomas, some leukemias, and SCLC. CD56 is widely expressed in SCLC, thus making the disease an ideal candidate for this therapeutic approach. The Phase I portion of a Phase I/II trial investigating huN901-DM1 in patients with recurrent or refractory lung cancer, metastatic carcinoid tumor, or other solid tumors was completed in November 2002; the Phase II portion is under way. Thirty-two patients (18 with SCLC and 14 with neuroendocrine tumors) who had received a maximum of three prior chemotherapy regimens and had an Eastern Cooperative Oncology Group PS of 0–2 have received therapy with huN901-DM1 (Tolcher A, 2002; Fossella FV, 2002). To determine the maximal tolerated dose, patients were split into cohorts of three to six and received an IV, dose-escalating, weekly dose of 5–75 mg/m2 for four weeks, followed by a twoweek rest period. Toxicities included mild sensory neuropathy in two patients and dose-limiting pancreatitis (at the 60 mg/m2 dose). No moderate or severe (greater than grade 2) hematologic toxicity was observed, and preliminary data showed two minor responses, one each in one neuroendocrine patient and one SCLC patient. The maximal tolerated dose for Phase II trials was 60 mg/m2 for four weeks, followed by a two-week rest period. In the absence of disease progression or unacceptable toxicity, therapy is repeated every six weeks for four courses for a maximum of six courses. A Phase I study investigating a more frequent dosing schedule is under way in the United Kingdom.

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Angiogenesis Inhibitors Overview. Angiogenesis (the formation of new blood vessels) allows tumors to develop their own vascular supply, thereby replenishing the tumor cells with nourishment and enabling them to grow, invade healthy tissue, and metastasize. Numerous studies have demonstrated that higher degrees of tumor vascularization portend poorer prognosis in cancer. Mechanism of Action. Angiogenesis inhibitors reduce the vascularization of tumors, thereby depriving the tumors of the nutrients required to sustain uncontrolled growth. The tumor cells may continue to show high rates of replication, but many more cells die in the nonvascularized, hypoxic state than in a tumor with abundant vascularization. ZD-6474. AstraZeneca’s ZD-6474 inhibits the vascular endothelial growth factor (VEGF) receptor tyrosine kinase, thereby inhibiting the vascularization of tumors that is required to sustain their growth. This agent also has activity against epidermal growth factor (EGF) receptor tyrosine kinases. ZD-6474 is in development for solid tumors; Phase II trials for SCLC, NSCLC, and myeloma are ongoing. Trials for SCLC are being carried out in Canada and the United Kingdom. Patients who responded to Þrst-line chemotherapy, with or without thoracic radiation, are eligible for inclusion in a Phase II study investigating ZD-6474 versus placebo. Patients are stratiÞed according to participating center, timing of prior radiotherapy (early versus late versus no prior radiotherapy), stage of disease at diagnosis (LD versus ED), and response at study entry (CR versus PR). Patients are randomized to one of two arms to receive either oral ZD-6474 or placebo; therapy continues for up to two years or in the absence of disease progression. This study aims to recruit 120 patients and is ongoing; no data are yet available. Bevacizumab. During the Þrst quarter of 2004, the FDA approved bevacizumab (Genentech/Roche’s Avastin; rhuMAb-VEGF), a humanized MAb that binds VEGF, for the treatment of metastatic colorectal cancer in combination with 5-ßuorouracil (5-FU). The compound is in Phase II development for SCLC in the United States. The Eastern Cooperative Oncology Group and the National Cancer Institute are recruiting patients with previously untreated ED-SCLC to receive EP plus bevacizumab. The primary end points of this multicenter, Phase II study are survival, response rates, and toxicity. Recruitment began in mid 2004; no clinical data are available. Thalidomide. Thalidomide (Celgene’s Thalomid) is an immunomodulator marketed for the treatment of leprosy. Thalidomide’s spectrum of activity has not yet been fully characterized. Available data from in vitro studies and preliminary clinical trials suggest the compound’s immunological effects can vary substantially

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under different conditions, but they may be related to the suppression of excessive tumor necrosis factor-alpha (TNF-α) production and downmodulation of select cell-surface adhesion molecules involved in leukocyte migration. Via inhibition of VEGF, thalidomide has an inhibitory effect on tumor angiogenesis (Shepherd FA, 2001). The compound is under investigation for several indications, including Crohn’s disease, multiple myeloma, and colorectal cancer. An ongoing, randomized, Phase III European trial initiated in 2001 is examining the role of the cisplatin/cyclophosphamide/epirubicin/etoposide (PCEE) regimen, with and without thalidomide. This trial plans to accrue approximately 200 ED-SCLC patients. Other clinical trials involving thalidomide include an ongoing Phase III trial in the United Kingdom, sponsored by the London Lung Cancer Group (LLGC), which was initiated in June 2003. This trial aims to enroll 400 previously untreated SCLC patients randomized to receive etoposide/carboplatin/thalidomide or etoposide/carboplatin/placebo. At the time of composing this reference, the enrollment is still continuing. This trial commenced following presentation of data from the Phase II portion of this study at ASCO 2002 (Lee SM, 2002). Twenty-six chemotherapy-n¨aive patients with LD-SCLC or ED-SCLC received AUC = 5 carboplatin on day 1, 120 mg/m2 IV etoposide on day 1, and 100 mg etoposide orally twice a day on days 2 and 3, every three weeks for six cycles. They also received 100 mg oral thalidomide daily for up to two years. The overall response rate in 23 evaluable patients was 65% (9% CR, 56% PR), and 22% patients achieved SD. Toxicities included grade 3/4 neutropenia in 58% of patients and thrombocytopenia in 19%. In addition, the Ireland Cancer Center and the National Cancer Institute are sponsoring a Phase II study of thalidomide at two centers in the United States involving 30 patients with ED-SCLC who achieved a CR or PR to induction chemotherapy. Bioreductive Agents Overview. Bioreductive, or hypoxia-selective, cytotoxins represent an interesting new class of drugs in the Þeld of anticancer biologics. Tirapazamine is a lead compound in this class and has been under investigation for solid tumors, including SCLC and NSCLC. Mechanism of Action. Bioreductive agents are unique because they exhibit selective cytotoxicity toward hypoxic cells (Peters KB, 2001). This pharmacological strategy is designed to exploit the fact that tumor cells are typically less well oxygenated than healthy cells. This characteristic contributes to tumor progression and to radiotherapy/chemotherapy resistance in a variety of ways, such as activation of certain signal transduction pathways and gene regulatory mechanisms. In the human body, metabolism of the active drug turns it into a toxic free radical when the compound undergoes one-electron reduction by cytochrome P450 reductase. In hypoxic cells, the resulting free radical induces single- and

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FIGURE 18. Structure of tirapazamine.

double-strand breaks in DNA, whereas in well-oxygenated tissues, the free radical is further oxidized and transformed into an inactive metabolite. Tirapazamine. Tirapazamine (SanoÞ-Aventis’ Tirazone) (Figure 18) is an intravenously administered bioreductive drug. The National Cancer Institute is collaborating with SanoÞ-Aventis on clinical trials of tirapazamine; the compound is in Phase II trials in the United States for SCLC. Disappointing Phase III trial data in NSCLC prompted SanoÞ-Aventis to terminate development of tirapazamine for this indication. Phase II data from a 26-patient trial (SWOG-0004) of tirapazamine, delivered concurrently with the EP/radiotherapy regimen in patients with LD-SCLC, were released at ASCO 2002 (le Q-T, 2002); these data were compared with data from an 86-patient historical control that examined EP/radiotherapy alone. Grade 3/4 neutropenia was evident in approximately 50% of patients in both groups. The dose-limiting toxicity of the tirapazamine study was esophagitis, experienced by 35% of patients in the tirapazamine study and by 10% of patients in the EP study. Grade 3/4 vomiting and febrile neutropenia were also higher in the tirapazamine study. An ongoing study in the United States (SWOG-0222) is recruiting patients with previously untreated LD-SCLC to receive tirapazamine plus EP/radiotherapy over a seven-week period as induction therapy. Within 28 days of completing induction, patients will begin the consolidation phase, which consists of EP. End points include overall survival, response rates, and toxicity. Receptor Tyrosine Kinase Inhibitors Overview. Small-molecule inhibitors of receptor tyrosine kinases (RTKs) have received much attention due to the use of the EGF RTK inhibitors erlotinib (OSI Pharmaceuticals/Genentech/Roche’s Tarceva) and geÞtinib (AstraZeneca’s Iressa) in NSCLC. Mechanism of Action. RTKs play a role in signal transduction, affecting essential processes such as cellular proliferation, migration, and survival, all of which are key events in metastasis (Murray N, 2004). Inhibition of speciÞc RTKs may therefore prevent the growth and spread of SCLC.

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FIGURE 19. Structure of imatinib.

Imatinib. Novartis is developing the orally administered agent imatinib for the treatment of several cancers, including SCLC. Imatinib (Figure 19) is an RTK inhibitor with activity against a variety of RTKs, including c-kit (CD117). The c-kit receptor and its ligand, stem-cell factor, have been identiÞed as important regulators of SCLC viability and are coexpressed in 40–80% of SCLC specimens (Kijima T, 2002; Dy GK, 2002). Imatinib was Þrst launched in the United States in 2001 for the treatment of chronic myeloid leukemia; it has since been launched in Europe and Japan. The agent is also marketed in the United States for gastrointestinal stromal tumors. Imatinib is undergoing Phase II investigation in the United States for SCLC treatment. Data from three clinical trials involving imatinib in SCLC were presented at ASCO 2004. In a Phase II study, patients with ED-SCLC received four cycles of imatinib 600 mg daily plus irinotecan 60 mg/m2 on days 1, 8, and 15 plus (AUC = 4) carboplatin on day 1 at four-week intervals (Raefsky EL, 2004). Tumor cells from these patients were analyzed for c-kit expression. Of 25 evaluable patients, 21 (84%) responded to treatment, including 20 PRs and 1 CR. This combined regimen induced grade 3 toxicities of 20% neutropenia, 3% thrombocytopenia, 6% anemia, 26% nausea/vomiting, 20% diarrhea, and 23% fatigue. Grade 4 neutropenia was observed in 6% of patients. Imatinib therapy was planned for two years in patients with responding and stable disease. With a follow-up of 2–14 months, further analysis is required to determine response and survival and to clarify the impact of adding imatinib to this regimen. In addition, these parameters will be analyzed with respect to the presence and degree of c-kit expression. Imatinib was combined with the irinotecan/cisplatin (IP) regimen in a Phase I trial that also investigated the expression of potential imatinib targets (including c-kit) by immunohistochemistry (Johnson FM, 2004). Six patients with ED-SCLC received 30 mg/m2 cisplatin and 65 mg/m2 irinotecan on days 1 and 8, every 21 days for four cycles, plus 300 mg of imatinib daily during chemotherapy and then as maintenance until disease progression. G-CSF support was added to the

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protocol after the Þrst three patients suffered from signiÞcant neutropenia, one of whom died from neutropenic fever. In the remaining three patients, G-CSF was given prophylactically, preventing signiÞcant neutropenia. Other toxicities included grade 2 (n = 5) and grade 3 (n = 1) diarrhea, deep vein thrombosis (n = 1), and asymptomatic pulmonary emboli (n = 2). Four of six patients had completed four cycles and were evaluable for response. All four of these patients achieved a PR, and all six patients showed marked tumor responses after the Þrst cycle. The addition of imatinib to IP resulted in an increased AUC of irinotecan by 67%, which probably accounted for the high incidence of neutropenia and diarrhea. The authors reported that the study had been extended to separate administration of imatinib from administration of irinotecan in an attempt to optimize tolerance and drug exposure. Researchers observed c-kit expression in only two of six (33%) patients in this study. Patients who had progressive disease after receiving one or two prior chemotherapy regimens and tested positive for c-kit protein expression were eligible to enter a Phase II study in which single-agent imatinib was administered at a dose of 400 mg/m2 twice a day (Krug LM, 2004); c-kit expression was observed in 78% of patient samples. Therapy was well tolerated, with 20% grade 3 edema. However, none of the ten patients who received treatment (including six with refractory disease) responded to imatinib, and no patient stayed on treatment for more than four weeks. Single-agent imatinib also produced negative results in a Phase II trial of 19 LD-SCLC and ED-SCLC patients, who received 600 mg imatinib daily as either Þrst- or second-line therapy. The agent was well tolerated, but no objective responses were observed (Johnson BE, 2003). Researchers examined c-kit expression retrospectively and found it in only 21% of patients’ tumor samples. Results to date show no clinical beneÞt of single-agent imatinib in either the Þrst- or second-line setting. Further analysis is required to determine whether this agent improves responses or survival when added to chemotherapy regimens and whether the presence or absence of c-kit and other potential imatinib targets is important. Synthetic Retinoids Overview. Retinoids are a group of natural and synthetic vitamin A analogues that exert their effects through receptors. They have multiple downstream effects, which are being exploited for the treatment of various diseases, including diabetes, psoriasis, and cancer. Mechanism of Action. Retinoids exert their effects by binding to retinoic acid receptors (RARs) and retinoic X receptors (RXRs), ligand-activated transcription factors that control expression of genes that modulate processes such as apoptosis, cell growth, and differentiation. RARs and RXRs have different ligandbinding properties, and agents targeted against these receptors (either agonists or antagonists) can therefore affect downstream gene expression.

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Fenretinide. Fenretinide is an orally active synthetic retinoid being developed by McNeil Pharmaceuticals and the National Cancer Institute. This agent is under investigation in a variety of cancers, including bladder and ovarian cancer and SCLC. A Phase II trial for SCLC was completed in the United States. Data presented at ASCO 2004 reported results from a Phase II trial of singleagent fenretinide in 18 SCLC patients with relapsed and refractory disease after one or two prior chemotherapy regimens (Schneider BJ, 2004). Patients received 900 mg/m2 fenretinide twice a day on days 1–7 of each 21-day cycle. Although therapy was well tolerated, with no hematologic toxicity, response rates were poor: 30% of patients (n = 18) achieved SD and median survival was 25 weeks. Proteasome Inhibitors Overview. The proteasome is a novel target for cancer drugs, and clinical studies in myeloma have generated intense excitement in this Þeld. Because the proteasome interacts with many proteins (described further on), research into the downstream consequences of drugs that inhibit this target is ongoing. Mechanism of Action. Proteasomes are enzyme complexes involved in the disposal of damaged cellular proteins and the degradation of short-lived regulatory proteins that regulate cell proliferation, apoptosis, adhesion, angiogenesis, and signal transduction. Accordingly, inhibition of proteasomes can stimulate apoptosis and suppress tumor growth and spread. Regulatory proteins degraded by proteasomes include p53, p21, p27, NF-kB, I-kB, and bcl-2. Down-regulation of the NF-kB survival pathway, activated by anthracyclines, can reverse drug resistance. In preclinical studies, cancer cells appear more susceptible than normal cells to the effects of proteasome inhibition. Bortezomib. Bortezomib (Millennium’s Velcade, formerly PS-341, MLN-341, and LDP-341) is a small-molecule proteasome inhibitor in Phase II trials for SCLC. The agent has been approved in the United States and Europe for the treatment of multiple myeloma and is under investigation for non-Hodgkin’s lymphomas. Bortezomib has high selectivity for the proteasome over other proteases (e.g., thrombin) and has demonstrated in vitro cytotoxicity against a wide range of tumor cell lines. Bortezomib inhibits the 26S proteasome that works through multiple pathways, including those that inßuence apoptosis and angiogenesis. Because NF-kB transcriptionally activates bcl-2 (a protein that is overexpressed in SCLC), its inhibition may induce tumor cell apoptosis or reduce bcl-2-associated drug resistance. The SWOG and the National Cancer Institute have initiated a Phase II trial of single-agent bortezomib (1.3 mg/m2 , twice weekly for two weeks, every 21 days) in patients with recurrent or refractory SCLC who have been treated previously with platinum-based therapies. At the time of composing this reference, single agent trial seems to have been suspended.

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However, the following trial appears to be ongoing: a Randomized, NonComparative; Multicenter; Open-Label; Phase 2 study of Tarceva (Erlotinib) Alone and of Tarceva Plus VELCADE (Bortezomib) for Injection in Patients With Relapsed or Refractory, Locally Advanced or Metastatic Non-Small Cell Lung Cancer. REFERENCES Agathangelou A, et al. Methylation associated inactivation of RASSF1A from region 3p21.3 in lung, breast and ovarian tumours. Oncogene. 2001;20(12):1509–1518. Agelaki S, et al. Phase II study of the combination of gemcitabine (G) and irinotecan (I) in previously treated patients with small-cell lung cancer: a multi-center Phase II study. Proceedings of the American Society of Clinical Oncology. 2003. Abstract 2788. Agelaki S, et al. A multi-center Phase II study of the combination of gemcitabine and docetaxel in previously treated patients with small-cell lung cancer. Lung Cancer. 2004;43(3):329–333. Akerley W, et al. Phase I trial of weekly paclitaxel in advanced lung cancer. Journal of Clinical Oncology. 1998;16(1):153–158. Akerley WL, et al. SWOG 0019: a Phase II study of gemcitabine and irinotecan for patients with untreated extensive-stage small-cell lung cancer (SCLC). Proceedings of the American Society of Clinical Oncology. 2004. Abstract 7202. Alavanja MC, et al. Preexisting lung disease and lung cancer among nonsmoking women. American Journal of Epidemiology. 1992;136(6):623–632. Argiris A, Murren JR. Advances in chemotherapy for small-cell lung cancer: single-agent activity of newer agents. Cancer Journal. 2001;7(3):228–235. Armstrong J. Prophylactic cranial irradiation. 2nd International Conference on New Perspectives in the Treatment of SCLC. 2002. Presentation. Arriagada R, et al. Initial chemotherapeutic doses and survival in patients with limited small-cell lung cancer. New England Journal of Medicine. 1993;329(25):1848–1852. Asai G, et al. Phase I and pharmacokinetic study of irinotecan and paclitaxel in patients with lung cancer. Proceedings of the American Society of Clinical Oncology. 2002. Abstract 2122. Assersohn L, et al. A randomized pilot study of SRL172 (Mycobacterium vaccae) in patients with small-cell lung cancer (SCLC) treated with chemotherapy. Clinical Oncology, Royal College of Radiologists (Great Britain). 2002;14(1):23–27. Asturias Tumor Registry, 2002. Personal communication. Received October 22, 2002. Auperin A, et al. Prophylactic cranial irradiation for patients with small-cell lung cancer in complete remission. Prophylactic Cranial Irradiation Overview Collaborative Group. New England Journal of Medicine. 1999;341:476–484. Baka S, et al. Palliative treatment. Hematology/Oncology Clinics of North America. 2004;18(2):417–482. Bandera EV, et al. Alcohol consumption and lung cancer in white males. Cancer Causes Control . 1992;3(4):361–369. Beat-Ris H. No, there is no role for surgery. 2nd International Conference on New Perspectives in the Treatment of SCLC. 2002. Presentation.

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Berghmans T, et al. A prospective study of infections in lung cancer patients admitted to the hospital. Chest. 2003;124:114–120. Bilello KS, et al. Epidemiology, etiology, and prevention of lung cancer. Clinics in Chest Medicine. 2002;23(1):1–25. Blot WJ, et al. Cancers of the lung and pleura. In: Schottenfeld D, Fraumeni JF, eds. Cancer Epidemiology and Prevention. 2nd ed. New York, NY: Oxford University Press; 1996:637–665. Bray F, et al. Estimates of cancer incidence and mortality in Europe in 1995. European Journal of Cancer. 2002;38:99–166. Buchet JP, Lison D. Mortality by cancer I-n groups of the Belgian population with a moderately increased intake of arsenic. International Archives of Occupational Environmental Health. 1998;71:125–130. Chute JP, et al. Twenty years of phase III trials for patients with extensive-stage smallcell lung cancer: perceptible progress. Journal of Clinical Oncology. 1999;17(6):1794– 1801. Clark R, Ihde DC. Small-cell lung cancer: treatment progress and prospects. Oncology. 1998;12:647–658. Dally H, et al. The CYP3A4*1B allele increases risk for small-cell lung cancer: effect of gender and smoking dose. Pharmacogenetics. 2003;13(10):607–618. D’Amico D, et al. Polymorphic sites within the MCC and APC loci reveal very frequent loss of heterozygosity in human small-cell lung cancer. Cancer Research. 1992;52(7): 1996–1999. De Marinis F, et al. Phase I/II trial of gemcitabine plus cisplatin and etoposide in patients with small-cell lung cancer. Lung Cancer. 2003;39(3):331–338. De Ruysscher D, Vansteenkiste J. Chest radiotherapy in limited-stage small-cell lung cancer: facts, questions, prospects. Radiotherapy Oncology. 2000;55(1):1–9. Domine M, et al. CPT-11–gemcitabine for refractory or relapsed small-cell lung cancer (SCLC): a Spanish multi-centric Phase II study. Proceedings of the American Society of Clinical Oncology. 2003. Abstract 2819. Dongiovanni V, et al. Second-line chemotherapy with weekly paclitaxel and gemcitabine in platinum-etoposide pretreated patients with small-cell lung cancer (SCLC). Proceedings of the American Society of Clinical Oncology. 2004. Abstract 7209. Dy GK, et al. Novel targets for lung cancer therapy: Part I. Journal of Clinical Oncology. 2002;20(12):2881–2894. East Anglia Cancer Registry. Cancer incidence in East Anglia 1999. http://ftp.srl.cam.ac. uk/ciu. Accessed November 7, 2002. Eckardt JR, et al. Single agent oral topotecan (PO) versus intravenous topotecan (IV) in patients (pts) with chemosensitive small-cell lung cancer (SCLC). An international phase III study. Proceedings of the American Society of Clinical Oncology. 2003. Abstract 2488. Eckardt JR, et al. A randomized Phase III trial (389): Oral topotecan/cisplatin (TC) vs. IV etoposide/cisplatin (PE) as treatment for chemotherapy-na¨õve patients (pts) with extensive disease small-cell lung cancer (ED-SCLC): interim tolerability results. Proceedings of the American Society of Clinical Oncology. 2004. Abstract 7046. Edwards BK, et al. Annual report to the nation on the status on cancer, 1973–1999, featuring implications of age, and aging on U.S. cancer burden. Cancer. 2002;94:2766–2792.

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Allergic Rhinitis

ETIOLOGY AND PATHOPHYSIOLOGY Introduction Allergic rhinitis (AR) is clinically deÞned as a symptomatic disorder of the nose induced by an immunoglobulin E (IgE)-mediated inßammation after the membranes of the nose are exposed to an allergen (Bousquet J, 2001). In clinical settings, diagnosis of AR is usually based on the patient’s medical history, although it may be conÞrmed by allergy skin testing or serum IgE determination. Hallmark symptoms of AR include sneezing, runny nose, itchy nose, and nasal congestion; patients may also experience non-nasal symptoms such as ocular irritation and itchy palate. Sneezing, rhinorrhea, and ocular irritation usually occur immediately following exposure to allergen and constitute the early-phase (acute) response. In many (but not all) AR sufferers, the early-phase response is followed by a late-phase allergic response that results when immune and inßammatory cells are recruited to the site of inßammation (i.e., nasal tissue); the late-phase response typically takes place 3–24 hours following allergen exposure and is characterized by a recurrence of sneezing, nasal congestion, and heightened sensitivity to subsequent allergen challenge. Both the early- and late-phase allergic responses are discussed in greater detail in the following sections. Etiology Allergen Sensitization. The cascade of events leading to AR begins with initial sensitization of the cells lining the nasal mucosa to a particular allergen. Allergen sensitization occurs when a susceptible (atopic) individual is exposed Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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to sufÞcient levels of airborne allergen (e.g., pollen, animal dander, house dust mites) to prime his or her system for an exaggerated T-helper 2 (TH 2)-mediated immune response to continued challenge. Upon inhalation, the allergen (antigen) comes in contact with cells in the airway mucosa (Figure 1). Epithelial and dendritic cells, which are found in the airway mucosa, and phagocytes (e.g., tissue macrophages, granulocytes), which are found in the airway submucosa, internalize and enzymatically degrade the offending antigen into short peptide chains. The peptides associate with major histocompatibility complex II (MHC II) on the surface of these cells, which now function as antigen-presenting cells (APCs). The role of APCs is to present antigen to the lymph system—speciÞcally, to CD4+ T lymphocytes, also known as TH 0 lymphocytes. With the help of cytokines like interleukin-4 (IL-4), the TH 0 lymphocytes become differentiated into peptide (antigen)-speciÞc TH 2 lymphocytes (the phenotype commonly associated with AR). TH 2 lymphocytes then release a variety of cytokines, including IL-4, IL-5, IL-9, IL-10, IL-13, and granulocyte-macrophage colony-stimulating factor (GM-CSF). IL-4 and IL-13, in particular, stimulate B lymphocytes (plasma cells) to produce IgE molecules. These immunoglobulins (antibodies) enter the circulation and inÞltrate airways, where they bind to high-afÞnity IgE receptors (FcεRI) on mast cells, basophils, and dendritic cells. Binding of IgE to airway cells—and mast cells in particular—sets the stage for an acute response to subsequent allergen exposure. Early-Phase Allergic Response. Subsequent allergen (antigen) exposure in the nasal mucosa of a sensitized individual results in cross-linking of IgE by antigen, which in turn causes mast cell degranulation (Figure 1). Mast cell degranulation releases a variety of presynthesized and membrane-derived mediators involved in the acute-phase allergic response. Mediators include the following: • •

Histamine, which is responsible for most acute-phase symptoms. Proteases, such as tryptase, chymotryptase, carboxypeptidase, and kininogenase.

Mediators that are synthesized de novo include the following: • • •

Arachidonic acid derivatives, such as leukotrienes B4 , C4 , D4 , and E4 ; prostaglandin D2 ; and thromboxane A2 . Cytokines, such as IL-4, IL-5, IL-13, GM-CSF, and tumor necrosis factoralpha (TNF-α). Platelet-activating factor (PAF).

Concomitant neural participation occurs as well, involving the release of neuropeptides such as substance P and neurokinin A from cholinergic and peptidergic nerves. The release of the soluble and membrane-derived mediators following mastcell degranulation initiates a host of physiological changes that lead to AR’s symptoms. These physiological effects include the following:

ETIOLOGY AND PATHOPHYSIOLOGY

FIGURE 1. The allergic cascade and druggable targets.

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Increased vascular permeability and mucus production. Bronchoconstriction. Vasodilation. Activation of nociceptive neurons (via neuronal histamine [H1 ] receptors) linked to parasympathetic reßexes. Upregulation of cellular adhesion molecules (CAMs). Recruitment of immune and inßammatory cells to the site of inßammation (i.e., nasal tissue).

Many current therapies employed in the treatment of AR target speciÞc inßammatory mediators involved in the early-phase allergic response. For example, antihistamines—the most widely prescribed agents for treating AR—bind to the inactive form of the H1 receptor (one of four histamine receptors in the human body—H1 , H2 , H3 , and H4 —although H1 is the receptor through which histamine exerts most of its effects in allergic disease) and stabilize this conformation, therefore shifting the equilibrium toward the inactive state, which is unable to initiate the signal transduction cascade that ultimately results in degranulation of mast cells and basophils. Additionally, leukotriene antagonists block the ability of leukotrienes to exert their proinßammatory effects and their ability to stimulate airway secretion production, enhance eosinophil chemotaxis, and cause microvascular leakage. Thus, the resultant effect of antihistamines and leukotriene antagonists is to block the release and activity of inßammatory mediators which play key roles in the development of AR’s early-phase symptoms (e.g., sneezing, itching, rhinorrhea, and ocular irritation) (Figure 1). (Currently marketed antihistamines and leukotriene antagonists are discussed in greater detail in “Current Therapies.”) Intranasal corticosteroids are regarded as the most effective drugs available for the treatment of AR thanks to their potent and extensive anti-inßammatory effects, which include inßuencing the release of cytokines and proinßammatory mediators (e.g., IL-1, IL-4, IL-5, IL-6, IL-8, and IL-13; TNF-α; GM-CSF; and regulated upon activation, normal T-cell expressed and secreted [RANTES]), downregulating the expression of adhesion molecules and chemokines, and reducing the recruitment and accumulation of inßammatory cells (e.g., eosinophils, mast cells, basophils, APCs) to the nasal mucosa (Figure 1). Through these multiple anti-inßammatory effects, intranasal corticosteroids provide excellent relief of symptoms arising from the early-phase, as well as late-phase (discussed in the following section), allergic response. (Currently marketed intranasal corticosteroids are discussed in greater detail in “Current Therapies.”) Late-Phase Allergic Response. The late-phase allergic response is characterized by recruitment of leukocytes (e.g., eosinophils, basophils, neutrophils, T lymphocytes) to the airway tissue (submucosa, epithelium, airway lumen). There, the leukocytes accumulate and propagate the proinßammatory actions initiated by mast-cell mediators (Figure 1).

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A number of substances have been implicated in leukocyte recruitment activity. Investigators have found that IL-5 acts as a chemoattractant for eosinophils, which play a major role in the late-phase response (as described further on in this section). In addition, IL-3 and IL-5 are chemoattractants for basophils. IL-16 has speciÞc chemoattractant activity for T cells, eosinophils, and macrophages. Studies also suggest that monocyte chemotactic proteins (MCPs)—in particular, MCP-3 and MCP-4—may play a role in recruiting inßammatory cells to the nasal mucosa during AR episodes. Other evidence indicates that leukotriene B4 acts as a chemoattractant for neutrophils and, to a lesser extent, eosinophils. Additionally, IL-4, IL-13, and TNF-α increase the expression of CAMs on endothelial cells, thereby facilitating the attachment and inÞltration of leukocytes into airway tissues. CAMs play a key role in the recruitment of immune and inßammatory cells to the airway tissue by “escorting” these cells from the bloodstream across endothelium and into tissues. CAMs—such as vascular cell adhesion molecule-1 (VCAM-1), platelet endothelial cell adhesion molecule-1, intercellular adhesion molecules 1 and 2 (ICAM-1 and -2), and E- and P-selectins—expressed on endothelial cells correspond to ligands (e.g., various integrins, L-selectin) on leukocytes; they use this relationship to guide the leukocytes’ migration to the site of inßammation. The accumulation of T lymphocytes (i.e., memory T cells) and other leukocytes—especially eosinophils—and their products in the airway tissues in the hours following allergen exposure is primarily responsible for the congestion and increased mucus production characteristic of the late-phase allergic response. Indeed, nasal eosinophilia, or the accumulation of eosinophils in the nasal tissue, is a hallmark of AR. Activated eosinophils release numerous inßammatory mediators that can damage airway endothelial cells and the extracellular matrix. They may also damage neurons if they come in contact with nerve endings in the airway lumina; such damage may result in bronchial hyperresponsiveness or airway hyperreactivity upon subsequent exposure to allergens. The following products of activated eosinophils are known to be involved in the late-phase allergic response (Pearlman DS, 1999):

•

•

•

Cytokines, such as IL-1α, IL-3, IL-4, IL-5, IL-8, IL-10, TNF-α, transforming growth factor-beta-1 (TGF-β1), and GM-CSF, which can cause autocrine enhancement, sustained inßammation, leukocyte chemoattraction, and adhesion marker expression. Basic polypeptides, such as major basic protein (MBP), eosinophil-derived neurotoxin (EDN), eosinophil peroxidase (EPO), eosinophil cationic protein (ECP), and TGF, which can cause degranulation of other inßammatory cells and local tissue damage (e.g., hypersensitivity, hyperresponsiveness). Leukotriene C4 , which can promote mucus secretion, vascular permeability, and bronchoconstriction.

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Basophil activation, rather than secondary mast-cell degranulation, appears to be responsible for the increased histamine levels noted during the late-phase response because neither tryptase nor prostaglandin D2 levels (both products of mast-cell degranulation) are simultaneously increased during this phase (Christodoulopoulos P, 2000). Investigators believe that a lymphokine called histamine-releasing factor (HRF) promotes the release of histamine from basophils activated by IgE+ during the late-phase allergic response; IgE+ is a form of IgE that, when attached to basophils, speciÞcally requires HRF for basophil degranulation (and consequent release of histamine). The late-phase allergic response just described occurs in as many as 50% of all AR sufferers and is correlated with the magnitude of the initial acute reaction. That is, patients who suffer from initially severe symptoms are more likely to experience a late-phase reaction than are individuals who suffer from mild symptoms early on. The late-phase reaction usually begins within 3–6 hours of allergen challenge, peaks at 6–8 hours, and subsides 12–24 hours after allergen challenge; it will, however, persist in the continuing presence of the offending allergen. Over time, patients who experience the late-phase response tend to grow more sensitive to nonallergic triggers such as cold air and smoke, simply because they have a higher degree of nasal hyperreactivity. Researchers hypothesize that the sensory nerves (C Þbers) in the nasal mucosa of AR sufferers exhibit a heightened responsiveness to allergens and other irritants, which in turn increases their propensity for neurogenic inßammation via neuropeptide (e.g., substance P, neurokinin A) release upon activation by inhaled particles (Togias A, 2000). Ultimately, a chronic inßammatory process such as that noted in AR could result in pathologic tissue remodeling. This remodeling may involve permanent modiÞcation of the histological and functional structure of this tissue. Researchers have shown that nonasthmatic AR patients have increased subepithelial deposition of type I and II collagens within their bronchial submucosa as compared with healthy controls. As stated previously, intranasal corticosteroids provide excellent relief from AR symptoms characteristic of the early-phase allergic response—rhinorrhea, sneezing, and itching nose. However, due to their extensive and potent antiinßammatory effects—especially their ability to downregulate adhesion molecule and chemokine expression and to reduce the leukocyte recruitment and accumulation in the nasal mucosa—intranasal corticosteroids are effective in reducing nasal congestion, which is characteristic of the late-phase allergic response (Figure 1). As a result, intranasal corticosteroids are recommended for Þrst-line use in AR patient with frequent or persistent symptoms or patients suffering from nasal congestion (Bousquet J, 2001). Predisposing Factors. Atopy.. The term atopic is used to describe individuals who display an inherited tendency to develop IgE-mediated immune responses that place them at risk for developing allergies after exposure to environmental allergens. IgE is the key

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antibody involved in potentiating both allergic and asthmatic reactions. Notably, levels of IgE are higher in atopic individuals than in nonatopic individuals. Diagnostic indicators of atopy include positive radioallergosorbent (RAST) tests, nasal eosinophilia, and the presence of ECP in nasal secretions. Although all AR sufferers are classiÞed as atopic, not all atopic individuals necessarily develop allergies. In other words, an atopic individual must be exposed to high enough levels of allergen to trigger an allergic reaction; if such a person is never exposed in this way, he or she will not suffer from the overactive immune response characteristic of AR. Studies estimate that atopy is present in 20–30% of the general population and in 10–15% of children (Fireman P, 2000; Nimmagadda SR, 1999). However, family history of atopy is one of the most important risk factors for developing AR. The risk of atopy is heightened in children with two atopic parents, especially if both parents’ atopy is manifested similarly (e.g., allergic rhinitis, asthma). Approximately 50% of children with two atopic parents are atopic themselves, and this percentage increases to 72% for children of patents with the same atopic manifestation (Fireman P, 2000; Nimmagadda SR, 1999). Lifestyle and Environment: The Hygiene Hypothesis. A widely held theory suggests that the increasingly sterile environment in which many people live in developed countries has greatly contributed to the rise in AR prevalence in these countries (as opposed to the situation found in many developing nations). In the industrialized world, rather than being exposed to a variety of childhood illnesses that allow their immune systems to mature and possibly develop immune tolerance, children tend to live in more protected environments. In these environments, they fail to encounter challenges to their immune systems (i.e., germs, microbes) that might stimulate a more mature immune response (via TH 1 lymphocytes) and possibly suppress the neonatal TH 2 lymphocytic response commonly associated with allergies (Figure 2). Researchers believe that the TH 1-like phenotype of T-cell-mediated immune responses is less likely to favor allergen sensitization than is the TH 2 phenotype. The aforementioned scenario is known as “the hygiene hypothesis.” It is based on studies indicating that exposure to microbial pathogens in early life offers protection against overzealous immune reactions to harmless offenders such as allergens later in life. In a study conducted in Italian military recruits, researchers found a negative correlation between atopy and childhood infection with hepatitis A (Matricardi PM, 1997). The same group published subsequent Þndings suggesting that respiratory allergy occurs less often in individuals who are repeatedly exposed to various orofecal and foodborne microbes (e.g., Helicobactor pylori, Toxoplasma gondii , hepatitis A) (Matricardi PM, 2000). Other studies suggest that factors such as attendance at day care facilities in infancy (Ball TM, 2000) or daily exposure to farm animals in early life (Riedler J, 2000) may help protect children from developing asthma and/or allergies in later years. In essence, researchers have found that the more exposure a child has to foreign microbes early in life, the better his or her chance of avoiding diseases

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FIGURE 2. The hygiene hypothesis.

involving hyperresponsive TH 2-mediated immune mechanisms (such as AR and asthma) later in life. Pathophysiology Disease Classification. Traditionally, AR has been classiÞed as either seasonal or perennial. Individuals who suffer from seasonal AR (SAR) are typically allergic to outdoor allergens (e.g., tree, grass, and weed pollens; mold spores) and experience symptoms at speciÞc times of the year when outdoor allergen levels are high. Patients who suffer from perennial AR (PAR) are typically allergic to allergens found indoors (e.g., house dust mites, animal dander, feces or skeletal debris from insects, mold spores/fungi) and can become symptomatic at any time of the year, depending on their exposure to allergens that are present in the environment year-round. Although different allergens are responsible for initiating SAR than are implicated in PAR, the symptoms and treatment approaches for both types of AR are basically the same. In 2001, the Allergic Rhinitis and Its Impact on Asthma (ARIA) report was published in collaboration with the World Health Organization (WHO) (Bousquet J, 2001). In addition to providing recommendations for the diagnosis and management of AR using an evidence-based approach, the ARIA report proposes alternative disease classiÞcation criteria—“intermittent” or “persistent” AR—to the more traditional SAR or PAR classiÞcation. According to the ARIA report, intermittent AR patients suffer symptoms for less than four days per week and for less than four weeks per year, while AR is considered persistent when the symptoms are present for more than four days per week and for more than four weeks per year. Figure 3 outlines the characteristics of SAR, PAR, intermittent AR, and persistent AR.

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FIGURE 3. Characteristics of seasonal, perennial, intermittent, and persistent allergic rhinitis.

In addition to being classiÞed as having SAR or PAR (or intermittent or persistent AR), patients with AR are often classiÞed as having mild, moderate, or severe disease. Although disease severity is typically linked to the severity of a patient’s symptoms and the degree to which symptoms affect their life, there are no uniform, deÞnitive deÞnitions of what constitutes mild, moderate, or severe AR among various treatment guidelines that exist within the seven countries under study (United States, France, Germany, Italy, Spain, United Kingdom, and Japan). For example, according to the ARIA guidelines, AR is characterized as mild when a patient experiences no sleep disturbance, no impairment of daily activities (e.g., sports, leisure, school, work), and no trouble-some symptoms,

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whereas it is moderate to severe when a patient experiences one or more of the following: abnormal sleep, impairment of daily activities, and troublesome symptoms (Figure 3). A similar classiÞcation is put forth in the European Academy of Allergy and Clinical Immunology (EAACI) guidelines (van Cauwenberge P, 2000), while Japanese allergy guidelines measure severity against three main symptoms: sneezing, nasal secretion, and nasal airßow obstruction (Nose Allergy Clinical Practice Guidelines, 2002). Link Between Allergic Rhinitis and Asthma. The link between AR and asthma has been repeatedly observed in the past 50 years, but the overall understanding of the pathogenesis of respiratory allergy has profoundly changed over the past decade. An increasing amount of epidemiological, pathophysiological, and clinical data indicate the following: • • •

The frequent coexistence of AR and asthma. The possible role of AR in the onset and evolution of asthma. The association of AR and nonspeciÞc bronchial hyperreactivity (Leynaert B, 2000; Bousquet J, 2001).

The same allergens that trigger AR can cause asthma. Studies from around the world implicate sensitivity to house dust mites as the most consistent allergenspeciÞc risk factor for the development of asthma in AR sufferers. Interestingly, asthma appears to be more often associated with PAR than with SAR, a link partially explained by the common allergen. Epidemiological studies have consistently shown that asthma and AR often coexist in the same patient (Sibbald B, 1991[a]; Houvinen E, 1999). AR has been described in more than 75% of patients with allergic asthma and in more than 80% of patients with nonallergic asthma (Sibbald B, 1991[a]; Leynaert B, 2000). Studies have also identiÞed a temporal relationship between the onset of AR and asthma: AR frequently precedes the development of the lower airways disease. In 1994, A.L. Wright and colleagues suggested that rhinitis that develops in the Þrst years of life is an early manifestation of an atopic predisposition and a risk for asthma. A study examining the coexistence of asthma and AR found, after a 23-year follow-up, that 48% of individuals with a clinical history of both diseases experienced the development of AR Þrst; 34.5% experienced the development of asthma Þrst; and 20.7% experienced the development of both diseases at the same time (Settipane RJ, 1994; Greisner WA, 1998). These data support the hypothesis that AR and asthma may be considered one entity, designated allergic rhinobronchitis or united airways disease (UAD), induced by a common inßammatory process (Simmons FE, 1994; Passalacqua G, 2000). Additionally, one study conducted by G.J. Braunstahl and colleagues demonstrated that nasal allergen provocation induced adhesion molecule expression and tissue eosinophilia in upper and lower airways, further supporting the hypothesis that local allergen exposure results in generalized airway inßammation (Braunstahl GJ, 2001).

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FIGURE 4. The link between allergic rhinitis and asthma.

The inßammatory process that takes place in the upper respiratory epithelium (i.e., nasal passageways) of AR sufferers is fundamentally the same as that noted in the lower respiratory epithelium (i.e., bronchial tubes) of asthma sufferers. Therefore, the difference in clinical presentation of disease is due primarily to the different functions of the end organs affected by the respective conditions. In asthma, this pathway is associated with bronchoconstriction (via smooth-muscle contraction in the lower airways) rather than nasal symptoms because of the distinct histological characteristics of the tissues involved. The end result of this inßammatory process is sneezing, congestion, and rhinorrhea in patients whose upper airways are affected, and coughing, breathlessness, and sputum production in those whose lower airways are affected. Researchers have yet to elucidate the speciÞc mechanisms through which AR inßuences the clinical course of asthma (i.e., why some AR sufferers become asthmatic and others do not). Nevertheless, numerous hypotheses have been proposed. Some investigators suggest that the nasal blockage experienced by many AR patients leads to increased mouth breathing, which in turn leads to increased airway exposure to “unÞltered” allergens and ensuing asthmatic symptoms (Assanasen P, 2001). Others suggest a role for AR-related postnasal drip, on the grounds that this problem may allow inßammatory mediators to spread to the lower airways. Alternatively, as shown in Figure 4, the stimulation of sensory nerves and nasobronchial reßexes noted in AR may lead to bronchoconstriction and consequent asthma (Nolte ND, 1983).

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Whatever the mechanisms involved, the link between AR and asthma is strengthening. In fact, drugs used to treat AR often ameliorate asthmatic symptoms in atopic individuals, and drugs for asthma have demonstrated efÞcacy in AR. For example, studies have shown that the administration of intranasal corticosteroids (in sufÞciently high doses) not only decreases nasal symptoms but also improves asthma symptoms, pulmonary function tests, and bronchial hyperresponsiveness of patients with AR and concomitant asthma. Studies also support the beneÞcial effects of histamine (H1 ) antagonists, which are generally reserved for use as AR therapy, in treating patients with coexisting mild seasonal asthma symptoms. Additionally, the leukotriene antagonists montelukast (Merck’s Singulair) and pranlukast (Ono’s Onon), initially approved as therapies for asthma, are now indicated for AR. Also, the anti-IgE monoclonal antibody omalizumab (Genentech/Novartis/Sankyo/Tanox Biosystems’ Xolair) is marketed in the United States for patients with moderate to severe persistent asthma who have demonstrated reactivity to a perennial aeroallergen. (The leukotriene antagonists and omalizumab are discussed in greater detail in “Current Therapies” and “Emerging Therapies,” respectively.) In addition to providing recommendations for the diagnosis and management of AR and deÞning alternative disease classiÞcation criteria, the ARIA report emphasizes the fact that AR is correlated with—and constitutes a risk factor for—the occurrence and severity of asthma. The report endorses the view that upper and lower airways must be considered a unique entity inßuenced by a common, evolving inßammatory process, and that inßammation of the lower airway may be sustained and ampliÞed by interconnected mechanisms occurring during AR. The ARIA report suggests that the optimal management of AR may therefore partially improve coexisting asthma. Additionally, as asthma patients with concomitant AR have been shown to have higher medication costs than patients suffering from asthma alone (Yawn BP, 1999), appropriate treatment of AR in asthma patients my reduce overall treatment costs. CURRENT THERAPIES The main goal of pharmacotherapy used in the treatment of allergic rhinitis (AR) is alleviating the hallmark symptoms of the disease (e.g., sneezing, rhinorrhea [runny nose], nasal congestion, nasal itching, ocular irritation). Although the triggers and allergic response durations may differ for patients with seasonal AR (SAR) and perennial AR (PAR), the symptoms of the two types of the disease are largely the same. Thus, similar agents are used to treat both forms of the disease. Numerous therapies, including antihistamines, intranasal corticosteroids, leukotriene antagonists, decongestants, anticholinergic agents, and mast cell stabilizers, are employed in the treatment of AR symptoms. While currently available AR therapies collectively are very effective in controlling the symptoms of the disease, certain drug classes are better than others in relieving particular AR symptoms. For example, antihistamines are effective in relieving many symptoms associated with mild to moderate AR (e.g., sneezing, rhinorrhea, nasal

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itching, ocular irritation), but these agents are less effective than intranasal corticosteroids in treating more-severe disease involving prolonged inßammation, and are less effective at controlling nasal congestion. In addition to differing efÞcacy for particular AR symptoms, current therapies also differ in routes of administration and dosing frequencies. Thus, physicians must balance desired efÞcacy with convenience when recommending pharmacotherapy to an AR patient. Table 1 summarizes the leading therapies available to treat AR in the seven major markets under study (United States, France, Germany, Italy, Spain, United Kingdom, and Japan), and Table 2 summarizes their relative advantages and disadvantages. Second-generation antihistamines and intranasal corticosteroids are the most commonly prescribed AR therapies, and there are numerous marketed products within these drug classes. Thus, the discussion here focuses primarily on agents within these two drug classes. The less numerous leukotriene antagonists are also discussed in greater detail in the following sections. Mast cell stabilizers and anticholinergics are also used in the treatment of AR symptoms. Mast cell stabilizers (e.g., cromolyn sodium [generics; also referred to as cromoglicic acid, disodium cromoglycate, or nedocromil sodium], pemirolast [Nikken Chemical’s Alegysal, generics], tranilast [Kissei’s Rizaban, generics]) are anti-inßammatory agents that address the symptoms of AR by preventing degranulation of mast cells and histamine release and, subsequently, synthesis of other inßammatory mediators. However, these agents require administration several times daily and take a period of days to weeks to reach maximum effect. Products containing cromolyn sodium are readily available over-the-counter (OTC) in all countries under study (AESGP, 2004[a]; AESGP, 2004[b]). The anticholinergic agent ipratropium bromide (Boehringer Ingelheim’s Atrovent) effectively treats rhinorrhea by inhibiting secretions from the serous and seromucous glands lining the nasal mucosa, but has no effect on any other nasal symptoms. Also, like mast cell stabilizers, ipratropium bromide requires administration multiple times daily. Because of their frequent dosing schedules and inferior efÞcacy compared with other commercially available allergy medications, mast cell stabilizers and anticholinergics are not widely prescribed for AR, so these drug classes are not discussed in detail here. Decongestants—administered orally or intranasally—are also routinely used in the treatment of nasal AR symptoms. (Ocular decongestant formulations can be used to treat eye redness and irritation associated with AR.) Therapies within this class are sympathomimetic agents that work by stimulating α-adrenergic receptors in vascular smooth muscle tissue. Oral and intranasal decongestants shrink mucous membranes and promote airßow and drainage, effectively reducing nasal congestion and, to a lesser extent, rhinorrhea. However, these agents do not address other common AR symptoms such as sneezing or itching. Consequently, oral and intranasal decongestants are most effective when used in combination with an antihistamine to treat AR. Decongestant products are readily available OTC, and many second-generation antihistamines are now formulated with a decongestant as a Þxed-combination product. Thus, this discussion of decongestants in the treatment of AR will be

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TABLE 1. Current Therapies Prescribed for Allergic Rhinitis Agent

Company/Brand

Daily Dose

Availability

60 mg bid or 180 mg qd (for patients aged 12 years or older); 30 mg bid (for patients aged 6–11 years) 5–10 mg qd (for patients aged 6 years or older); 2.5 mg bid or 5 mg qd (for patients aged 2–5 years); 2.5 mg bid or qd (for patients aged 6–23 months) 5 mg qd (for patients aged 12 years and older); 2.5 mg qd (for patients aged 6–11 years); 1.25 mg qd (for patients aged 1–5 years); 1.0 mg qd (for patients aged 6–11 months) 5 mg qd (for patients aged 6 years or older)

US, F, G, I, S, UK, J

Second-generation antihistamines Fexofenadine +/− pseudoephedrine

Sanofi-Aventis’ Allegra/Telfast, Allegra-D 12 Hour

Cetirizine +/− pseudoephedrine

Pfizer/UCB/Daiichi/ Sumitomo’s Zyrtec/Virlix/ Cirrus, Zyrtec-D 12 Hour, generics

Desloratadine +/− pseudoephedrine

Schering-Plough’s Clarinex/ Aerius/ Neoclarityn, Clarinex-D 24 Hour

Levocetirizine

UCB’s Xyzal/Xusal

US, F, G, I, S, UK, J

US, F, G, I, S, UK

F, G, I, S, UK

Intranasal corticosteroids Fluticasone propionate

GlaxoSmithKline’s Flonase/ Flixonase, generics

Mometasone furoate

Schering-Plough’s Nasonex

2 sprays (50 µg per spray) in each nostril qd or one spray in each nostril bid (for patients aged 12 years or older); one spray (50 µg per spray) in each nostril qd (for patients aged 4–11 years) 2 sprays (50 µg per spray) in each nostril qd (for patients aged 12 years or older); one spray (50 µg per spray) in each nostril qd (for patients aged 2–11 years)

US, F, G, I, S, UK, J

US, F, G, I, S, UK

Leukotriene antagonists Montelukast

Merck’s Singulair

10 mg qd (for patients aged 15 years or older); 5 mg qd (for patients aged 6–14 years); 4 mg qd (for patients aged 2–5 years)

US, F, G, I, S, UK, Ja

a Montelukast is approved for the treatment of asthma in all seven countries under study. As of this writing,

however, the drug is only indicated for treating seasonal allergic rhinitis in the United States and the United Kingdom. bid = Twice daily; qd = Once daily. US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan.

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TABLE 2. Comparison of Current Therapies for Allergic Rhinitis Compound

Advantages

Second-generation antihistamines Fexofenadine

Cetirizine

Desloratadine

• Classified as a non-drowsy antihistamine, with no label precautions cautioning against driving while taking the drug. • Fixed fexofenadine/pseudoephedrine combination product is available in a 12-hour formulation, and a 24-hour formulation has been approved by the FDA. • Currently marketed in all seven countries under study.

Disadvantages • Label does not include PAR as an approved indication. • Only approved for children as young as age six. • Only available in oral tablet or capsule formulations.

• Indicated for both SAR and PAR. • Approved for use in children as young as six months in age. • Available in oral tablet, chewable tablet, and oral syrup formulations. • Lower-priced generic versions are currently available in most European countries. • Currently marketed in all seven countries under study.

• Not classified as a ‘‘nonsedating’’ antihistamine. • Due to the higher potential for somnolence, product label includes a warning stating that patients taking cetirizine should use caution when driving, operating machinery, or consuming alcohol or other agents that can depress the CNS. • Fixed cetirizine/pseudoephedrine combination product is only available in a 12-hour formulation.

• Classified as a non-drowsy antihistamine, with no label precautions cautioning against driving while taking the drug. • First second-generation antihistamine to demonstrate proven, consistent efficacy in relieving nasal congestion. • Indicated for both SAR and PAR. • Approved for use in children as young as six months in age. • Available in oral tablet, rapidly disintegrating tablet, and oral syrup formulations. • Fixed desloratadine/pseudoephedrine combination product is available in a 24-hour formulation.

• Currently not marketed in Japan.

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TABLE 2. (continued) Compound Levocetizine

Advantages • Indicated for both SAR and PAR. • First second-generation antihistamine to be indicated for PER, which reflects the new classification put forth in the ARIA report. • Has demonstrated proven, consistent efficacy in relieving nasal congestion.

Intranasal corticosteroids • Superior efficacy in relieving nasal Intranasal symptoms compared with oral fluticasone second-generation antihistamines. propionate • Significantly reduced systemic bioavailability compared with other intranasal corticosteroid products. • Currently marketed in all seven countries under study. Intranasal mometasone furoate

• Superior efficacy in relieving nasal symptoms compared with oral second-generation antihistamines. • Significantly reduced systemic bioavailability compared with other intranasal corticosteroid products. • Indicated for children as young as age two. • Only currently available intranasal corticosteroid that is indicated for the prophylaxis of nasal symptoms in SAR patients.

Disadvantages • Only approved for children as young as age six. • Product label acknowledges that no evidence from comparative clinical trials has indicated levocetirizine negatively affects mental alertness, reactivity, or ability to drive, but the label does state patients should consider their potential response to the drug if they intend to drive, operate machinery, or participate in hazardous activities, as sedating effects have been noted in clinical trials involving levocetirizine. • Only available in oral tablet formulation. • Currently not marketed in the United States and Japan. • Indicated for children as young as age four. • Must be delivered via the nasal route, which is less desirable to patients than oral formulations. • ‘‘Steroid stigma’’ precludes some patients from using this agent unless absolutely necessary. • Must be delivered via the nasal route, which is less desirable to patients than oral formulations. • ‘‘Steroid stigma’’ precludes some patients from using this agent unless absolutely necessary. • Currently not marketed in Japan.

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TABLE 2. (continued) Compound

Advantages

Leukotriene antagonists Montelukast

• Due to its alternative mechanism of action (leukotriene inhibition), the drug is often used in combination with oral second-generation antihistamines for patients who fail to achieve adequate symptom relief with antihistamine monotherapy and who seek an alternative to intranasal corticosteroids. • Indicated for AR in children as young as age two. • Available in oral tablet and chewable tablet formulations for AR patients. (An oral granule formulation is also available, but is only indicated for use in treating asthma in children ages 12-23 months.)

Disadvantages • Efficacy is only comparable to oral second-generation antihistamines, and has been shown to be inferior to that of intranasal fluticasone propionate. • Marketed for asthma in all countries under study, but only formally approved for SAR in the United States and the United Kingdom.

AR = Allergic rhinitis; ARIA = Allergic Rhinitis and Its Impact on Asthma; CNS = Central nervous system; PAR = Perennial allergic rhinitis; PER = Persistent allergic rhinitis; SAR = Seasonal allergic rhinitis.

limited to their mention as Þxed-combination products with second-generation antihistamines; decongestants are not discussed as a separate class here. Second-Generation Antihistamines Overview. Antihistamines have been employed in the treatment of AR since the discovery of the Þrst histamine receptor antagonists in 1937 (Slater JW, 1999). First-generation antihistamines—including diphenhydramine (PÞzer’s Benadryl, generics), chlorpheniramine (Schering-Plough’s Chlor-Trimeton, generics), and brompheniramine maleate (Wyeth’s Dimetane, generics)—are very effective in treating many common AR symptoms (e.g., rhinorrhea, sneezing, itchy nose, itchy eyes). However, Þrst-generation antihistamines are highly lipophilic and can readily cross the blood-brain barrier. Once they penetrate the CNS, Þrst-generation antihistamines bind H1 receptors, as well as cholinergic, dopaminergic, and serotonergic receptors, in the brain. This action triggers undesirable and troublesome CNS side effects, including fatigue, drowsiness, performance impairment, and anticholinergic effects (e.g., dryness of the mouth and eyes, constipation, increased ocular pressure). Since 1981, numerous second-generation antihistamines (e.g., fexofenadine [SanoÞ-Aventis’ Allegra/Telfast], cetirizine [PÞzer/UCB/Daiichi//Sumitomo’s Zyrtec/Virlix/Cirrus, generics], desloratadine [Schering-Plough’s Clarinex/ Aerius/Neoclarityn], levocetirizine (UCB’s Xyzal/Xusal]) have entered the

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market, and these agents have all but replaced Þrst-generation antihistamines in the prescription AR market. Second-generation antihistamines are equally effective in controlling the common symptoms of AR, but they offer numerous advantages over their predecessors. Because second-generation antihistamines are larger molecules, possess a different ionic charge, and are more lipophobic than the Þrst-generation agents, they do not cross the blood-brain barrier as readily as the original antihistamines. Thus, they lack the unwanted CNS effects of the Þrst-generation agents. Additionally, second-generation antihistamines are highly speciÞc for histamine H1 receptors, with little afÞnity for cholinergic, dopaminergic, and serotonergic receptors. As a result, they cause mild or no sedation, do not affect performance, and have no anticholinergic effects. The discussion here focuses on four of the top-selling second-generation antihistamines—fexofenadine, cetirizine, desloratadine, and levocetirizine—as representatives of this crowded class. These four antihistamines offer comparable efÞcacy. Some of the product characteristics that differ between agents include the following: indications and age of patients for which each drug is approved; formulations in which the drug is available; and the potential to cause sedation. These characteristics are discussed in greater detail for each drug in the following sections and are outlined in Table 2. Mechanism of Action. There are four types of histamine receptors in the human body—H1 , H2 , H3 , and H4 —although H1 is the receptor through which histamine exerts most of its effects in allergic disease. The H1 receptor demonstrates constitutive activity in the human body—even in the absence of ligand binding—because two states of the H1 receptor (the active state and the inactive state) exist in equilibrium (Simons FER, 2004). Histamine preferentially binds to the active form of the receptor, shifting the balance between the two receptor states toward the active form. The active form of the receptor then initiates a G protein-mediated signal transduction cascade that results in the degranulation of FcεRI+ cells. Antihistamines bind to the inactive form of the H1 receptor and stabilize this conformation, thereby inhibiting the signal transduction cascade that ultimately results in degranulation of mast cells and basophils. H1 antihistamines, previously viewed as H1 receptor antagonists, have been reclassiÞed as inverse agonists owing to enhanced understanding of their molecular pharmacology (Simons FER, 2004). The degranulation of mast cells and basophils initiates a host of physiological changes leading to AR’s early-phase symptoms (e.g., sneezing, itching, rhinorrhea, ocular irritation). However, antihistamines are less effective at controlling nasal congestion, as histamine is not the primary mediator of this symptom. Instead, mediators such as kinins, prostaglandins, leukotrienes, and, in the latephase allergic response, leukocyte inÞltration are more intimately involved in nasal congestion. Numerous in vitro and in vivo studies have shown that antihistamines possess a range of anti-inßammatory properties, including down-regulation of adhesion protein expression, inhibition of eosinophil and other inßammatory cell migration

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and activation, and inhibition of the generation and activity of various proinßammatory cytokines and mediators (Simons FER, 2004). However, further studies are needed to determine the clinical relevance of these anti-inßammatory effects in AR patients during natural exposure to allergens at typical therapeutic doses of oral antihistamines. Formulation. Most second-generation antihistamines are delivered orally; the exceptions are azelastine (MedPointe’s Astelin, Viatris’ Allergodil/Rhinolast/ Aßuon) and levocabastine (Janssen Cilag’s [a Johnson & Johnson subsidiary] Livostin), which are administered intranasally. Because of patient preference for oral formulations, azelastine and levocabastine are used far less frequently than oral antihistamines for the treatment of AR. Fexofenadine. The oral second-generation antihistamine fexofenadine (Allegra/Telfast) (Figure 5) is marketed in all countries under study by SanoÞ-Aventis. The drug was originally launched in the United States in 1996 by Hoechst Marion Roussel (HMR). (HMR merged with Rhone-Poulenc Rorer in 1999 to form Aventis, and Aventis merged with SanoÞ-Synth´elabo in 2004 to form SanoÞ-Aventis.) The drug was subsequently launched in its Þrst European market, the United Kingdom, in 1998 and in Japan in 2000. Fexofenadine, which can be administered once- or twice-daily, is indicated for the treatment of SAR and chronic idiopathic urticaria (CIU; hives) in all regions under study, and it is also indicated for dermatitis and pruritis (itching) in Japan. Fexofenadine is approved for use in children as young as age six. The drug was originally developed by Sepracor, but the company licensed its U.S. patent to the drug to HMR in 1993. Sepracor later licensed remaining worldwide patents to HMR in 1999. As a result of these agreements, Sepracor now earns royalties from SanoÞ-Aventis. In 1998, HMR launched a 12-hour version of Allegra-D, a Þxed combination of fexofenadine and the decongestant pseudoephedrine, in the United States for relief of SAR symptoms in patients aged 12 years and older. SanoÞ-Aventis received FDA approval for a once-daily, 24-hour formulation of Allegra-D in October 2004. Fexofenadine is the active metabolite of terfenadine (HMR’s Seldane/ Teldane/Triludan), one of the Þrst second-generation antihistamines to enter the

FIGURE 5. Structure of fexofenadine.

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AR market. Terfenadine has the potential to cause serious or life-threatening cardiac events such as QTC prolongation and ventricular arrhythmias when taken at high doses or coadministered with other drugs metabolized by the cytochrome P450 system (e.g., macrolide antibiotics, azole antifungals), leading to an accumulation of terfenadine in a patient’s plasma. Consequently, terfenadine was withdrawn from the French and U.S. markets in 1997 and 1998, respectively. The European Commission recommended the withdrawal of high-dose terfenadine tablets in 1998, and further recommended that additional label warnings be added to low-dose formulations of the drug. In all countries under study where terfenadine remains on the market, the drug’s follow-on compound, fexofenadine, which does not cause these cardiac adverse events, has replaced its predecessor. Like other second-generation antihistamines proÞled in this section, fexofenadine’s predominant therapeutic effects are exerted through the same means as discussed in the “Mechanism of Action” section for the drug class. Numerous studies have demonstrated fexofenadine’s efÞcacy in relieving AR symptoms. In one two-week, randomized, placebo-controlled trial, 572 patients with moderate to severe ragweed SAR were treated with one of three fexofenadine doses (60 mg, 120 mg, or 240 mg) or placebo twice-daily (Bernstein DI, 1997). At the end of the 14-day treatment period, all three fexofenadine treatment groups demonstrated signiÞcantly greater improvements in the primary efÞcacy measure—patient-assessed evening reßective total symptom scores (TSS)—compared with placebo recipients. (The evening-reßective TSS was the sum of individual symptom scores for the previous 12 hours, assessed by the patient prior to receiving their second daily dose of study medication. Symptoms included in the sum were sneezing; rhinorrhea; itchy nose, palate, and/or throat; and itchy, watery, and red eyes.) Patients treated with placebo demonstrated a mean 16.9% improvement over baseline evening reßective TSS, compared with mean improvements of 28.1%, 25.5%, and 28.1% for the three fexofenadine groups (60 mg, 120 mg, and 240 mg twice-daily, respectively). Additionally, all three fexofenadine treatment groups demonstrated signiÞcantly greater improvements over placebo in evening-reßective scores for all individual symptoms assessed; the smallest mean improvements were noted for nasal congestion. Fexofenadine was well tolerated in this study, and the incidence of adverse events was 14.2%, 6.9%, and 11.7% for the 60 mg, 120 mg, and 240 mg fexofenadine twice-daily groups, respectively, compared with 9.2% for the placebo group. The adverse events most commonly reported in the study were headache (2.8%, 0.7%, 0.7%, and 2.8%, respectively) and leucopenia (reduced white blood cell count; 2.1%, 0.7%, 1.4%, and 0.7%, respectively). Treatment with fexofenadine, regardless of dose, was not associated with any electrocardiographic abnormalities. The 180 mg once-daily dose of fexofenadine has been shown to offer efÞcacy comparable to 10 mg once-daily cetirizine (Zyrtec). In a two-week study conducted by F. Hampel and colleagues involving 495 patients with moderate to severe SAR, patients treated with fexofenadine demonstrated a mean 19% improvement over baseline reßective TSS compared with a mean improvement

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of 21.6% for cetirizine-treated patients. These improvements were considered statistically equivalent, as were the improvements for the two treatment groups on the measures of morning instantaneous TSS (sum of patient-assessed scores for symptom severity over the previous hour) and morning instantaneous scores for individual symptoms (Hampel F, 2003). Another key end point examined in the Hampel study was the difference between fexofenadine and cetirizine’s effects on drowsiness and motivation. (Although cetirizine is a second-generation antihistamine, the drug is not classiÞed as a “nonsedating” antihistamine like fexofenadine, because sedative effects have been noted with use of cetirizine in clinical studies. Cetirizine’s sedative effects are discussed in greater detail in the section devoted to this drug.) In this study, treatment with fexofenadine was associated with signiÞcantly greater improvements over baseline drowsiness scores, as measured on a 100-point visual analog scale (VAS), compared with cetirizine at all time points assessed—7 a.m., 10 a.m., and 3 p.m. Treatment with fexofenadine was also associated with a trend toward greater improvement in motivation scores, as measured on a 100-point VAS, compared with cetirizine, but the between-group difference on this measure did not reach statistical signiÞcance (Hampel F, 2003). The results from the Hampel study were comparable to those from another large-scale study comparing the efÞcacy and safety of fexofenadine and cetirizine in SAR patients. A study conducted by P.H. Howarth and colleagues determined that there was no signiÞcant difference between patients treated with fexofenadine and cetirizine on the measures of 24-hour reßective TSS, 24-hour reßective individual symptom scores, and 24-hour instantaneous TSS. Also, the incidence of drowsiness or fatigue was signiÞcantly higher for cetirizine-treated patients (8.6%) compared with fexofenadine-treated patients (4.0%); the incidence of drowsiness or fatigue for fexofenadine-treated patients was comparable to that for placebo recipients (4.3%) (Howarth PH, 1999). Fexofenadine has also demonstrated efÞcacy comparable to that of loratadine (Schering-Plough/Shionogi’s Claritin/Clarityn, generics) in improving 24-hour reßective TSS in a two-week study involving 688 SAR patients. Patients in the fexofenadine group demonstrated signiÞcantly superior improvements over baseline on the measures of nasal congestion and itchy, watery, red eyes compared with patients treated with either loratadine or placebo (van Cauwenberge P, 2000[a]). Fexofenadine has also demonstrated signiÞcantly superior efÞcacy in relieving nasal congestion compared with placebo in some studies, but this beneÞcial effect has not been consistently demonstrated in all trials. In a meta-analysis of six placebo-controlled, double-blinded, randomized studies evaluating the efÞcacy of fexofenadine in patients with moderate to severe SAR, fexofenadine-treated patients demonstrated signiÞcantly superior improvements in nasal congestion scores compared with placebo in three studies, but a signiÞcant difference was not noted in three other studies (Meeves SG, 2000; Meeves SG, 2003). The product label for fexofenadine reports that the incidence of adverse events among nearly 2,500 AR patients aged 12 years or older treated with 20–240 mg

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fexofenadine twice daily in clinical trials was comparable to that for placebotreated patients. In studies in which more than 1,300 AR patients were treated with either 60 mg fexofenadine twice-daily or placebo, the most commonly reported adverse events were viral infection (2.5% for fexofenadine-treated patients versus 1.5% for placebo-treated patients), nausea (1.6% versus 1.5%, respectively), drowsiness (1.3% versus 0.9%, respectively), and fatigue (1.3% versus 0.9%, respectively). The product label also reports the incidence of adverse events among nearly 600 AR patients aged 12 years or older treated with either 180 mg fexofenadine once daily or placebo in clinical trials, and the adverse events most commonly reported in these studies were headache (10.6% of patients treated with fexofenadine versus 7.5% of patients treated with placebo), upper respiratory tract infection (3.2% versus 3.1%, respectively), and back pain (2.8% versus 1.4%, respectively). The adverse events most commonly reported among nearly 450 pediatric AR patients (aged 6–11 years) treated with 30 mg fexofenadine twice daily or placebo were headache (7.2% versus 6.6%, respectively), upper respiratory tract infection (4.3% versus 1.7%, respectively), coughing (3.8% versus 1.3%, respectively), and accidental injury (2.9% versus 1.3%, respectively). The original U.S. patent covering fexofenadine expired in 2001. However, SanoÞ-Aventis holds numerous other patents pertaining to the drug, including method of use, formulation, composition, and process patents; the last of these patents expires in 2018. In 2001, Barr Laboratories initiated legal action challenging Aventis’s (now SanoÞ-Aventis’s) remaining patents for fexofenadine and Þled an abbreviated new drug application (ANDA) seeking FDA approval for a generic version of fexofenadine. Aventis subsequently sued Barr Laboratories for patent infringement, prompting a Hatch-Waxman 30-month stay on the launch of any generics. Since then, additional companies have Þled ANDAs with the FDA for generic formulations of either fexofenadine or the fexofenadine/pseudoephedrine combination product, and Aventis has Þled a number of patent infringement lawsuits against those companies, which include Impax Laboratories, Teva Pharmaceuticals, Mylan Laboratories, and Dr. Reddy’s Laboratories. On June 30, 2004, a summary judgment was handed down by the United States District Court in New Jersey in the patent infringement lawsuit brought about by Aventis against these Þve generic drug manufacturers (Barr Laboratories, Impax Laboratories, Teva Pharmaceuticals, Mylan Pharmaceuticals, and Dr. Reddy’s Laboratories). The judgment stated that the defendants’ products do not infringe upon three of SanoÞ-Aventis’s fexofenadine patents. Five additional patents included in the original lawsuit remain in litigation, and, in March 2004, Aventis and Albany Molecular Research (AMR) Þled lawsuits against the Þve aforementioned generic drug companies pertaining to two additional patents owned by AMR and licensed to SanoÞ-Aventis. Patents covering fexofenadine and the fexofenadine/pseudoephedrine combination product are not set to expire in Europe and Japan until 2014. Cetirizine. Cetirizine (PÞzer/UCB/Daiichi/Sumitomo’s Zyrtec/Virlix/Cirrus, generics) (Figure 6) is an oral second-generation antihistamine that has been marketed by UCB Pharmaceuticals in Europe since the late 1980s. The drug is

CURRENT THERAPIES

OH

OH

O N

O O

N

N

Cl

389

O

N

Cl

FIGURE 6. Structure of R- and S-cetirizine.

available over the counter in all Þve European countries under study (AESGP, 2004[a]). The drug received FDA approval in the United States in 1995, where it is jointly marketed by UCB and licensee PÞzer. UCB received regulatory approval in Japan for cetirizine in 1998; currently, Daiichi and Sumitomo distribute the drug in this region. However, in April 2005, UCB announced that the distribution agreement with Sumitomo will be terminated as of June 30, 2005, and that UCB will begin co-promoting Þve and 10 mg tablets of cetirizine in Japan with GlaxoSmithKline K.K. (GlaxoSmithKline Japan) beginning on July 1, 2005. Like fexofenadine, cetirizine is indicated for treating SAR and chronic idiopathic urticaria (CIU). However, unlike fexofenadine, cetirizine is also indicated for treating PAR. Cetirizine is, unlike fexofenadine, approved for use in younger patients; cetirizine is indicated for SAR in patients as young as two years of age, and PAR and CIU in patients as young as six months in age. Cetirizine is also indicated for eczema and dermatitis in Japan. Cetirizine is available in oral tablet, chewable tablet, and oral syrup formulations. This availability represents another advantage over fexofenadine, which is available only as oral capsules or tablets; chewable tablet and oral syrup formulations can be administered to young children who would otherwise have a difÞcult time swallowing oral tablets or capsules. In 2001, UCB and PÞzer received FDA approval for Zyrtec-D, a 12-hour Þxed combination of cetirizine and the decongestant pseudoephedrine, for the treatment of SAR and PAR in patients aged 12 or older. Cetirizine is the carboxylated metabolite of the Þrst-generation antihistamine hydroxyzine (PÞzer’s Atarax/Visaril, generics). Cetirizine’s predominant therapeutic effects are exerted through the same mechanism of action as other second-generation antihistamines, as discussed in the “Mechanism of Action” section for the drug class. Like fexofenadine, cetirizine’s efÞcacy in relieving AR symptoms has been demonstrated in numerous placebo-controlled studies. In one such double-blind study involving 419 adult SAR participants, patients were randomized to receive either placebo or one of three cetirizine doses (5, 10, or 20 mg once daily) for one week (Curran MP, 2004; Falliers CJ, 1991). EfÞcacy was measured by

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improvement over baseline in the patient-assessed TSS at the end of the sevenday treatment period, as well as the physician’s global assessment of treatment efÞcacy. (TSS, in this case, was the sum of individual severity scores for the following seven symptoms: sneezing, rhinorrhea, itchy eyes, itchy nose, itchy palate, eye redness, and watery eyes.) Treatment with 20 mg cetirizine daily was associated with the largest reduction in TSS over baseline (approximately 62%), compared with improvements of approximately 52%, 39%, and 26% for patients treated with 10 mg cetirizine, 5 mg cetirizine, and placebo, respectively. Additionally, signiÞcantly larger proportions of patients were deemed effectively treated (according to physicians’ global assessment of treatment efÞcacy) with cetirizine than with placebo. Overall, 73%, 79%, and 77% of patients treated with 5 mg, 10 mg, and 20 mg cetirizine daily, respectively, were deemed effectively treated, compared with only 53% of patients treated with placebo. Cetirizine treatment was shown to improve both patient- and physician-rated nasal congestion scores (which were also assessed in this study), but the difference in improvement from that of the placebo group was not signiÞcantly different. The prominent side effects in this study were somnolence (9% of patients treated with 5 mg cetirizine, 23% of patients treated with 10 mg cetirizine, 25% of patients treated with 20 mg cetirizine, and 6% of patients treated with placebo), headache (10%, 11%, 15%, and 15%, respectively), dry mouth (5%, 7%, 6%, and 0%, respectively), and fatigue (3%, 7%, 5%, and 4%). Although cetirizine is a second-generation antihistamine and is associated with less fatigue, drowsiness, and performance impairment than Þrst-generation antihistamines, the drug is not classiÞed as a “nonsedating” antihistamine, unlike fexofenadine. According to cetirizine’s product label, the incidence of somnolence in clinical studies was higher among patients treated with the drug compared with patients treated with placebo, and this trend was dose related. In controlled and uncontrolled clinical studies involving more than 6,000 adult patients (aged 12 years or older), 14% of patients treated with 10 mg cetirizine daily experienced somnolence, compared with 11% of patients treated with 5 mg cetirizine daily and 6% of patients treated with placebo. A similar dose-related trend was noted in studies involving pediatric patients (ages 6–11) treated with cetirizine in clinical studies, but the incidence of somnolence was not as high as in studies involving adults. According to the product label for cetirizine, the incidence of somnolence was 4.2%, 1.5%, and 1.3% for pediatric patients treated with 10 mg cetirizine, 5 mg cetirizine, and placebo daily, respectively. Because of the higher incidence of somnolence among cetirizine-treated patients, the drug’s product label includes a precaution stating that patients taking cetirizine should use caution when driving a car or operating machinery, and that concurrent use of the drug with alcohol or other agents that depress the CNS should be avoided to prevent additional impairments of alertness and performance. As described in the aforementioned studies conducted by F. Hampel, P.H. Howarth, and their respective colleagues, cetirizine demonstrated efÞcacy comparable to that of fexofenadine in large-scale clinical trials involving SAR patients (Hampel F, 2003; Howarth PH, 1999). However, in each of these studies, the

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incidence of drowsiness was signiÞcantly greater in patients treated with cetirizine compared with that in patients treated with fexofenadine. Cetirizine has also demonstrated comparable efÞcacy in improving AR symptoms as other second-generation antihistamines, including ebastine (Almirall/Dainippon/Meiji Seika’s Ebastel), mizolastine (SanoÞ-Aventis’ Mizollen, Schwarz Pharma’s Mizollen/Zolim), astemizole (Janssen Pharmaceutica [a Johnson & Johnson subsidiary] Hismanal, which has since been withdrawn from all markets under study), and terfenadine (Curran MP, 2004). Generic versions of cetirizine are available in all European countries under study, excluding Italy (where the drug is patent protected until 2009). Cetirizine faces patent expiry in the United States and Japan in 2007. In the United States, the FDA has already granted tentative approval for several companies’ ANDAs for generic versions of cetirizine. Desloratadine. In the face of the 2002 patent expiry for its blockbuster second-generation antihistamine loratadine, Schering-Plough launched desloratadine (Clarinex/Aerius/Neoclarityn), the principal active metabolite of loratadine, as a follow-on product. Desloratadine received its Þrst approval in January 2001 when the European Commission granted marketing authorization to all 15 European Union member states for an oral tablet formulation for the treatment of SAR in adults and children 12 years or older. Since this initial authorization, desloratadine’s European label has been expanded to include the indications of PAR and CIU. A rapidly disintegrating pill formulation and an oral syrup formulation have been approved as well, the latter of which is indicated for children as young as age two. Following a long delay due to manufacturing problems, Schering-Plough launched desloratadine in the United States in January 2002, a year after it received an approvable letter from the FDA for a tablet formulation for the treatment of SAR. The delayed U.S. approval hampered Schering-Plough’s efforts to convert as many patients taking loratadine to desloratadine as possible before the former lost patent protection and was subsequently switched from prescription to OTC status in the United States in December 2002. (Loratadine has been available OTC in Germany and the United Kingdom since the early 1990s, and was switched to OTC status in Italy in 2004. Desloratadine received FDA marketing clearance for the broadened indication of AR (encompassing SAR and PAR) and CIU in February 2002. In June of the same year, the FDA granted Schering-Plough approval to market a rapidly disintegrating tablet formulation of desloratadine for SAR, PAR, and CIU in patients aged 12 or older. In September 2004, the FDA approved an oral syrup formulation of desloratadine for treating SAR in patients as young as 2 years and for treating PAR and CIU in patients aged 6 months or older. The FDA granted approval to Schering-Plough’s Clarinex-D 24 Hour, a Þxed combination of desloratadine and the decongestant pseudoephedrine, for SAR in patients aged 12 or older in March 2005. Desloratadine was originally developed by Sepracor, which in 1997 licensed worldwide rights to the drug’s patents to Schering-Plough. As a result of this

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agreement, Sepracor now earns royalties on desloratadine sales from ScheringPlough. The drug is currently not marketed in Japan. Like other second-generation antihistamines proÞled in this section, desloratadine’s predominant therapeutic effects are exerted through the same means as discussed in the “Mechanism of Action” section for the drug class. As is the case with fexofenadine and cetirizine, desloratadine’s beneÞcial effects in treating AR symptoms have been demonstrated in numerous clinical studies, including two similarly designed, double-blind, placebo-controlled trials. In both studies, patients were randomized to receive either 5 mg desloratadine once daily or placebo for two weeks, and the mean change from baseline in the average of morning and evening reßective TSS served as the primary endpoint. (TSS, in this case, was the sum of total nasal symptom score [TNSS; sum of scores for rhinorrhea, nasal congestion/stufÞness, nasal itching, and sneezing] and total non-nasal symptom score [TNNSS; sum of scores for itching/burning eyes, tearing/watering eyes, redness of eyes, and itching of ears or palate].) In one of the studies involving 328 SAR patients, desloratadine-treated patients demonstrated a mean 30.0% improvement over baseline TSS, compared with an improvement of 22.2% for placebo-treated patients. This difference was statistically signiÞcant, as was the difference between groups in TNSS improvements (31.5% versus 22.8%, respectively) and TNNSS improvements (35.9% and 26.6%, respectively) (Meltzer EO, 2001[a]; Murdoch D, 2003). Similar Þndings were found in the second 346-patient study; TSS improvements were 30.3% and 18.2% for desloratadine- and placebo-treated patients, respectively. Corresponding improvements for TNSS were 24.4% and 11.7%, respectively, and were 26.6% and 11.5%, respectively, for TNNSS. The improvements in the desloratadine groups over the placebo group were statistically signiÞcant for all measures—TSS, TNSS, and TNNSS. Desloratadine was the Þrst second-generation antihistamine to demonstrate proven, consistent efÞcacy in relieving nasal congestion (Bachert C, 2001). The drug acts as a subtype-selective antagonist of muscarinic receptors (at M1 and M3 receptor subtypes), and this antagonism may contribute to its decongestant effects. Desloratadine’s beneÞcial effect on nasal congestion was demonstrated in three large-scale, placebo-controlled studies involving 278 to 346 patients. In each of the three studies, which ranged from two to four weeks in duration, patients treated with desloratadine demonstrated a mean 21.3–23.5% improvement over baseline on the average morning and evening reßective congestion score, compared with improvements of 13.6–16.2% for patients treated with placebo. In each of the studies, the difference between the desloratadine and placebo groups was statistically signiÞcant (Geha RS, 2001). Desloratadine is well tolerated among AR patients. The product label for the drug reports compiled safety data for 3,300 AR patients aged 12 or older treated with either the recommended dosage of desloratadine (5 mg once daily) or placebo during clinical studies. The label also reports that the following adverse events occurred in more than 2% of desloratadine-treated patients and occurred more frequently among patients treated with the drug than among patients treated

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with placebo: pharyngitis (4.1% of desloratadine recipients vs. 2.0% of placebo recipients), dry mouth (3.0% versus 1.9%, respectively), myalgia (2.1% versus 1.8%, respectively), fatigue (2.1% versus 1.2%, respectively), somnolence (2.1% versus 1.8%, respectively), and dysmenorrhea (2.1% versus 1.6%, respectively). The product label for desloratadine reports consolidated adverse event rates for desloratadine-treated pediatric patients with either AR or CIU; the adverse event rates for pediatric AR sufferers are not reported separately from those for pediatric CIU patients. In general, the incidence of adverse events was lower among older children (ages 6–11 years) compared with younger children (ages 2–5) and infants (ages 6–23 months). A postmarketing surveillance study of desloratadine-treated patients found a lower incidence of adverse events than that reported on the product’s label. Among nearly 48,000 SAR patients treated with desloratadine for 1–232 days (mean 38.4 days), the overall incidence of adverse events was 0.44%. The most commonly reported adverse events deemed possibly or probably related to the drug were fatigue, headache, nausea, dry mouth, and diarrhea, each of which occurred in ≤0.07% of patients (Murdoch D, 2003). Desloratadine has market exclusivity in the United States until 2007 and is patent-protected in all European countries under study through 2010. Although desloratadine is currently not marketed in Japan, the drug’s patent in this region expires in 2005. It will likely be granted six years of data exclusivity when it launches in Japan (similar to when loratadine launched). Levocetirizine. Levocetirizine (UCB’s Xyzal/Xusal), the (R) enantiomer of cetirizine, was originally developed by Sepracor. In May 1999, Sepracor licensed all issued and pending patent rights pertaining to levocetirizine to UCB, excluding patent rights in the United States and Japan, where Sepracor retained its rights to the drug. In January 2001, UCB received marketing authorization in Germany for levocetirizine for the treatment of SAR, PAR, and CIU in patients as young as age six. The drug was subsequently approved by the other European Union (EU) member states in September of the same year. By the end of 2001, levocetirizine had launched in Germany and the United Kingdom; the drug did not launch in the other EU countries under study (France, Italy, and Spain) until 2003. In December 2004, levocetirizine’s European label was expanded further when it became the Þrst antihistamine to be approved for treating persistent allergic rhinitis (PER). The indication of PER reßects the new classiÞcation of AR put forth in the Allergic Rhinitis and Its Impact on Asthma (ARIA) guidelines. (The ARIA guidelines are discussed in greater detail in “Etiology and Pathophysiology.”) As stated previously, levocetirizine is the (R) enantiomer of cetirizine. Studies have demonstrated that levocetirizine accounts for cetirizine’s afÞnity to the H1 receptor rather than the (S) enantiomer, dextrocetirizine (Devalia JL, 2001; Wang DY, 2001). According to the product label for levocetirizine, the drug has a twofold higher afÞnity for the H1 receptor compared to that of its predecessor compound, cetirizine. Like other second-generation antihistamines proÞled in this section, levocetirizine’s predominant therapeutic effects are exerted through the same means as discussed in the “Mechanism of Action” section for the drug class.

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Levocetirizine’s approval for PER was based on the Xyzal in Persistent Rhinitis Trial (XPERT), a six-month, double-blind, placebo-controlled study involving 551 patients. Study participants were randomized to receive either 5 mg levocetirizine daily or placebo, and efÞcacy was measured by two primary end points: mean improvement over baseline Total 5 Symptom Score (T5SS, the sum of individual scores for rhinorrhea, sneezing, nasal congestion, itchy nose, and itchy eyes) and change from baseline in the overall score on the Rhinoconjunctivitis Quality of Life Questionnaire (RQLQ, a disease-speciÞc measure of health-related quality of life). After four weeks of treatment (the predeÞned timing for both primary endpoint assessments), levocetirizine-treated patients demonstrated a 39% improvement over baseline T5SS and a 49% improvement over baseline RQLQ scores; corresponding improvements for placebo-treated patients were 27% and 33%, respectively. Additionally, patients treated with levocetirizine demonstrated signiÞcantly greater improvements over patients treated with placebo on individual symptom scores for sneezing, rhinorrhea, itchy nose, and itchy eyes as early as one week into the trial, and these signiÞcant differences were evident after six months of treatment. The difference between the levocetirizine and placebo groups on the assessment of nasal congestion did not reach statistical signiÞcance until 12 weeks into the trial. During the course of the entire six-month trial, 69.1% of levocetirizine-treated patients reported at least one adverse event, compared with 70.7% of patients in the placebo group. The most commonly reported adverse events among study participants were headache (24.5% of patients treated with levocetirizine versus 23.2% of patients treated with placebo), pharyngitis (19.8% versus 20.5%, respectively), inßuenza-like symptoms (14.0% versus 13.9%, respectively), fatigue (8.6% versus 7.0%, respectively), somnolence (6.8% versus 1.8%, respectively), and gastroenteritis (2.9% versus 5.1%, respectively) (Bachert C, 2004). Levocetirizine’s beneÞcial effect in treating AR symptoms was also demonstrated in a double-blind, placebo-controlled study involving 294 PAR patients. Study participants were treated with either 5 mg levocetirizine or placebo for six weeks, and efÞcacy was measured as the mean improvement over baseline for the Total 4 Symptom Score (T4SS, the sum of scores for itchy nose, itchy eyes, rhinorrhea, and sneezing); improvements over baseline for the four individual symptoms, as well as for nasal congestion, were also reported. After six weeks of treatment, patients treated with levocetirizine demonstrated a 48.1% improvement over baseline T4SS, compared with an improvement of 32.6% for placebo-treated patients. Additionally, patients treated with levocetirizine demonstrated signiÞcantly greater improvements over placebo-treated patients on all individually assessed symptoms, including nasal congestion. The improvements in nasal congestion scores were 22.3% and 8.8% for levocetirizine- and placebo-treated patients, respectively. The overall incidence of adverse events for levocetirizine recipients (60.0%) was lower than that for placebo recipients (68.1%) in this study. Headache (34.7% for levocetirizine-treated patients versus 34.7% for placebotreated patients), inßuenza-like symptoms (16.7% versus 13.9%, respectively),

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pharyngitis (8.7% versus 4.2%, respectively), and upper respiratory tract infections (6.7% versus 9.0%, respectively) were the most commonly reported adverse events (Potter PC, 2003). According to the product label for levocetirizine, which reports the pooled safety data from more than 1,700 patients treated with either the drug or placebo in clinical studies, the overall incidence of adverse events associated with levocetirizine use appears to be lower than that reported in the aforementioned studies conducted by C. Bachert, P.C. Potter, and their respective colleagues. According to the drug’s product label, 15.1% of patients treated with 5 mg levocetirizine in clinical studies experienced at least one adverse event, compared with 11.3% of patients treated with placebo; nearly 92% of these adverse events were mild to moderate. The most commonly reported adverse events were similar to those in the study conducted by Bachert and colleagues: headache (2.6% for levocetirizine-treated patients versus 3.2% of placebo-treated patients), somnolence (5.2% versus 1.4%, respectively), dry mouth (2.6% versus 1.6%, respectively), and fatigue (2.5% versus 1.2%, respectively). Like its predecessor compound, cetirizine, the overall incidence of sedating adverse events (i.e., somnolence, fatigue, and asthenia [loss of energy or strength]) associated with levocetirizine use (8.1%, as reported by the product label) is higher than that for placebo-treated patients (3.1%). The product label for levocetirizine acknowledges that although no evidence has been revealed in comparative clinical trials to indicate the drug negatively affects mental alertness, reactivity, or ability to drive, patients should consider their potential response to levocetirizine if they intend to drive, operate machinery, or participate in hazardous activities, as sedating effects have been noted in clinical trials involving the drug. Although levocetirizine is currently not on the market in the United States, the U.S. patent for this drug is not expected to expire until 2012. Levocetirizine will not face patent expiration in Europe during the study period. Intranasal Corticosteroids Overview. Intranasal corticosteroids are regarded as the most effective drugs available for the treatment of AR because of their potent and extensive antiinßammatory effects. However, physicians often prescribe oral second-generation antihistamines as Þrst-line therapy because patients generally prefer oral medications to drugs that are administered intranasally. Although they are not always as effective as oral antihistamines at alleviating ocular irritation, nasal corticosteroids provide excellent relief from other early- and late-phase symptoms, including sneezing, itchy or runny nose, and nasal congestion. As a result, agents in this class are recommended for Þrst-line use in AR patients with frequent or persistent symptoms or patients suffering from nasal congestion (Bousquet J, 2001). Intranasal corticosteroids are most effective when administered prophylactically on a daily basis. Patients may begin to experience symptomatic relief within approximately 12 hours, but several days of administration may be required

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before the maximal beneÞt is achieved. This onset of action is much longer than that of oral second-generation antihistamines, which typically ranges from 30 minutes to three hours. Several intranasal corticosteroid products are marketed for the treatment of AR: ßuticasone propionate (GlaxoSmithKline’s Flonase/Flixonase, generics), mometasone furoate (Schering-Plough’s Nasonex), budesonide (AstraZeneca’s Rhinocort/Rhinocort AQ), triamcinolone acetonide (Aventis’ Nasacort/Nasacort AQ), beclomethasone (GlaxoSmithKline’s Beconase/Beconase AQ, generics), and ßunisolide (IVAX’s Nasarel). Clinical studies have demonstrated that available intranasal corticosteroid products offer comparable clinical efÞcacy in relieving AR symptoms (Nielsen LP, 2001; Waddell AN, 2003), but agents in this class differ with respect to other product attributes, such as systemic bioavailability, dosing frequency, fragrance, and use of preservatives. Therefore, the choice of intranasal corticosteroid is often based on a balance of safety, convenience, and patient preference rather than clinical efÞcacy. As stated previously, intranasal corticosteroids are prescribed less often than second-generation antihistamines, largely because of patient preference for orally administered drugs. In addition, although clinical trials have shown that intranasal corticosteroids are very safe—even when taken over the long term—the stigma associated with steroid use leads many patients to avoid using intranasal corticosteroids unless absolutely necessary. Indeed, long-term use of oral, systemic steroids is associated with many well-known and serious side effects (e.g., adrenal insufÞciency, osteoporosis, growth retardation), and most orally inhaled steroids have been associated with suppression of the hypothalamic-pituitaryadrenal (HPA) axis because these drugs can pass into the systemic circulation and mimic the effects of naturally occurring corticosteroids. However, because of the localized activity in the nose, lower corticosteroid doses are required to achieve a therapeutic effect when the drug is delivered intranasally. Additionally, intranasal corticosteroids are associated with fewer side effects than other corticosteroid formulations because of their lower systemic absorption from the nasal mucosa and/or extensive Þrst-pass metabolism. As a result of systemic safety concerns associated with corticosteroid products, systemic bioavailability is probably the most important characteristic distinguishing one intranasal corticosteroid from another. The systemic bioavailability of currently available intranasal corticosteroids varies greatly; systemic bioavailabilities for intranasal formulations of triamcinolone acetonide, beclomethasone, ßunisolide, and budesonide are reported to be 46%, 44%, 40–50%, and 31%, respectively, while intranasal formulations of two newer corticosteroids—ßuticasone propionate and mometasone furoate—have signiÞcantly reduced systemic bioavailabilities of 0.42–0.51% and 0.46%, respectively (Salib RJ, 2003[b]). Because of their superior systemic bioavailabilities and convenient once-daily recommended dosing schedule, this section proÞles intranasal formulations of ßuticasone propionate and mometasone furoate as representatives for other agents in this class; these two agents are also the top-selling intranasal corticosteroids for the treatment of AR in the major markets under study.

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Mechanism of Action. All corticosteroids exert their therapeutic effect by binding to intracellular glucocorticoid receptors, which are found in all cells. Following translocation to the nucleus, activated glucocorticoid receptors either promote or inhibit transcription of genes that are involved in the inßammatory process. In this way, corticosteroids inßuence the release of cytokines and proinßammatory mediators, including the interleukins (IL)-1, -4, -5, -6, -8, and -13; tumor necrosis factor-alpha (TNF-α); granulocyte-macrophage colonystimulating factor (GM-CSF); and regulated upon activation, normal T cell expressed and secreted (RANTES). By lowering the levels of these inßammation mediators, corticosteroid therapy downregulates the expression of adhesion molecules and chemokines and thus reduces the recruitment and accumulation of inßammatory cells—including eosinophils, mast cells, basophils, and antigen-presenting cells (APCs)—to the nasal mucosa. By reducing the inßux of inßammatory cells into the nasal mucosa, intranasal corticosteroids effectively reduce nasal hyperresponsiveness to allergic stimuli (Mygind N, 2001; Salib RJ, 2003[b]). Other key mechanisms by which corticosteroids help alleviate the symptoms of AR include promotion of apoptosis of eosinophils, inhibition of mediator release from basophils and eosinophils, inhibition of T-helper 2 (TH 2) cell formation, inhibition of immunoglobulin E (IgE) antibodies, and reduction in mucus secretion by submucosal gland cells (Mygind N, 2001). Intranasal Fluticasone Propionate. The intranasal formulation of ßuticasone propionate (GlaxoSmithKline’s Flonase/Flixonase, generics) (Figure 7) launched in Europe in 1991. The drug was subsequently approved for use in Japan and the United States in July and October of 1994, respectively. Intranasal ßuticasone propionate is indicated for the treatment of both SAR and PAR in patients aged 4 years or older; the drug is also indicated for the management of nasal symptoms associated with nonallergic rhinitis. Intranasal ßuticasone propionate is formulated as an aqueous solution that is administered via a metering, atomizing spray pump. A nasal drop formulation of

F S H3C

HO H3C

CH3

O O CH3

H F

O

H

O F FIGURE 7. Structure of fluticasone propionate.

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ßuticasone propionate is also available in Europe for treating AR, but is not as widely used as is the aqueous solution. Like many other intranasal corticosteroids, intranasal ßuticasone propionate contains the preservative benzalkonium chloride, which can cause allergic or irritant reactions in some patients. The recommended initial dosing for adult patients is two sprays in each nostril once daily, and the dose may be reduced to one spray per nostril once daily after a few days if a patient’s symptoms continue to respond to a lower dose. The recommended initial dose for children and adolescents is one spray in each nostril once daily; two sprays per nostril once daily can be administered if a child fails to achieve adequate symptom control with the lower dose. Like other intranasal corticosteroids, intranasal ßuticasone propionate is a potent anti-inßammatory agent. The anti-inßammatory effects of these drugs are numerous (as described in the previous section). Through these multiple antiinßammatory effects, intranasal ßuticasone propionate effectively reduces nasal hyperresponsiveness to allergic stimuli. The efÞcacy of intranasal ßuticasone has been demonstrated in multiple clinical studies. In one trial involving 227 SAR patients, study participants were treated with either 200 µg intranasal ßuticasone propionate once-daily, 100 µg intranasal ßuticasone propionate twice-daily, or placebo (Nathan RA, 1991). At the end of the two-week treatment period, patients treated with 200 µg intranasal ßuticasone propionate once daily demonstrated a 46.8% improvement over baseline in physician-rated mean TNSS, compared with an improvement of 26.8% for placebo-treated patients. A greater improvement was seen in patients treated with 100 µg intranasal ßuticasone propionate twice-daily (54.9% improvement). Treatment with intranasal ßuticasone propionate—regardless of dose—was also associated with signiÞcantly greater improvements over baseline scores for each of the four individual nasal symptom scores. Patients treated with 200 µg intranasal ßuticasone propionate once daily demonstrated improvements of 33.8%, 46.0%, 56.4%, and 53.8% for physician-rated nasal obstruction, rhinorrhea, sneezing, and nasal itching, respectively. Corresponding improvements for patients treated with 100 µg intranasal ßuticasone propionate once daily were 40.8%, 53.7%, 67.9%, and 61.3%, respectively, and were 17.4%, 26.9%, 30.2%, and 34.4%, respectively, for placebo-treated patients. The efÞcacy of intranasal ßuticasone propionate has also been demonstrated in PAR patients. In one 365-patient, six-month study, patients were treated with either 200 µg intranasal ßuticasone propionate once daily, 100 µg intranasal ßuticasone propionate twice daily, or placebo (Banov CH, 1994). As in the aforementioned study conducted by R.A. Nathan and colleagues, efÞcacy was primarily measured by the improvement over baseline in clinician-rated mean TNSS (sum of scores for nasal congestion, rhinorrhea, sneezing, and itching). At the end of 24 weeks, the patients treated with 200 µg intranasal ßuticasone propionate once daily, 100 µg intranasal ßuticasone propionate twice daily, and placebo demonstrated improvements of 47%, 51%, and 33% in clinician-rated mean TNSS. Additionally, treatment with intranasal ßuticasone propionate was superior to placebo on a second primary efÞcacy end point: clinician-rated overall

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evaluation of treatment efÞcacy. At the conclusion of the study, 59% of patients treated with 200 µg intranasal ßuticasone propionate once daily and 67% of patients treated with 100 µg intranasal ßuticasone propionate twice-daily demonstrated moderate or signiÞcant improvements according to physicians, compared with only 41% of placebo-treated patients. Intranasal ßuticasone propionate has demonstrated superior efÞcacy compared with a number of oral second-generation antihistamines, including cetirizine, loratadine, terfenadine, and astemizole (Wiseman LR, 1997). In one large-scale, two-week comparative trial, 600 SAR patients were randomized to one of four treatment groups: 200 µg intranasal ßuticasone propionate once daily, 200 µg intranasal ßuticasone propionate and 10 mg loratadine administered together once daily, 10 mg loratadine once daily, or placebo. At the end of the 14-day treatment period, improvements over baseline scores for clinician-rated TNSS were 61.3%, 61.0%, 32.6%, and 33.7% for patients treated with intranasal ßuticasone propionate monotherapy, intranasal ßuticasone propionate and loratadine combination, loratadine monotherapy, and placebo, respectively. These results were statistically signiÞcant for the two groups receiving intranasal ßuticasone propionate. These two groups also demonstrated signiÞcantly greater improvements over patients treated with loratadine monotherapy and placebo on each of the individual nasal symptom scores. Patients treated with intranasal ßuticasone propionate monotherapy demonstrated improvements of 54.5%, 55.9%, 67.2%, and 69.4% on the clinician-rated measures of nasal blockage, discharge, itching, and sneezing, respectively, while the corresponding improvements for the intranasal ßuticasone propionate and loratadine combination group were 52.9%, 59.8%, 65.5%, and 67.4%, respectively. The improvements for the loratadine monotherapy group on the four aforementioned individual nasal symptoms (24.9%, 31.6%, 38.4%, and 36.9%, respectively) were comparable to the improvements demonstrated by placebo-treated patients (26.0%, 33.3%, 37.4%, and 39.1%, respectively) (Ratner PH, 1998). In this comparative study, patients treated with intranasal ßuticasone propionate (either as monotherapy or in combination with loratadine) also demonstrated signiÞcantly greater improvements on the RQLQ than patients treated with loratadine monotherapy and placebo recipients. Global RQLQ scores improved 53.6% and 57.5% over baseline for the intranasal ßuticasone propionate monotherapy and intranasal ßuticasone propionate and loratadine combination groups, respectively, while corresponding improvements for the loratadine monotherapy and placebo groups were 31.7% and 32.5%, respectively (Ratner PH, 1998). The improvements on each of the seven individual RQLQ domains (nasal symptoms, ocular symptoms, activities, practical problems, sleep, emotional issues, and other non-nasal, non-ocular symptoms) were also signiÞcantly greater for the two intranasal ßuticasone propionate groups compared with the loratadine monotherapy and placebo groups. According to the product label for intranasal ßuticasone propionate, the most common adverse events that occurred in more than 1,500 patients treated with either the drug or placebo in multiple clinical trials were headache (16.1% of

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patients treated with 200 µg intranasal ßuticasone propionate once daily versus 14.6% of patients treated with placebo), pharyngitis (7.8% versus 7.2%, respectively), epistaxis (bloody nose, 6.9% versus 5.4%, respectively), and nasal burning/irritation (3.2% versus 2.6%, respectively). The product label for intranasal ßuticasone propionate also reports the incidence of adverse events among 167 children (ages 4–11 years) treated with 100 µg intranasal ßuticasone propionate once daily. The most commonly reported adverse events among these pediatric patients were asthma symptoms (7.2%), headache (6.6%), pharyngitis (6.0%), and epistaxis (6.0%). In March 2003, IVAX Þled an ANDA with the FDA seeking approval for a generic version of intranasal ßuticasone propionate. However, following expiry of intranasal ßuticasone propionate’s patent in May 2004, GlaxoSmithKline Þled a citizen’s petition in June 2004 asking the FDA to delay the approval of any generic versions of intranasal ßuticasone propionate until the FDA issues Þnal guidelines on establishing bioavailability and bioequivalency for products that act locally rather than systemically, such as inhaler products. The FDA issued draft guidance on bioavailability and bioequivalence studies for nasal aerosols and nasal sprays for local action in April 2003.. European and Japanese patents pertaining to intranasal ßuticasone propionate expire between 2005 and 2006. In May 2005, IVAX became the Þrst company to receive marketing authorization for a generic version of intranasal ßuticasone propionate in the United Kingdom, and the company plans to seek additional approvals for this generic equivalent in additional European countries. Intranasal Mometasone Furoate. Intranasal mometasone furoate (ScheringPlough’s Nasonex) (Figure 8) received European and U.S. regulatory approval in 1997. The drug is currently not available in Japan, but Phase III trials are under way in this region. Intranasal mometasone furoate is indicated for the treatment of SAR and PAR; it is the only marketed intranasal corticosteroid indicated for use in children as young as age two. Additionally, the drug is the only currently available intranasal corticosteroid that is indicated for the prophylaxis of nasal symptoms in SAR patients aged 12 years or older; it is recommended that patients begin taking the drug two to four weeks before the start of an allergy season to prevent the development of nasal symptoms. Intranasal mometasone furoate is also indicated for the treatment of nasal polyps, a complication of AR, in patients aged 18 years or older. The FDA approved a new scent-free formulation of intranasal mometasone in August 2004. Like intranasal ßuticasone propionate, intranasal mometasone furoate is formulated as an aqueous solution that is delivered via a metered-dose, manual pump spray. Intranasal mometasone furoate also contains the preservative benzalkonium chloride, which can cause allergic or irritant reactions in some patients. Similar to intranasal ßuticasone propionate, the recommended initial dosing for intranasal mometasone furoate for adult patients is two sprays in each nostril once daily, and the recommended initial dose for children aged two to 11 is one spray in each nostril once daily.

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FIGURE 8. Structure of mometasone furoate.

Intranasal mometasone furoate exerts its potent anti-inßammatory effects through the same mechanisms of action as other intranasal corticosteroids, as discussed in the “Mechanism of Action” section for the drug class. The efÞcacy of intranasal mometasone furoate has been demonstrated in numerous clinical trials. In one four-week study, the efÞcacy of intranasal mometasone furoate was compared with that of intranasal beclomethasone and placebo in 501 SAR patients (Hebert JR, 1996). Study participants were randomized to receive one of four treatments: either 100 µg or 200 µg intranasal mometasone furoate once daily, 200 µg intranasal beclomethasone twice daily, or placebo. Physician-rated improvement over baseline for TNSS on day 8—the predeÞned primary efÞcacy assessment—were 53% for the 100 µg intranasal mometasone furoate group, 59% for the 200 µg intranasal mometasone furoate group, 59% for the intranasal beclomethasone group, and 34% for the placebo group. The three active treatment groups also demonstrated signiÞcantly greater improvements in the four individual nasal symptom scores on day 8 compared with placebo recipients. The three groups treated with intranasal corticosteroids demonstrated mean improvements of 51–58% for rhinorrhea, 41–52% for nasal stufÞness/congestion, 56–59% for nasal itching, and 63–71% for sneezing; corresponding improvements for the placebo group were 26%, 28%, 31%, and 32%, respectively. Additionally, the percentage of patients who were deemed by physicians to have no or mild symptoms on day 8 was 66%, 71%, 75%, and 43% for patients treated with 100 µg intranasal mometasone furoate, 200 µg intranasal mometasone furoate once, intranasal beclomethasone, and placebo, respectively. The incidence of adverse events was comparable between the four treatment groups (from a low of 25% for patients treated with 100 µg intranasal mometasone furoate to a high of 30% for patients treated with intranasal beclomethasone). Headache, nasal burning, and epistaxis were the three most commonly reported adverse events in this study. As stated previously, intranasal mometasone furoate is indicated for the prevention of nasal symptoms in SAR patients. This beneÞcial effect was demonstrated in one eight-week study involving 349 SAR patients who were randomized

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to one of three treatment groups: 200 µg intranasal mometasone furoate once daily, 168 µg intranasal beclomethasone twice daily, or placebo (Graft D, 1996). Treatment was initiated four weeks before the estimated start of the ragweed allergy season, and the average proportion of minimal symptom days served as the primary end point. (A minimal symptom day was deÞned as a day on which morning and evening averaged TNSS was less than or equal to 2 out of a maximum score of 12.) During the ragweed season, the proportion of minimal symptom days was 83% for patients treated with intranasal mometasone furoate, 77% for patients treated intranasal beclomethasone, and 64% for patients treated with placebo. Intranasal mometasone furoate and intranasal ßuticasone propionate demonstrated comparable efÞcacy on multiple end points in a large-scale, placebo-controlled comparative trial. A total of 550 PAR patients were randomized to receive either 200 µg intranasal mometasone furoate once daily, 200 µg intranasal ßuticasone propionate once daily, or placebo for a period of three months (Mandl M, 1997). The three treatment groups demonstrated improvements of 37%, 39%, and 22%, respectively, on the primary efÞcacy end point—the change from baseline in cumulative morning plus evening TNSS on day 15, as recorded in patient diaries. The improvements for the two active treatment groups were signiÞcantly greater compared with placebo recipients but were not signiÞcantly different from one another. A signiÞcant difference between the intranasal mometasone furoate and intranasal ßuticasone propionate groups was noted on the secondary efÞcacy measure of physician-rated improvement in nasal congestion on day 29 (53% versus 45%, respectively) and at week 8 (56% versus 48%, respectively), as well as on physician-rated improvement in nasal discharge at weeks 8 and 12 (64% versus 58%, respectively). The product label for intranasal mometasone furoate reports consolidated safety data from nearly 4,000 adult and adolescent AR patients (ages 12 years or older) who were treated with either intranasal mometasone furoate or placebo in clinical trials. The incidence of adverse events was comparable between intranasal mometasone furoate-treated patients and placebo recipients. The most commonly reported adverse events in clinical trials were headache (26% of patients treated with 200 µg intranasal mometasone furoate once daily versus 22% of patients treated with placebo), viral infection (14% versus 11%, respectively), pharyngitis (12% versus 10%, respectively), epistaxis/blood-tinged mucus (11% versus 6%, respectively), and coughing (7% versus 6%, respectively). The product label also reports the incidence of the most commonly reported adverse events among nearly 800 pediatric patients (ages 3–11 years) treated with either 100 µg intranasal mometasone furoate once daily or placebo. The most commonly reported adverse events among pediatric patients were similar to those reported in adult studies: headache (17% of patients treated with 100 µg intranasal mometasone furoate once daily versus 18% of patients treated with placebo), coughing (13% versus 15%, respectively), pharyngitis (10% versus 10%, respectively), viral infection (8% versus 9%, respectively), and epistaxis/blood-tinged mucus (8% versus 9%, respectively).

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Branded intranasal mometasone furoate will not face competition from generic competitors until the end of the study period, when the drug faces loss of patent protection and/or market exclusivity in 2012 in Japan and Europe and in 2014 in the United States. Leukotriene Antagonists Overview. Although histamine is the principal mediator of the early-phase allergic response, other proinßammatory mediators—including leukotrienes—play a role in the development of an allergic reaction. Derived from arachidonic acid, leukotrienes are lipid mediators that are released from most of the inßammatory cells present in (or recruited to) the airways. Most development activity pertaining to leukotriene antagonists has been for asthma, as leukotrienes directly mediate bronchoconstriction and have proinßammatory effects. Four leukotriene antagonists have been approved for the treatment of asthma—montelukast (Merck’s Singulair), pranlukast (Ono’s Onon), zaÞrlukast (AstraZeneca’s Accolate), and zileuton (Critical Therapeutics’ Zyßo). Two of these four leukotriene antagonists—montelukast and pranlukast—have also been approved for the treatment of AR. Pranlukast, a product that is exclusive to the Japanese market, received formal approval for the treatment of AR in 2000. (The drug received approval for asthma in Japan in 1995.) In 1998, SmithKline Beecham (now GlaxoSmithKline), which had licensed rights to the drug for Europe and the United States, discontinued development of pranlukast after it had reached Phase III trials for asthma. Because of its limited geographic availability, this agent is not proÞled in greater detail here. Instead, montelukast is discussed as the representative for this class because the drug is currently marketed in all countries under study for the treatment of asthma and is either marketed or used off-label for the treatment of AR in the major markets. Mechanism of Action. Leukotrienes are potent proinßammatory mediators that are derived from arachidonic acid, an unsaturated fatty acid produced from membrane phospholipids. Leukotrienes are generated during the immediate response to antigen provocation and their concentrations increase during the late inßammatory phase of the allergic cascade. Mast cells and basophils generate a mixture of leukotrienes, which act to stimulate the production of airway secretions, enhance the migration of eosinophils, and cause microvascular leakage. Blocking the effect of leukotrienes with therapeutic intervention provides an alternative strategy for treating AR, especially in patients whose allergies involve a more pronounced late-phase response. Currently, two modes of leukotriene inhibition have been developed for respiratory conditions, including AR and asthma: •

Leukotriene receptor antagonists. Leukotriene receptor antagonists are also known as cysteinyl leukotriene receptor antagonists. Cysteinyl leukotrienes exert their biological effect by binding to receptors located on airway smooth

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muscles (Drazen JM, 1999). Therefore, blocking the interaction between leukotrienes and their respective receptors might inhibit the inßammatory effects. Leukotriene synthesis inhibitors. The leukotriene synthesis inhibitors include inhibitors of 5-lipoxygenase (5-LO) and 5-LO-activating protein (FLAP), which are components of the 5-lipoxygenase pathway. This pathway produces several leukotrienes (e.g., leukotriene B4 , cysteinyl leukotrienes) that can cause edema, eosinophil migration, and airway secretion (Drazen JM, 1999). Theoretically, inhibitors of this pathway might reduce airway inßammation.

Montelukast. In 1998, montelukast (Merck’s Singulair) (Figure 9) received approval in the United States and Europe for the treatment of asthma, and, in 2001, the drug received regulatory approval for asthma in Japan, where the drug is marketed by Merck’s Japanese subsidiary, Banyu. In December 2002, montelukast was approved by the FDA for the treatment of SAR in adults and children as young as age two. In March 2005, the drug was approved for the treatment of SAR in its Þrst European market, the United Kingdom. Currently, montelukast is undergoing Phase II trials in Japan for AR. Montelukast is available in three formulations—oral tablet, chewable tablets, and oral granules. Regular oral tablets are indicated for use in AR patients aged 15 years or older, while chewable tablets are indicated for use in pediatric patients ages 2–14 years of age. The oral granule formulation, which can be administered directly or mixed with soft foods, is indicated for asthma patients ages 12–23 months; currently, montelukast is not approved for use in treating AR in children under the age of two. Merck and Schering-Plough were jointly developing a Þxed-dose, once-daily combination therapy comprising Merck’s leukotriene antagonist montelukast and Schering-Plough’s oral second-generation antihistamine loratadine. In January 2002, however, the companies announced that the Þxed-combination tablet did not demonstrate a statistically signiÞcant improvement in the treatment of SAR when compared with each product administered separately in Phase III clinical trials. Although the drug is still listed in Phase III development on the ScheringPlough company Web site, no further development has been reported since 2002.

FIGURE 9. Structure of montelukast sodium.

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Montelukast is a leukotriene D4 antagonist. The drug binds to the CysLT1 receptor—which is found on airway smooth muscle cells, airway macrophages, and proinßammatory cells (e.g., eosinophils)—with high afÞnity and selectivity. By blocking the CysLT1 receptor, montelukast blocks the ability of leukotriene D4 to exert its proinßammatory effects and its ability to stimulate airway secretion production, enhance eosinophil chemotaxis, and contribute to the development of AR symptoms. The efÞcacy of montelukast in treating SAR symptoms was demonstrated in a large, double-blind, placebo- and active-controlled study. A total of 1,214 SAR patients were randomized to receive either 10 mg montelukast, 10 mg loratadine, or placebo once daily in the evening during the spring allergy season. Improvements in mean daytime TNSS (D-TNSS, as measured in the evening before administration of the next dose of study medication) served as the primary efÞcacy endpoint. At the end of the two-week treatment period, montelukast-treated patients demonstrated a mean 18.1% improvement in D-TNSS, which was statistically signiÞcant compared with the mean 13.5% improvement demonstrated by the placebo group and comparable to the 21.8% improvement demonstrated by the loratadine group (van Adelsberg J, 2003). Patients treated with montelukast also demonstrated signiÞcantly greater improvements over placebo-treated patients on a number of secondary endpoints, including the mean nighttime symptom score (mean of scores for difÞculty falling asleep, nighttime awakenings, and nasal congestion on waking); patient and physician global evaluations of treatment efÞcacy; improvements on overall RQLQ scores; mean daytime eye symptoms score (mean of scores for ocular tearing, redness, itching, and pufÞness); and composite scores (mean of D-TNSS and nighttime symptom scores). Loratadine-treated patients demonstrated signiÞcant improvements over placebo-treated patients on each of these secondary end points as well, excluding the nighttime symptoms score assessment. Additionally, treatment with montelukast was associated with a signiÞcant reduction in peripheral blood eosinophil counts compared with placebo; peripheral blood eosinophil counts remained unchanged for both the placebo and loratadine treatment groups. The incidence of adverse events was comparable among all three treatment groups—17%, 15%, and 16% for patients treated with montelukast, loratadine, and placebo, respectively. Comparable results were found in a similar study involving 907 SAR patients randomized to one of four treatment groups—10 mg montelukast, 10 mg loratadine, 10 mg montelukast and 10 mg loratadine given together, or placebo—during the fall allergy season. Primary and secondary end points used in this study were similar to the ones in the aforementioned trial conducted by J. van Adelsberg and colleagues. At the end of the two-week treatment period, mean improvements over baseline D-TNSS for patients treated with montelukast, loratadine, montelukast and loratadine, and placebo were 23.3%, 24.9%, 28.8%, and 12.9%, respectively (Nayak AS, 2002). The difference between each of the three active treatment groups was statistically signiÞcant compared with placebo for this endpoint, as well as on several secondary endpoints including nighttime

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symptom scores, eye symptoms scores, and overall RQLQ scores. In general, patients treated with both montelukast and loratadine demonstrated numerically greater, but not statistically signiÞcant, improvements on symptom scores compared with patients treated with either montelukast or loratadine monotherapy. Although montelukast has demonstrated efÞcacy comparable to that of the antihistamine loratadine in clinical trials, the leukotriene inhibitor has demonstrated efÞcacy inferior to that of the intranasal corticosteroid ßuticasone propionate in a large double-blind study. A total of 705 SAR patients were randomized to receive either 10 mg montelukast or 200 µg intranasal ßuticasone propionate once daily in the evening for 15 days (Ratner PH, 2003). Patients’ improvements over baseline in D-TNSS (as measured by a 100-point VAS) served as the primary end point of the trial, and key secondary end points included the improvement over baseline in daytime individual nasal symptom scores (D-INSS), nighttime total nasal symptom score (N-TNSS, deÞned as the sum of individual scores for nasal congestion upon awakening, difÞculty in going to sleep due to nasal symptoms, and nighttime awakenings attributable to nasal symptoms, as measured by a fourpoint scale for each symptom), and nighttime individual nasal symptom scores (N-INSS). At the end of the treatment period, patients treated with intranasal ßuticasone propionate demonstrated signiÞcantly greater improvements than montelukasttreated patients on all key primary and secondary end points assessed—D-TNSS, D-INSS, N-TNSS, and N-INSS. After 15 days of treatment, patients treated with intranasal ßuticasone propionate demonstrated a 44.0% improvement over baseline D-TNSS compared with an improvement of 31.4% for montelukast-treated patients. Improvements in D-INSS for the intranasal ßuticasone propionate group ranged from a low of 40.5% for nasal congestion to a high of 47.5% for sneezing; corresponding D-INSS for the montelukast group ranged from a low of 29.0% for nasal congestion to a high of 32.9% for sneezing. Improvements in N-TNSS for intranasal ßuticasone propionate- and montelukast-treated patients were 48.7% and 38.9%, respectively, and the improvements in N-INSS were also higher for the intranasal ßuticasone propionate group (ranging from a low of 42.4% for nasal congestion upon waking to a high of 55.1% for nighttime awakenings because of nasal symptoms) compared with the montelukast group (ranging from a low of 32.3% to a high of 44.4% for the same measures). The overall incidence of adverse events was comparable between the two treatment groups—29% and 28% for the intranasal ßuticasone propionate and montelukast groups, respectively—and the most commonly reported adverse events were headache (5% and 7%, respectively), sore throat (3% and 2%, respectively), and diarrhea (3% and 2%, respectively). The product label for montelukast reports that, among 2,200 adult patients (aged 15 years or older) treated in placebo-controlled clinical studies, upper respiratory infection was the only adverse event that occurred in 1% or more of patients and occurred more often in patients treated with montelukast (1.9%) than in placebo recipients (1.5%). The product label further reports that headache, otitis media, pharyngitis, and upper respiratory infection were the only adverse events

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that occurred in 2% or more of patients and occurred more often in montelukast patients than in placebo recipients in a placebo-controlled clinical trial involving 280 pediatric (aged 2–14 years) patients. Montelukast is patent-protected until 2012 in the United States and Europe. The Japanese patent for montelukast does not expire until after the conclusion of the study period. Nonpharmacological Approaches For patients with severe AR who fail to achieve adequate control of symptoms with typical pharmacotherapy, physicians may recommend allergen immunotherapy as a treatment option. Allergen immunotherapy aims to decrease a patient’s allergic sensitivity over time by introducing to the patient’s immune system increasingly larger doses of the substance to which the patient is allergic. Allergen immunotherapy has been shown to decrease patients’ symptoms and reduce the need for AR pharmacotherapy, even for several years after allergen immunotherapy is discontinued (Bousquet J, 1998). A consensus statement issued in August 2000 by experts involved in the American College of Allergy, Asthma, and Immunology’s Immunotherapy in Allergic Asthma conference urged more widespread use of allergen immunotherapy, especially in patients with comorbid asthma and allergy. The experts concluded that speciÞc allergen immunotherapy has been shown to be highly effective in the treatment of allergic asthma, that it can help prevent the development of new allergies and curtail progression of illness, and that it is an effective means of preventing the development of asthma in susceptible individuals. Traditional subcutaneous (SC) allergy immunotherapy, which was Þrst introduced in the early 1900s, involves a series of weekly subcutaneous injections using standardized allergen extracts that are delivered at escalating doses until a maximal dose is achieved; induction of clinical tolerance of the maximal dose usually takes three to six months. Once a patient reaches the maximal, or “maintenance,” dose, the frequency of injections tapers, eventually reaching once per month. Most allergy experts recommend that a patient remain on immunotherapy for a period of at least three years to achieve the best results. Compliance is a major issue with traditional SC immunotherapy because of the need for multiple SC injections, frequent ofÞce visits—which require that the patient remain in a medical facility for at least 20 minutes after receiving the allergy shot to be monitored for serious allergic reactions—and the extended period of time over which the patient must be treated. Although the risk of severe systemic adverse reactions is low with traditional SC immunotherapy—it is estimated at less than 1% of U.S. patients receiving such treatment (Huggins JL, 2004; Lockey RF, 2001)—the risk of death because of anaphylactic reactions is a possibility. In an effort to overcome the drawbacks of traditional SC immunotherapy, new immunotherapy techniques and formulations have been developed, including sublingual immunotherapy. Sublingual immunotherapy was Þrst introduced

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in the early 1990s by the Danish company ALD-Abell´o, and this form of allergen immunotherapy involves the self-administration of allergen extract under the tongue, where it is kept for a couple of minutes before being spit out or swallowed. Like traditional SC immunotherapy, sublingual immunotherapy has been shown to provide long-lasting efÞcacy even after discontinuation, and has been shown to reduce the occurrence of asthma in AR patients (Passalacqua G, 2004). Sublingual immunotherapy is available in Europe and is gaining in popularity, but this more convenient form of allergen immunotherapy is not yet available in the United States or Japan. In November 2004, the FDA granted Greer Laboratories approval to begin Phase I safety and dosing studies with a sublingual formulation of immunotherapy for dust mites. A drawback to noninjectable routes of immunotherapy is that they are not always covered by health insurance providers because evidence to support their efÞcacy is insufÞcient. Despite the effectiveness of allergen immunotherapy, not all AR patients are appropriate candidates for this treatment, and patient selection is a key factor in the ultimate outcome and efÞcacy of such therapy. Allergen immunotherapy should only be employed in AR patients who have documented sensitivity (either with a positive skin test or radioallergosorbent [RAST] test) to speciÞc antigens, as the response to treatment is directly related to the administered allergen to which a patient is allergic. Allergens that are available for immunotherapy include a variety of pollens, pet dander, dust mites, molds, and insect venom, and typically, patients who demonstrate sensitivity to a few dominant allergens will respond more positively than patients who have nonspeciÞc hyperreactivity (DuBuske LM, 2001). For patients with sensitivities to multiple allergens, several allergens can be combined in a single vaccine; however, this combination may affect the concentration of each allergen, and certain allergens may interact with others, therefore impacting efÞcacy (Huggins JL, 2004). Optimally, allergen immunotherapy should be initiated when patients are young or have had the condition for a short period of time, as such patients may have the best response to such therapy (DuBuske LM, 2001). Additionally, allergen immunotherapy should not be administered to AR patients with impaired ability to survive a systemic allergic reaction (e.g., patients with chronic lung disease, unstable angina, myocardial infarction, uncontrolled hypertension, major organ failure), patients taking beta-blocker or angiotensinconverting enzyme (ACE) inhibitor pharmacotherapy (as these agents may mask the early signs of anaphylaxis), or young children under the ages of three or four (Huggins JL, 2004). Also, pregnant women who were tolerating and beneÞting from allergen immunotherapy prior to pregnancy can maintain treatment without dose escalation, but allergen immunotherapy should not be newly initiated during pregnancy (Huggins JL, 2004). Physicians occasionally consider surgical intervention to alleviate obstructive symptoms in very severe AR sufferers with complicating conditions, such as chronic sinusitis, severe septal deviation (causing severe obstruction), or nasal polyps that persist despite maximal pharmacotherapy. However, the need for surgical intervention is very rare in the overall treatment of AR.

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EMERGING THERAPIES Allergic rhinitis (AR) patients in the seven major pharmaceutical markets (United States, France, Germany, Italy, Spain, United Kingdom, and Japan) and the physicians who treat them can choose from a wide selection of available antihistamine and intranasal corticosteroid products that offer good efÞcacy in controlling AR symptoms. However, novel therapies are being pursued as potential therapies for AR, including new antihistamine products, novel intranasal corticosteroids, immunoglobulin E (IgE)-mediated signaling inhibitors, and chemokine antagonists. Many of these emerging drugs—especially those with novel mechanisms of action—are in early Phase II development, and limited clinical trial information for these therapies is available in peer-reviewed journals. This chapter discusses emerging drugs in clinical development for AR and presents the latest Þndings from clinical studies for these therapies. Table 3 summarizes the drug therapies discussed in this chapter. As discussed in “Current Therapies,” immunotherapy is used to decrease a patient’s allergic sensitivity over time by exposing the patient’s immune system to increasingly larger doses of the substance to which the patient is allergic. Despite its proven effectiveness, only a small minority of the AR prevalent population actually undertakes this approach, as traditional immunotherapy requires inconvenient weekly to monthly ofÞce visits for subcutaneous injections and a considerable commitment of time is required to achieve maximal results. In recent years, advances in the Þeld of immunotherapy have notably included the advent of sublingual immunotherapy in Europe. Other novel approaches to immunotherapy include the use of synthetic CpG motifs; high-level expression systems to produce recombinant allergens in bacteria, yeast, and insect cells; and peptide-based approaches (e.g., small peptide fragments of allergens and altered peptide ligands). These novel approaches to immunotherapy may ultimately offer a simpler, more rapid way to build up immunity in AR patients compared with traditional immunotherapy approaches. However, because subcutaneous and sublingual immunotherapy preparations are not classiÞed as traditional pharmaceutical agents, these emerging immunotherapy approaches are not discussed further here. Intranasal Corticosteroids Overview. Intranasal corticosteroids are the most effective drugs available for the treatment of AR. Although they are not always as effective as oral antihistamines at alleviating ocular irritation, intranasal corticosteroids provide excellent relief from other early- and late-phase symptoms. In a meta-analysis of randomized, controlled trials published between 1966 and 1997, intranasal corticosteroids were shown to provide signiÞcantly greater relief of nasal blockage, nasal discharge, sneezing, nasal itch, postnasal drip, and total nasal symptoms compared with oral antihistamines (Weiner JM, 1998). Therefore, intranasal corticosteroids are the recommended Þrst-line treatment for patients experiencing nasal congestion or if AR symptoms occur frequently or are persistent (Bousquet J, 2001).

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TABLE 3. Emerging Therapies in Development for Allergic Rhinitis Compound

Development Phase

Intranasal corticosteroids Intranasal ciclesonide United States Europe Japan 685698 United States Europe Japan NS-126 United States Europe Japan IgE-mediated signaling inhibitors Omalizumab United States Europe Japan R-112 United States Europe Japan Second-generation antihistamines Intranasal olopatadine United States Europe Japan Chemokine inhibitors Bertilimumab (CAT-213) United States Europe Japan

Marketing Company

III III I

Altana Altana Teijin

III III —

GlaxoSmithKline GlaxoSmithKline —

— — III

— — SS Pharmaceuticals/Nippon Shinyaku

—a —a III

—a —a Novartis/Sankyo

II I —

Rigel Pharmaceuticalsb Rigel Pharmaceuticalsb —

PR — —

Alcon — —

— II —

— Cambridge Antibody Technology —

a Omalizumab is currently marketed in the United States by Genentech and Novartis (Tanox also receives

royalties on sales) for use in patients 12 years or older with moderate to severe persistent asthma who have demonstrated reactivity to a perennial aeroallergen. Additionally, the drug is currently preregistered in Europe for allergic asthma. b In January 2005, Rigel Pharmaceuticals entered into a research and license agreement with Pfizer to develop spleen tyrosine kinase inhibitors for allergic asthma and other respiratory disease. The agreement between Rigel and Pfizer currently does not cover R-112, but following completion of additional Phase II studies, Pfizer will have a limited option to license R-112 under a different financial agreement. IgE = Immunoglobulin E; PR = Preregistered.

However, despite their superior efÞcacy, intranasal corticosteroids are prescribed less often than oral antihistamines, primarily because most patients prefer taking oral drugs over intranasal formulations. There are several intranasal corticosteroids currently on the market for the treatment of AR. Popular agents in this class include ßuticasone propionate (GlaxoSmithKline’s Flonase/Flixonase, generics), mometasone furoate (Schering-

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Plough’s Nasonex), beclomethasone dipropionate (GlaxoSmithKline’s Beconase/ Beconase AQ, generics), budesonide (AstraZeneca’s Rhinocort/Rhinocort AQ), and triamcinolone acetonide (Aventis Nasacort/Nasacort AQ). Available intranasal corticosteroid products offer comparable clinical efÞcacy, yet agents in this class differ from one another with regard to other product attributes (e.g., systemic bioavailability, dosing frequency, and use of preservatives). (Differences between the leading intranasal corticosteroids currently on the market are discussed in greater detail in “Current Therapies.”) Because several highly effective intranasal corticosteroids are already available for AR, limited activity—as measured in terms of clinical development of new agents—is taking place within this class. In fact, only three intranasal corticosteroids—Altana/Teijin’s intranasal ciclesonide, GlaxoSmithKline’s 685698 (Avamys/Allermist), and SS Pharmaceuticals (a subsidiary of Boehringer Ingelheim)/Nippon Shinyaku’s NS-126—are in late-stage clinical development for AR. These agents are discussed in greater detail in the following sections. Viatris (formerly ASTA Medica) is conducting Phase II studies with an intranasal formulation of loteprednol etabonate, a “soft steroid” designed to be quickly inactivated upon entering the systemic circulation. (Bausch & Lomb Pharmaceuticals currently markets a 0.5% ophthalmic suspension of loteprednol etabonate as Lotemax for inßammatory eye disorders, and a 0.2% ophthalmic suspension as Alrex for allergic conjunctivitis.) A Phase I pharmacokinetic study has been conducted in healthy male volunteers (Hermann R, 2004), but no clinical efÞcacy data in AR patients have been published to date for the intranasal formulation of the drug. Mechanism of Action. Intranasal corticosteroids ameliorate the symptoms of AR by reducing edema and local inßammation and inducing vasoconstriction. Their anti-inßammatory action stems from their ability to inhibit cytokine and chemokine production and to reduce cellular inÞltration of antigen presenting cells (APCs), T cells, and eosinophils within the nasal mucosa. These agents also inhibit mediator release from basophils and prevent accumulation of mucosal mast cells (although they have little effect on mast-cell mediator release). By reducing the inßux of inßammatory cells into the nasal mucosa, intranasal corticosteroids effectively reduce nasal hyperresponsiveness to allergic stimuli. Intranasal Ciclesonide. Altana (formerly Byk Gulden) and Teijin are developing an intranasal formulation of ciclesonide (an anti-inßammatory corticosteroid) for the potential treatment of SAR and perennial AR (PAR). Altana is conducting Phase III trials in the United States and Europe, and Phase I studies are under way in Japan with Teijin. In 1998, Byk Guilden and Teijin entered into an agreement granting the latter Japanese development and marketing rights to an inhaled formulation of ciclesonide (discussed in the following paragraph). This agreement was extended in 2000 to include the intranasal formulation of the drug. Altana, Aventis (now SanoÞ-Aventis), and Teijin have also developed an inhaled formulation of ciclesonide (Alvesco) for treatment of persistent asthma.

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In January 2005, this product was launched in the United Kingdom, which serves as the reference member state for the European Mutual Recognition Procedure. In October 2004, the FDA issued an approvable letter to Altana and SanoÞ-Aventis for the inhaled formulation of ciclesonide, and in January 2004, Teijin Þled for regulatory approval in Japan. In 2001, Byk Gulden and Aventis entered into an agreement to codevelop and copromote the inhaled formulation of ciclesonide. However, it does not appear that the agreement between Altana and SanoÞ-Aventis has been extended to include the intranasal formulation. Intranasal ciclesonide, like other intranasal corticosteroids, ameliorates the symptoms of AR by reducing edema, inducing vasoconstriction, inhibiting cytokine and chemokine production, and reducing cellular inÞltration of antigenpresenting cells, T cells, and eosinophils within the nasal mucosa. Ciclesonide is not directly active, but is cleaved intracellularly to the active drug, desisobutyrylciclesonide (des-CIC), by endogenous esterases. Des-CIC has demonstrated a 100-fold greater binding afÞnity at the rat glucocorticoid receptor compared with its pro-drug, ciclesonide (Stoeck M, 2004). Additionally, the pro-drug is poorly absorbed into the systemic circulation, thus the “on-site activation” prevents systemic side effects of corticosteroid therapies, which are a concern to physicians and patients. Results from a Phase II study were announced in a March 2004 Altana company press release and were presented at the 2004 Annual Meeting of the American Academy of Allergy, Asthma, and Immunology (AAAAI) in San Francisco (Susman E, 2004). A total of 736 patients with SAR were randomized to receive one of four doses of intranasal ciclesonide—25 µg/day, 50 µg/day, 100 µg/day, or 200 µg/day—or placebo for two weeks. At the end of 14 days, all groups treated with intranasal ciclesonide demonstrated greater improvements over baseline total nasal symptom score (TNSS) than the group treated with placebo. The improvements were −4.81 points, −4.79 points, −5.33 points, and −5.83 points for the 25 µg/day, 50 µg/day, 100 µg/day, and 200 µg/day treatment groups, respectively; placebo recipients experienced an improvement of −4.19 points. The improvements were statistically signiÞcant for the 100 µg/day and 200 µg/day intranasal ciclesonide treatment groups, but not for the groups treated with 25 µg/day and 50 µg/day. The most commonly reported adverse events were headache and pharyngitis, but the incidence of adverse events in this study was relatively low. The efÞcacy of intranasal ciclesonide was evaluated in a randomized, placebocontrolled, double-blind crossover trial involving 24 people with a history of AR who were asymptomatic at the time of the study (Schmidt BM, 1999). Study participants were treated with either 200 µg intranasal ciclesonide per nostril per day or placebo for seven days; a washout period of at least 14 days separated the treatment periods. Study participants were subjected to intranasal allergen challenge with pollen extracts for two days prior to treatment initiation and at two hours after administration of study medication on each of the treatment days. In this study, rhinal airßow was measured by anterior rhinomanometry 5 and 30

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minutes after allergen challenge; nasal obstruction, itching, and rhinorrhea were measured on a 10 cm visual analogue scale (VAS). At the conclusion of the trial, researchers found that rhinal airßow improved signiÞcantly from day 5 and nasal obstruction improved signiÞcantly from day 2 in patients treated with intranasal ciclesonide, compared with placebo recipients. A signiÞcant improvement in itching and rhinorrhea was also noted in the intranasal ciclesonide group. Intranasal ciclesonide was shown to be well tolerated both systemically and locally in this trial, and no adverse events were considered to be related to the drug. One case of each of the following adverse events was noted in this study: lumbar pain, infectious rhinitis, biliary pain, tonsillitis, and diarrhea. Use of intranasal ciclesonide did not cause irritation of the nasal mucosa in this study. The safety and tolerability of repeated intranasal ciclesonide doses were evaluated in a placebo-controlled, double-blind, modiÞed sequential-dose study involving 40 healthy volunteers and 8 asymptomatic SAR patients (Nave R, 2004). Patients were randomized to receive either 50, 100, 200, or 400 µg intranasal ciclesonide or placebo once or twice daily for a period of 14 days. During the course of the study, 41 study participants experienced a total of 134 adverse events, 74 of which were deemed to be possibly or probably related to study medication. Of the side effects experienced in this study, 97% were considered to be mild in intensity, and the remaining 3% were considered to be moderate. The most common adverse events in this study were headache, fatigue, and rhinitis. Intranasal ciclesonide was shown to have low systemic bioavailability and no effect on adrenal function. Very few detailed data pertaining to the efÞcacy or safety of intranasal ciclesonide in AR patients have appeared in peer-reviewed journals. However, less hypothalamic-pituitary-adrenal (HPA) axis suppression was observed in asthma patients treated with high-dose inhaled ciclesonide compared with highdose ßuticasone propionate (Lee DKC, 2004). This fact suggests that intranasal ciclesonide may provide a safe corticosteroid therapy for treatment of AR. However, the drug would be entering a highly competitive intranasal corticosteroid market served by established agents that allergy experts regard as highly effective with low incidences of side effects. In particular, two newer nasal steroids, ßuticasone propionate and mometasone furoate, both possess extremely low systemic bioavailability (less than 1%) (Salib RJ, 2003[b]). 685698. GlaxoSmithKline is developing a novel intranasal corticosteroid product, 685698 (Avamys/Allermist), for the treatment of AR. (GlaxoSmithKline is also developing an inhaled combination product containing 685698 and a longacting beta-2 adrenergic agonist, 159797, as a potential treatment for asthma and chronic obstructive pulmonary disease [COPD].) The intranasal formulation of 685698 is currently in Phase III development in the United States and Europe; there does not appear to be any ongoing development for 685698 in Japan. According to the GlaxoSmithKline company Web site, the company expects to Þle a new drug application (NDA) in the United States and a marketing authorization application (MAA) in Europe in 2006.

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The only information pertaining to the clinical efÞcacy and/or safety of 685698 for AR that has been released to date is a comment in a December 2003 GlaxoSmithKline company press release regarding several of the company’s pipeline projects. According to the release, intranasal 685698 has demonstrated superior efÞcacy to intranasal ßuticasone propionate in relieving nasal symptoms after allergen exposure. However, the press release did not include detailed efÞcacy and safety information for the drug, nor have such data been published in a peer-reviewed journal. NS-126. SS Pharmaceuticals (a subsidiary of Boehringer Ingelheim) and Nippon Shinyaku are codeveloping the intranasal corticosteroid NS-126. Phase III trials are under way in Japan; there does not appear to be any ongoing development with this agent in the United States and Europe. The two companies are also developing an inhaled formulation of NS-126 as a potential treatment for asthma. IgE-Mediated Signaling Inhibitors Overview. IgE is the key antibody involved in potentiating allergic reactions. Cross-linking of IgE by antigen is a key step in the initiation of the earlyphase allergic response and the subsequent development of AR symptoms. Thus, targeting IgE and intracellular signaling mediated by IgE within effector cells (e.g., mast cells, basophils) represents a potential therapeutic approach to treating AR. Currently, one agent in this class, the anti-IgE monoclonal antibody (mAb) omalizumab (also known as rhuMAb-E25; Genentech/Novartis/Sankyo/Tanox Biosystems’ Xolair), is marketed for the treatment of allergic asthma and has shown beneÞt in treating AR in clinical studies. A second agent, Rigel Pharmaceuticals’ R-112, targets the spleen tyrosine kinase (SYK), an intracellular regulator of cell IgE-mediated signaling, and is in development for AR. Both of these agents are discussed in greater detail in the following sections. Mechanism of Action. IgE plays a major role in allergic reactions by binding cell-surface IgE receptors and thereby forming receptor-IgE complexes. Crosslinking of these complexes with neighboring complexes initiates a signal transduction pathway that ultimately results in the release of histamine and other inßammatory mediators (e.g., proteases, leukotrienes [LTs], prostaglandins, cytokines) from mast cells and basophils. Agents in this class aim to block the binding of IgE to its receptor on effector cells or to disrupt the subsequent signal transduction pathway that results in the release of inßammatory mediators from mast cells and basophils. Omalizumab. Omalizumab (Genentech/Novartis/Sankyo/Tanox’s Xolair) is a recombinant, humanized anti-IgE mAb. The drug received FDA approval in June 2003 for use in patients 12 years or older with moderate-to-severe persistent asthma who have demonstrated reactivity to a perennial aeroallergen. In July 2004, Novartis Þled omalizumab for regulatory approval in Europe for treatment of allergic asthma. The drug is currently in Phase III development in Japan for bronchial asthma and AR. A U.S. Phase III trial began in May 2004 to evaluate

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omalizumab in pediatric allergic asthma patients, and a U.S. Phase II study was initiated in July 2004 to evaluate omalizumab’s potential as a therapy for peanut allergy. Omalizumab was jointly developed by Genentech, Novartis, and Tanox. In the United States, Genentech and Novartis share marketing rights for the drug, and Novartis has marketing rights in regions outside the United States. Tanox receives royalties based on worldwide sales of the product and will also share development and marketing responsibilities with Novartis in certain Asian markets (excluding Japan). In February 2003, Sankyo signed a licensing agreement with Novartis in which both companies agreed to codevelop and comarket omalizumab in Japan. The development status of omalizumab for AR is uncertain. Genentech, Novartis, and Tanox originally Þled in June 2000 for U.S. and European regulatory approval of omalizumab for asthma and AR. However, in July 2001, the FDA issued a complete response letter requesting additional preclinical, clinical, and pharmacokinetic data for omalizumab. In a December 2002 company press release, Genentech announced that the company (with Novartis) had submitted an amendment to the original biologics licensing application (BLA) for omalizumab to the FDA, and that additional data would be submitted to the European Medical Evaluations Agency (EMEA, now European Medicines Agency) as part of a resubmitted marketing application in 2004. When the December 2002 company press release was issued, Genentech and Novartis were seeking approval for omalizumab for the allergic asthma indication, not for asthma or AR independently. In July 2003, the FDA approved omalizumab for allergic asthma, and in July 2004, Novartis Þled for European approval for omalizumab for the same indication. Currently, no mention of active development of omalizumab for AR appears on the Genentech, Novartis, or Tanox company Web sites. Additionally, at the 2004 annual meeting of the AAAAI, Novartis company representatives said there were no plans to seek formal approval for omalizumab as a treatment for AR. However, according to the Sankyo company Web site, the drug is in Phase III development in Japan for bronchial asthma and AR. Omalizumab binds free IgE at the same site as the high-afÞnity IgE receptor (FcεRI) found on effector cells such as mast cells and basophils, thereby preventing IgE binding to these cells. Importantly, omalizumab is nonanaphylactogenic (i.e., it cannot cause serious allergic reactions) because it cannot bind IgE that is already bound to mast cells or basophils. The anti-IgE antibody also binds any IgE that dissociates from FcεRI or FcεR2 receptors on effector cells, a behavior that means that it essentially “mops up” unbound IgE molecules. Omalizumab also down-regulates IgE receptor expression by reducing IgE levels (MacGlashan DW, 1997). The dual action of omalizumab on IgE and its receptor contributes to the symptom improvement observed in the clinical trials. Researchers have shown that FcεRI densities on basophils from both allergic and nonallergic subjects range from 104 to 106 per cell, and hundreds of thousands of IgE receptors are usually occupied with IgE (MacGlashan DW, 1997). Additionally, only a few thousand IgE molecules are required to produce a half maximal release of histamine from basophils exposed to speciÞc allergens. IgE

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levels must therefore be nearly undetectable before anti-IgE mAb therapy will produce therapeutic effects. To achieve optimal response, dosing of the agent has to be individualized to match an AR patient’s total IgE level. Additionally, according to omalizumab’s product label, its mean serum elimination half-life in asthma patients is 26 days; IgE’s half-life is one to two days. This extended half-life allows omalizumab to be dosed once every two to four weeks, a fairly convenient dosing schedule over a six- to eight-week pollen season. A Phase II clinical trial showed that omalizumab signiÞcantly reduced serum levels of free IgE in a dose-dependent manner in AR patients (Casale TB, 1997). However, the authors were unable to demonstrate statistically signiÞcant improvement in symptoms within the dose range tested (0.15–0.50 mg/kg). A subsequent 12-week, double-blind, dose-ranging (50–300 mg), placebo-controlled trial in patients with moderate-to-severe ragweed-induced SAR provided evidence that lowering systemic free IgE levels with omalizumab provides clinical beneÞt in patients (Casale TB, 2001). A total of 536 patients were randomized to receive either 50 mg, 150 mg, or 300 mg omalizumab or placebo via subcutaneous injection. Patients with baseline IgE levels of 150–700 IU/mL were treated every three weeks; patients with baseline levels of 30–150 IU/mL were treated every four weeks. Treatment was initiated approximately two weeks prior to onset of the ragweed pollen season to assess whether blocking IgE binding before and during the pollen season could reduce SAR symptoms. During the course of the pollen season, the average daily nasal symptom severity score (DNSSS)—the primary efÞcacy end point—was 23% lower in the 300 mg omalizumab group than in the placebo group. (DNSSS was the average of patient scores for self-assessed severity and duration of nasal symptoms [sneezing, rhinorrhea, itchy nose, and stuffy nose].) This difference was statistically signiÞcant. Patients receiving 300 mg of omalizumab also experienced almost no difference in nasal symptom severity score between the Þrst treatment day and peak season. There was a signiÞcant association between the severity of nasal symptoms and free IgE levels in this study: lower free IgE levels were associated with the lowest symptom severity scores. In addition, the 300 mg omalizumab group experienced lower scores (indicating improved quality of life) on the Rhinoconjunctivitis Quality of Life Questionnaire (RQLQ) than placebo recipients throughout the 12-week treatment period. (The RQLQ measures overall well-being of rhinoconjunctivitis patients on seven domains [activity limitations, sleep disturbances, non-hay fever symptoms, practical problems, nose symptoms, and emotional function] and is based on a 0 to 6 unit scale; lower scores indicate better AR-related quality of life.) The most commonly reported adverse events in this study were headache (11.6% of the 300 mg omalizumab group, 15.7% of the 150 mg omalizumab group, 15.3% of the 50 mg omalizumab group, and 16.2% of the placebo group); upper respiratory tract infection (6.2%, 8.2%, 9.5%, and 8.1%, respectively); and viral infection (5.4%, 8.2%, 6.6%, and 6.6%, respectively). A follow-up to this 12-week study in ragweed-induced SAR patients demonstrated that readministration of omalizumab is safe during a second ragweed

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pollen season (Casale TB, 2000). The rate and pattern of adverse events observed in the follow-up study were similar to those observed in the primary study. There were no serious adverse events related to omalizumab retreatment, and no patients developed anti-omalizumab antibodies. Omalizumab was shown to provide effective symptom control and to be a safe and well-tolerated therapy for patients with moderate-to-severe symptomatic PAR (Chervinsky P, 2003). In a randomized, double-blind Phase III study involving 289 patients, study participants (ages 12–70 years) were randomized to receive subcutaneous injections of either placebo or omalizumab (at least 0.016 mg/kg/IgE [IU/mL] either once or twice monthly) throughout the 16-week trial. Omalizumab doses were tailored to each patient’s baseline body weight and serum free IgE levels. Throughout the entire 16-week study period, omalizumabtreated patients demonstrated a lower average DNSSS—the primary efÞcacy end point—compared with placebo recipients. A decrease in symptom severity category (i.e., controlled, mild, moderate, and severe, depending on DNSSS) was noted in 69% of omalizumab-treated patients compared with 49% of placebo recipients. Additionally, 28% of patients in the omalizumab group and 10% of placebo-treated patients achieved a DNSSS of less than 0.75, corresponding to a severity category of “controlled.” Omalizumab was also shown to be superior to placebo on improvements in RQLQ scores and nasal congestion (stuffy nose), two of the secondary efÞcacy measures evaluated in this study. Mean RQLQ scores improved 51% and 38% for omalizumab- and placebo-treated patients, respectively, during the course of the 16-week study. Additionally, omalizumabtreated patients demonstrated a mean 31% improvement over baseline score on the measure of daily stuffy nose severity during the previous 24 hours, and a 30% improvement over baseline score on the measure of daily stuffy nose severity upon waking. Corresponding improvements for placebo-treated patients on these two measures were 14% and 11%, respectively. Omalizumab was well tolerated by PAR patients (Chervinsky P, 2003). At least one adverse event was reported by 77% of omalizumab-treated patients and 85% of placebo recipients; most adverse events were of mild to moderate intensity. The most commonly reported adverse events in this study were upper respiratory tract infections (16.7% of omalizumab recipients and 13.1% of placebo recipients); headache (15.3% and 23.4%, respectively); and nasopharyngitis (12.5% and 13.8%, respectively). Three cases of serious adverse events occurred during this trial—one in the omalizumab groups (infectious mononucleosis) and two in the placebo group (appendicitis and recurrent disk herniation)—but none of the three cases was deemed treatment-related. R-112. Rigel Pharmaceuticals is developing R-112, an intranasal SYK inhibitor, as a potential treatment for AR. Phase II studies are currently underway in the United States. According to an October 2002 company press release, an initial Phase I safety study was conducted in the United Kingdom, but no further European development has been reported. Additionally, no development has been reported in Japan.

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In 2004, Rigel Pharmaceuticals stated that it was seeking a development partner for R-112. In January 2005, Rigel entered into a research and license agreement with PÞzer; the focus of the agreement is to develop SYK inhibitors for allergic asthma and other respiratory disease. The agreement between Rigel and PÞzer currently does not cover R-112, but, following Rigel’s completion of additional Phase II studies, PÞzer will have a limited option to license R-112 under a different Þnancial agreement. SYK is an intracellular protein that is expressed in a number of leukocytes, including mast cells, macrophages, eosinophils, neutrophils, T cells, and B cells (Stenton GR, 2002). The protein induces tyrosine phosphorylation of numerous other proteins involved in the allergic inßammation signaling cascade. Studies have demonstrated that SYK-deÞcient mast cells neither degranulate nor synthesize leukotrienes and secrete cytokines following stimulation via the FcεRI receptor (Costello PS, 1996). Additionally, SYK-deÞcient variant cell lines from rat basophilic leukemia RBL-2H3 cells fail to release histamine following FcεRI receptor aggregation (Zhang J, 1996). In addition, transfection of wild-type rat SYK into these variant cells resulted in reconstitution of FcεRI receptor-mediated histamine release. These studies demonstrate the essential role that SYK plays in allergic reactions and how inhibition of SYK can preclude the synthesis and release of mediators involved in the allergic response. A Phase II, randomized, placebo-controlled, two-day study was conducted to evaluate the efÞcacy, safety, and onset of action of R-112 in 319 SAR patients (Meltzer EO, 2005). The study was conducted at two outdoor locations (one in Atlanta and one in San Diego), and patients were treated with either two sprays R-112 per nostril (for a total dose of 3 mg per nostril) or placebo twice daily at intervals four hours apart. The primary efÞcacy end point in this study was the effect of treatment on the combined Global Symptom Complex (GSC) score, which was the sum of scores for sneezing, nasal congestion, rhinorrhea, and nasal itch at three separate time points. Treatment with R-112 was associated with a mean 7.0-point reduction (38% improvement) in GSC score on day 1, compared with a mean 5.4-point reduction (29% improvement) for patients treated with placebo. The difference between the two groups was statistically signiÞcant. Similar signiÞcant results were observed on day 2; the group treated with R-112 demonstrated a mean 5.6-point reduction (35% improvement) in GSC score compared with a mean 4.3-point reduction (27% reduction) for placebo-treated patients. Additionally, treatment with R-112 was associated with signiÞcant improvements compared with placebo recipients in individual symptom scores for nasal congestion, itchy nose, sneezing, rhinorrhea, nose blows, postnasal drip/throat clearing, and cough on both days of the study. Notably, patients treated with R-112 began to demonstrate a signiÞcant improvement over placebo recipients as early as 30 minutes after dosing, and the reduction in symptoms persisted for the entire duration of the day in the outdoor park setting. The incidence of adverse events was similar for both treatment groups (21.3% of patients treated with R-112 and 20.6% of patients treated with placebo), and no serious drug-related adverse events were noted during the

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trial. The most commonly reported adverse events were somnolence (1.3% of R-112 recipients versus 0% of placebo recipients), epistaxis (bloody nose; 0.6% versus. 1.3%, respectively), and burning at the site of application (1.3% versus 0.6%, respectively). In a December 2003 company press release, Rigel announced the results from a ten-day study that evaluated the safety of three doses of R-112 compared with placebo. Each of the three R-112 dosing groups included six study participants. Although no speciÞc safety data were included in the company press release, Rigel reported that participants treated with R-112 and placebo demonstrated comparable results on a number of safety parameters. No abnormalities in blood counts, blood chemistry, electrocardiography, or spirometry (airway function tests) were noted, and patients treated with R-112 did not demonstrate local nasal irritation. Rigel announced the results of a Phase I/II study in a July 2003 company press release. This study evaluated the efÞcacy and safety of a single intranasal dose of R-112 in 20 patients with asymptomatic AR. Although the company press release did not include speciÞc efÞcacy or safety results, Rigel reported that treatment with R-112 was associated with a signiÞcant or consistently positive trend in reducing inßammatory mediator release. No signiÞcant adverse events were noted in this study. R-112 may have a signiÞcant advantage over other AR pharmacotherapies due to its novel mechanism of action and its rapid onset of action. R-112 blocks mast cell activation and subsequent release of multiple inßammatory mediators by disrupting the signal transduction pathway following binding of IgE to the FcεRI receptor. Blocking the release of multiple inßammatory mediators represents a signiÞcant improvement on the action of many other allergy medications (e.g., antihistamines, leukotriene antagonists), which target a single inßammatory mediator after it has already been released from cells. In an August 2004 company press release, Rigel acknowledged that intranasal corticosteroids—a mainstay of AR therapies—can inhibit multiple inßammatory mediators, but the company points out that intranasal corticosteroids may require multiple days of administration before a positive effect on symptoms is noted. In the Phase II study conducted at two outdoor locations, R-112’s beneÞcial effect on AR symptoms was noted as early as 30 minutes after dosing. Therefore, the company claims that R-112’s rapid onset of action may offer a signiÞcant advantage over intranasal corticosteroids for patients who want rapid relief of their AR symptoms. However, the price of R-112 may prevent the product from achieving widespread use among AR sufferers. It has been estimated that the annual cost of R-112 therapy may reach $1,800 per year, a Þgure that is much higher than the annual cost of antihistamines and intranasal corticosteroids (Carrel L, 2004). Therefore, R-112 may be relegated to third-line use behind antihistamines and intranasal corticosteroids, especially if cost-conscious third-party payers in the United States continue to remove more expensive AR therapies from their formularies as additional agents are switched to OTC status.

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Second-Generation Antihistamines Overview. Second-generation antihistamines are the most commonly prescribed class of drugs for AR because these agents are very effective at ameliorating many of the symptoms associated with AR (e.g., sneezing, nasal itching, rhinorrhea, ocular irritation). Numerous second-generation antihistamines are currently on the market. As discussed in “Current Therapies,” Þrst-generation antihistamines, which are readily available as OTC products, are associated with a number of side effects—most notably fatigue, drowsiness, and performance impairment—that ultimately make them less attractive than second-generation agents. Second-generation antihistamines, by comparison, are equally as effective as Þrst-generation agents in controlling AR symptoms, yet offer many advantages over their predecessors (i.e., they are associated with mild or no sedation, do not affect performance, have no anticholinergic effects, and have longer durations of action for reduced dosing frequency). The antihistamine market is saturated with numerous product offerings and there is very little ongoing clinical development in this class. Currently, only one second-generation antihistamine product is in late-stage clinical development: Alcon is developing an intranasal formulation of olopatadine, which is currently on the market as an ophthalmic formulation (Patanol) and as an oral formulation in Japan (Kyowa’s Allelock). The intranasal formulation of olopatadine is discussed in greater detail in the following section. Mechanism of Action. Subsequent allergen (antigen) challenge in the nasal mucosa of a sensitized AR patient results in cross-linking of IgE by antigen, which in turn causes degranulation of high-afÞnity IgE receptor (FcεRI)+ cells (e.g., mast cells, basophils). Degranulation of these cells releases a variety of inßammatory mediators involved in the acute-phase allergic response. One of the key inßammatory mediators released is histamine, the primary mediator of early AR symptoms. There are four types of histamine receptors in the human body—H1 , H2 , H3 , and H4 —and H1 is the receptor through which histamine exerts most of its effects in allergic disease. The H1 receptor demonstrates constitutive activity in the human body, even in the absence of ligand binding, as two states of the H1 receptor—the active states and the inactive state—exist in equilibrium (Simons FER, 2004). Histamine preferentially binds to the active form of the receptor, shifting the balance between the two receptor states toward the active form. The active form of the receptor then initiates a G protein-mediated signal transduction cascade that results in the degranulation of FcεRI+ cells. Antihistamines used to treat AR bind to the inactive form of the H1 receptor and stabilize this conformation, therefore shifting the equilibrium toward the inactive state. The inactive state of the receptor is unable to initiate the signal transduction cascade that ultimately results in degranulation of mast cells and basophils. Thus, the resultant effect of antihistamines is to block the release of inßammatory mediators which initiate a host of physiological changes that lead to AR’s early-phase symptoms (e.g., sneezing, itching, rhinorrhea, ocular irritation). Antihistamines

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are less effective, however, at controlling nasal congestion, because histamine is not the primary mediator of this symptom. An enhanced understanding of the molecular pharmacology of H1 antihistamines has led to their reclassiÞcation as inverse agonists; they were previously viewed as H1 receptor antagonists (Simons, FER, 2004). However, for the purpose of this section, this discussion will refer to H1 antihistamines as antihistamines. Intranasal Olopatadine. Alcon is developing Patanase, an intranasal formulation of the second-generation antihistamine olopatadine, as a potential treatment for SAR in the United States. No ongoing development has been reported in Europe and Japan. In an April 2005 company press release, Alcon announced that the FDA has accepted its new drug application for the intranasal formulation of olopatadine. Kyowa currently markets an oral formulation of olopatadine (Allelock) in Japan, and the company has licensed U.S. rights for ophthalmic and nasal formulations of the drug to Alcon. Alcon currently markets an ophthalmic formulation of olopatadine (Patanol). Olopatadine exerts its therapeutic beneÞt by blocking the effects of histamine released from mast cells during an allergic reaction. Topical administration through a nasal formulation aims to diminish the nasal symptoms characteristic of AR: rhinorrhea, sneezing, and itchy and stuffy nose. In a March 2004 company press release, Alcon announced the results of a large-scale Phase III study evaluating the efÞcacy of the nasal formulation of olopatadine. These results were also presented at the 2004 Annual Meeting of the AAAAI in San Francisco. In this randomized, double-blind, placebo-controlled study, 677 patients with SAR were randomized to receive twice-daily administrations of either 0.6% or 0.4% olopatadine nasal spray, or placebo. Patients were asked to record both an instantaneous and reßective assessment of nasal symptom severity in the morning and the evening for two weeks. At the end of the study, patients treated with 0.6% olopatadine and 0.4% olopatadine demonstrated a 30.1% and 27.6% improvement, respectively, on the primary efÞcacy end point—the change from baseline in the total nasal symptom score (TNSS), deÞned as an average of morning and evening reßective severity scores for stuffy nose, runny nose, itchy nose, and sneezing. The corresponding improvement for the placebo group was 18.7%. These differences were statistically signiÞcant for both olopatadine treatment groups. A signiÞcant difference over placebo was also noted for both olopatadine groups on the measure of TNSS for instantaneous assessments of nasal symptoms. On this secondary measure, treatment with 0.6% olopatadine and 0.4% olopatadine were associated with 26.2% and 24.3% improvements over baseline, respectively, compared with a 15.8% improvement for placebo-treated patients. Olopatadine nasal spray was shown to be safe and well tolerated in this study. Patients who participated in this Phase III study were also asked to Þll out the RQLQ. At the end of the two-week study period, the groups treated with 0.6% and 0.4% olopatadine nasal sprays both demonstrated a signiÞcant 1.1

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unit improvement on the RQLQ compared with a 0.8 unit improvement for the placebo group. Olopatadine nasal spray was shown to have a rapid onset of action in a doseranging study involving 320 SAR patients (Patel P, 2004). Three different dose concentrations of olopatadine nasal spray—0.6%, 0.4%, and 0.2%—were shown to improve nasal symptoms within 30 minutes following drug administration. Additionally, a dose response was noted in this trial, with the most pronounced improvement in nasal symptoms occurring with 0.6% olopatadine nasal spray treatment. Other intranasal antihistamine formulations are currently on the market, however. According to the available information for intranasal olopatadine, this drug does not appear to offer any clear advantages over such competing products. Chemokine Inhibitors Overview. The late-phase allergic response is characterized by recruitment of leukocytes (e.g., eosinophils, basophils, neutrophils, T cells) to the airway tissue (submucosa, epithelium, airway lumen). The accumulation of leukocytes—especially eosinophils—and their products in the airway tissues in the hours following allergen exposure is primarily responsible for the congestion and increased mucus production characteristic of the late-phase allergic response. A number of cytokines have been implicated in leukocyte recruitment activity in AR, including the chemokine family of proteins. These agents, along with their respective receptors (members of the G protein-coupled receptor (GPCR) superfamily), play a critical role in mediating chemotaxis (directional movement) of leukocytes to sites of inßammation and infection in the body. T-helper 2 (TH 2) cells, mast cells, and eosinophils—three leukocytes that play prominent roles in allergic responses—preferentially express three cysteine-cysteine chemokine receptors (CCRs): CCR3, CCR4, and CCR8 (Chantry D, 2002). Therefore, blocking these three receptors, or inhibiting the chemokines that bind to these receptors, represents potential therapeutic targets in the treatment of AR. Two chemokine inhibitors—Cambridge Antibody Technology’s bertilimumab (CAT-213) and GlaxoSmithKline’s 766994—have reached Phase II development for AR. Bertilimumab is an mAb that neutralizes eotaxin1 , a potent, highly speciÞc chemokine that attracts eosinophils from the bloodstream into the airway tissue, and is discussed in greater detail in a following section. 766994 is an oral antagonist of CCR3, a chemokine receptor found on the surface of numerous cell types, including eosinophils, TH 2 cells, mast cells, basophils, macrophages, and airway epithelial cells (Erin EM, 2002). Eotaxin1 binds to CCR3, thus, 766994 acts to prevent the interaction between CCR3 and its ligand, eotaxin1 . Inhibiting this receptor-ligand interaction is a novel approach to preventing chemotaxis of leukocytes into nasal tissue and the subsequent AR symptoms that develop during the late-phase allergic reaction. 766994 is in early Phase II development, and no data have been published to date in peer-reviewed journals demonstrating 766994’s effect on AR symptoms in human subjects.

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Mechanism of Action. Two general approaches can be employed to inhibit the effects of chemokines: neutralize the chemokine through antibody inhibition, or block the chemokine receptor. Binding of a chemokine-speciÞc antibody to a given chemokine can inhibit the latter’s ability to bind with its receptor, thus precluding subsequent physiologic effects. Alternatively, chemokine receptor inhibitors act by binding the receptor and disrupting the conformation of its extracellular domain, thereby inhibiting ligand binding to the receptor. Bertilimumab (CAT-213). Cambridge Antibody Technology (CAT) is developing bertilimumab (CAT-213), a human IgG4 mAb that targets the chemokine eotaxin1 (also known as CCL11), as a potential treatment for AR. Phase II studies are underway in the United Kingdom. CAT is also conducting trials to evaluate bertilimumab’s potential in other allergic conditions: the drug is in Phase II for allergic conjunctivitis and in preclinical development for asthma. In its 2003 annual report, CAT announced plans to seek a development partner for bertilimumab. CAT further reported in a September 2004 earnings conference call that the company is still seeking a licensing partner for the drug. Eosinophils are a type of leukocyte that plays a key role in the late-phase allergic reaction, and eotaxin1 is a potent, highly speciÞc chemokine that attracts eosinophils from the bloodstream into the airway tissue. (Eotaxin1 also plays a role in the chemotaxis and activation of other leukocytes, including basophils, mast cells, and T cells [Amerio P, 2003].) Once eosinophils (along with other leukocytes) accumulate in airway tissues, these cells release numerous inßammatory mediators—including cytokines, basic polypeptides, and leukotrienes—that can damage airway endothelial cells and the extracellular matrix. The accumulation of eosinophils (eosinophilia) in the nasal tissue is a hallmark of AR, and the accumulation of these cells and their inßammatory products is primarily responsible for the congestion and increased mucus production characteristic of the late-phase allergic response. Bertilimumab binds to and neutralizes eotaxin1 , thereby preventing this chemokine from recruiting eosinophils and other leukocytes into airway tissues. Inhibiting eosinophilia in turn leads to a reduction in AR symptoms. In a double-blind Phase II study, treatment with bertilimumab prior to allergen challenge was shown to reduce the number of inÞltrating eosinophils and mast cells in the nasal submucosa of patients with SAR (Salib R, 2003[a]). A total of 48 patients were treated with single intranasal doses of 10 mg bertilimumab or placebo, or single intravenous (IV) doses of 50 mg, 200 mg, or 500 mg bertilimumab or placebo, 30 minutes prior to being subjected to grass pollen allergen challenge. Nasal lavage (irrigation of the nasal cavity and nasal mucosa) was performed at Þve time points following allergen challenge (at 30 minutes, one hour, two hours, six hours, and eight hours); a nasal tissue sample was also obtained through biopsy six hours following allergen challenge. Compared with placebo, patients treated with 10 mg intranasal, 50 mg IV, or 500 mg IV bertilimumab demonstrated signiÞcant reductions of 42%, 40%, and 46%, respectively,

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in the number of submucosal mast cells. Patients treated with 200 mg IV bertilimumab demonstrated a 24% decrease in the number of submucosal mast cells compared with placebo, but this difference was not statistically signiÞcant. The group treated with 10 mg intranasal bertilimumab also demonstrated a signiÞcant reduction in submucosal eosinophil inÞltrate, but the corresponding reduction was not signiÞcant for any of the IV bertilimumab groups. In a second similarly designed double-blind Phase II study, pretreatment with bertilimumab was shown to attenuate nasal obstruction in patients with SAR (Pereira S, 2003). A total of 52 patients were treated with single doses of either intranasal bertilimumab or placebo, or IV bertilimumab or placebo, 30 minutes prior to grass pollen allergen challenge; doses used in this study were the same as those used in the aforementioned study conducted by R. Salib and colleagues. Treatment with 10 mg intranasal bertilimumab was associated with the greatest attenuation of nasal obstruction compared with placebo, as assessed by acoustic rhinometry measurements taken prior to allergen challenge and six hours thereafter. However, neither intranasal or IV bertilimumab was shown to signiÞcantly alter nasal symptoms or peak inspiratory ßow in this study. Three cases of adverse events were deemed to be possibly related to bertilimumab treatment in this study: one case of ßulike symptoms and fever, one case of vasovagal attack and wheezing, and one case of nosebleed. While the mechanism of action for this agent seems promising—targeting a key chemokine involved in the chemotaxis of eosinophils—it is important to note that other substances have been implicated in leukocyte recruitment activity characteristic of AR. For example, investigators have found that interleukin (IL)-5, IL-16, and leukotriene B4 have chemoattractant activity for eosinophils, and IL-4, IL-13, and tumor necrosis factor-alpha (TNF-α) increase the expression of cellular adhesion molecules (CAMs) on endothelial cells, thereby facilitating the attachment and inÞltration of leukocytes into airway tissues. Therefore, although eotaxin1 is a potent eosinophilic chemokine, numerous other cytokines and inßammatory mediators need to be targeted to completely inhibit eosinophilia that occurs in AR. Very little information is available on bertilimumab. The only clinical efÞcacy data in AR patients come from a small, early-phase, single-dose studies involving approximately 50 patients.

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Asthma

ETIOLOGY AND PATHOPHYSIOLOGY Disease Definition In 1995, the Global Initiative for Asthma (GINA)—a collaborative effort by the World Health Organization (WHO), the U.S. National Institutes of Health (NIH), and the National Heart, Lung and Blood Institute (NHLBI)—published the following comprehensive deÞnition of asthma: Asthma is a chronic inßammatory disorder of the airways in which many cells and cellular elements play a role—in particular, mast cells, eosinophils, and T lymphocytes. In susceptible individuals, this inßammation causes recurrent episodes of wheezing, breathlessness, chest tightness, and cough, particularly at night and/or in the early morning. These symptoms are usually associated with widespread but variable airßow obstruction that is at least partly reversible either spontaneously or with treatment. The inßammation also causes an associated increase in the airway hyperresponsiveness to a variety of stimuli (Pearce N, 1998). Currently, asthma is understood to be a disease that can occur, regress, and recur at any age. Because knowledge of asthma pathophysiology and treatment approaches changes frequently (Bousquet J, 2000), diagnosis and classiÞcation practices vary (Pearce N, 1998). Etiology The etiology of asthma is widely thought to be multifactorial, being inßuenced by both environmental and genetic mechanisms. The majority of asthma cases Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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ETIOLOGY AND PATHOPHYSIOLOGY

437

arise when a person becomes hypersensitive to allergens in the general environment, the home, or the workplace. Moreover, 66–75% of asthma patients are believed to be atopic—that is, they have a genetically predetermined sensitivity to such environmental antigens (McFadden ER, 1992). Other asthma attack triggers include exercise, drug exposure, and even lying down (in the case of nocturnal asthma). Table 1 summarizes the various types of asthma. From the 1950s through the 1990s, asthma prevalence increased throughout the world, most notably in industrialized countries. The origins of this trend remain controversial, but considerable attention has been directed toward understanding the environmental and genetic factors that contribute to the development of asthma. Environmental Risk Factors. Asthma is often classiÞed as either extrinsic or intrinsic. Extrinsic asthma—considered to be synonymous with allergic asthma—may be caused by stimuli such as airborne allergens, animal dander, speciÞc food products, and perhaps most commonly, house dust or mites. This form of asthma is much more common in children than in adults, and disease onset typically occurs before 40 years of age. Intrinsic asthma has long been thought to arise from exposure to nonspeciÞc irritants such as tobacco smoke, chemical fumes, and pollution or from other pathological conditions, such as viral infection or gastroesophageal reßux, that damage the airways. However, the possibility that intrinsic asthma is a response to undetected allergens, either environmental or autoimmune, has not been ruled out. Stimuli known to trigger asthma—whether the extrinsic or intrinsic form—can be categorized as speciÞc antigen factors, which cause inßammation, or as nonspeciÞc antigen factors, which exacerbate bronchial hyperresponsiveness (BHR). Examples of speciÞc antigens are pollen, dust mites, animal dander, fungal spores, feathers, detergents, and metals. NonspeciÞc antigens include infections, smoking, sulfur dioxide, airborne particulates, reduced ventilation, and select foods or preservatives. Two major theories have been proposed to explain the roles played by environmental factors in the development of asthma: the urbanization model and the allergic sensitization model. Urbanization Model. Some researchers have suggested that the increase in asthma prevalence, morbidity, and mortality is the result of increased exposure to allergens in the modern indoor environment. In fact, indoor allergen exposure is recognized as the most important risk factor for asthma in children. The severity of a particular asthma case is also known to reßect the degree of allergen exposure. During the past 70–80 years, the concentration of allergens found in the home has increased because the average rate of air exchange has fallen by one-third. Factors contributing to reduced air exchange include the installation of double-glazed windows, the switch from open Þres to central heating, and the blocking of chimneys.

438

ASTHMA

TABLE 1. Types of Asthma Asthma Type Allergic asthma

Drug-induced asthma

Exercise-induced asthma

Near-fatal and hyperacute asthma

Nocturnal asthma

Occupational asthma

Comments Also known as extrinsic asthma, this form of the disease is thought to afflict people who are genetically predisposed to developing hypersensitivity to environmental allergens. It accounts for most childhood asthma cases and a growing proportion of adult cases. It is theorized that sensitization to specific allergens in childhood may lead to a chronic form of the disease. An estimated one in ten cases of adult-onset asthma is caused by sensitivity to aspirin or other cyclooxygenase inhibitors. No clear link between this phenomenon and immune system processes or other biochemical pathways has been established. Classic symptoms of aspirin-induced asthma are rhinitis and flushing, followed by exacerbation manifested by severe bronchoconstriction. This form of asthma occurs with exercise in asthmatics whose disease is poorly controlled. In a subset of asthmatics (particularly children), exercise is the sole cause of asthma attacks. When exercise is the initiating factor, symptoms usually become apparent after termination of the physical activity. Exacerbations brought on by exercise are similar to those of regular asthma attacks, and spontaneous recovery occurs, usually within an hour. The etiology of exercise-induced asthma is not fully understood. It appears that the principal factors behind such attacks are a greater volume of air passing through the bronchioles and the cooler temperature of this air. The effect of increased venous congestion on the airways may also contribute to this type of asthma. Some data suggest that patients with near-fatal and hyperacute asthma exhibit different histology and pathophysiology from other asthmatic patients. Near-fatal cases have a large proportion of neutrophils and few eosinophils, suggesting a different pathogenesis from other asthmatics. Furthermore, the central airways of fatal cases exhibit greater amounts of smooth muscle and submucosal glands and may be more reactive to bronchoconstrictive stimuli. Nocturnal asthma involves increased frequency of asthma symptoms at night, caused by greater bronchial hyperresponsiveness and inflammation. The heightened asthma symptoms may be partly attributable to passive venous congestion associated with lying down. Nocturnal asthma may be a continuation of daytime symptoms and an indication that daytime asthma control is inadequate. This type is caused by prolonged exposure and sensitization to chemicals used in the workplace. Symptoms typically disappear if the chemical stimulant is removed within six months of initial exposure; longer-term exposure often causes the development of persistent asthma.

Source: Based on MacKay IR, 2001; Staton GW, 2002.

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Allergic Sensitization Model. A second theory suggests that a person’s propensity to develop asthma may be increased by exposure to speciÞc allergens at some time between birth and the Þrst two to three years of life. As a result, people develop sensitivities to the allergens typically found in the region where they lived early in life. For example, a person born in a mountainous area would more likely become sensitized to grass pollen, whereas someone living in a densely populated area would have a greater chance of developing sensitivity to dust mites and pets. On the other hand, some researchers believe that more frequent exposure to childhood infections (e.g., viruses) reduces the likelihood of developing asthma. Early exposure to a wide variety of infectious agents is thought to reduce the odds that the immune system will become sensitized to a particular allergen (Holt PG, 1999). For example, the practice of childhood immunizations reduces the exposure to infections and thus increases the likelihood of developing asthma. Conversely, the more siblings a child has, the greater the child’s exposure to viruses and the less likely that he or she will develop asthma. Moreover, a younger sibling is less likely than a Þrst-born child to develop asthma because the younger sibling has greater exposure to viruses earlier in life. Scientists hypothesize that the immune system of a newborn is skewed toward the TH2 immune response (i.e., antibody response) and therefore needs environmental stimuli (such as those just described) that enhance the TH1-cell-mediated immune response to achieve balance between TH1- and TH2-mediated immune responses (Busse WW, 2001). Therefore, practices that reduce exposure to childhood infections also reduce the TH1 immune response and create an immune system imbalance that researchers believe may promote asthma in children. The immune response is discussed further in the “Pathophysiology” section. Genetic Risk Factors. People with a family history of asthma have a slightly increased relative risk of developing the disease, and a greater concordance of asthma exists in monozygotic twins than in dizygotic twins (Clarke JR, 2000). The genetics of asthma are complex, involving different subsets of genes in different patients. Pointing to the complex pathways involved in asthma, researchers emphasize that all the genes involved in the etiology of this disease have not yet been identiÞed. Pathophysiology. Multiple pathological events occurring in the lungs produce the clinical manifestations of asthma. Whatever the type of asthma, however, the disease is always characterized by pulmonary inßammation leading to bronchial hyperreactivity (i.e., increased bronchial smooth-muscle contraction and bronchospasm) and reduced lung function. This section reviews the basic pathological mechanisms of asthma, with particular emphasis on allergic asthma, which has been the subject of most research in this Þeld. Inflammation Allergen-Induced Inflammation. The immune response begins when dendritic cells in the lungs bind, process, and present allergen, as illustrated in Figure 1.

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FIGURE 1. The allergic cascade.

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Although the bronchial epithelium normally serves as a protective barrier against inhaled allergens, chronic inßammation renders the bronchial epithelium of asthmatics unusually permeable to certain allergens, allowing them access to the subepithelial dendritic cells. Bronchial epithelial cells in asthmatics have been shown to secrete granulocyte-macrophage colony-stimulating factor (GM-CSF), which enhances the proliferation and antigen-presenting capabilities of dendritic cells. The ultimate result of these aberrations in the bronchial epithelium is increased presentation of the allergen to hypersensitivity-mediating T cells and B cells. Upon initial exposure to an allergen, a person’s CD4+ helper T cells begin to differentiate into TH1 or TH2 cells. TH1 lymphocytes predominantly secrete the cytokines interleukin-2 (IL-2), interferon-gamma (IFN-γ ), and IL-12; TH2 cells are programmed to release a different class of cytokines: IL-3, IL-4, IL-9, IL-10, IL-13, and GM-CSF. This TH2 cytokine cascade ultimately leads to the antibody-driven allergic inßammatory reaction that characterizes the asthmatic phenotype. Many researchers believe that the initiation and evolution of asthma are caused by an imbalance in the immune response that favors the development of TH2 cells. Neurogenic Inflammation. The nervous system itself can trigger an inßammatory response known as neurogenic inßammation. The neurogenic response ampliÞes ongoing inßammation and may be responsible for the chronic nature of asthma. In neurogenic inßammation, the release of growth factors stimulates sensory nerve endings. When sensitized by mediators such as bradykinin, thromboxane, prostaglandin D2 (PGD2), IL-1, tumor necrosis factor-alpha (TNF-α), and histamine, the nerve endings modulate the release of inßammatory neuropeptides. These neuropeptides may include neurokinin a, calcitonin-gene-related peptide, and substance P. Neurogenic inßammation may be caused by the reduced production of enzymes that degrade neuropeptides, which allows neuropeptide levels to increase unchecked. Early-Phase Asthmatic Reactions Immunoglobulin E. Two particularly important cytokines in the TH2 response are IL-4 and IL-13, both of which act on B cells and facilitate antibody production. Upon activation by allergen, TH2-type T cells secrete IL-4 and IL-13, which act as signals to induce the isotype switching (the mechanism by which allergenspeciÞc immunoglobulin E [IgE] antibodies are developed) and the production of allergen-speciÞc IgE by B lymphocytes (Figure 1). Once synthesized and released into the circulation, secreted IgE binds to allergen. The resulting complexes, in turn, bind to certain high-afÞnity IgE receptors (FcεRI) on mast cells in tissues or peripheral blood basophils. In addition, the allergen-IgE complexes bind to low-afÞnity receptors (FcεRII) on the surfaces of lymphocytes, eosinophils, platelets, and macrophages, but the precise effect of this interaction remains unclear.

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The Role of Mast Cells. Mast cells activated by IgE release histamine and the leukotrienes LTC4, LTD4, and LTE4, as shown in Figure 1. These potent smooth-muscle constrictors cause immediate bronchospasm and vasoconstriction of bronchial blood vessels. Leukotrienes also increase blood vessel permeability, leading to edema and hypersecretion of mucus into the airways. In addition, platelet-activating factor (PAF) released by mast cells may increase local inßammation. These effects constitute the initial early-phase asthmatic reaction, which is characterized by wheezing, coughing, difÞculty breathing, and production of sputum. Typically, the early-phase reaction resolves within one hour. Late-Phase Asthmatic Reactions. Over time, mast cells (along with macrophages and epithelial cells) release mediators (cytokines and chemokines) that attract more inßammatory cells to the lungs, leading to a late-phase reaction that occurs within four to six hours of the early-phase reaction, as depicted in Figure 1. The cells predominantly recruited during this phase include eosinophils, basophils, neutrophils, and certain lymphocytes, predominantly CD4+ TH2 cells. The Role of Eosinophils. Eosinophils have been strongly implicated in the pathology of asthma. One of the most notable cytokines secreted by activated mast cells (and TH2 cells) is IL-5, which stimulates the differentiation of eosinophils from bone marrow precursors and allows these cells to inÞltrate the lung (Figure 1). With the assistance of cell adhesion molecules (CAMs), eosinophils adhere to blood vessels in the lung and extravasate (i.e., pass between the endothelial cells lining blood vessel walls) into lung tissue. This process is enhanced by the increased level of CAMs expressed by microvascular endothelial cells. The principal CAMs expressed by endothelial cells include intercellular cell adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). Various cytokines—but particularly IL-1 and TNF-α —induce the expression of these CAMs. VCAM-1 binds to the cell-surface receptor of verylate-activation-4 integrin (VLA-4; also known as alpha1beta2 integrin) and is thought to reduce the threshold for eosinophil activation by cytokine mediators. Eosinophil adhesion to ICAM-1 is believed to trigger transendothelial migration of the eosinophil into the lung. Once activated in the lung, the eosinophils produce mediators such as PAF, TH2 cytokines, leukotrienes, and toxic granule proteins. Together with oxygenderived radicals, these mediators cause signiÞcant deterioration of epithelial integrity and may lead to epithelial cell death. The loss of these cells is associated with reduced control of smooth-muscle relaxation, resulting in a tendency to increased bronchoconstriction and hyperreactivity. Abnormalities in automatic neural control of airway tone may also occur (discussed in the “Airway Remodeling” section). Chemokines also activate and recruit eosinophils into the airway. These molecules include a broad and diverse group of soluble proteins; the most important chemokines present in allergic inßammation during acute-stage asthma are macrophage inßammatory protein-1α (MIP-1α), RANTES (regulated upon activation, normal T cells expressed and secreted), and monocyte chemotactic protein

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(MCP). Eotaxin is another chemokine thought to be necessary for eosinophil accumulation during the chronic stages of the asthma response. Epithelial Injury. Researchers note that asthma patients routinely show damage to the bronchial epithelium. This injury may range from a minor loss of ciliated cells to complete denudation of the epithelium. Epithelial damage contributes to increased sensitivity by allowing inhaled allergens and irritants to permeate the endothelium more rapidly. In addition to leaving subepithelial layers unprotected, epithelial breakdown eliminates various protective anti-inßammatory mediators and smooth-muscle relaxants. Moreover, the detached epithelial cells collect in bronchoalveolar ßuid. The epithelial degeneration and shedding are associated with physiological effects that aggravate and prolong asthma attacks by reducing mucociliary clearance (resulting in mucus buildup and airßow obstruction) and increasing exposure of submucosal cells and nerve endings to allergens. Disruption of the epithelium also reduces the clearance of inßammatory substances and respiratory secretions, in turn exacerbating the overproduction of mucus in the lungs. Furthermore, inßammatory mediators can trigger overproduction of mucus by submucosal glands (goblet epithelial cells), leading to the formation of potentially fatal plugs in the airways. Airway Remodeling. Persistence of the inßammatory response leads to the development of chronic inßammation, the third inßammatory phase of asthma. Over a period of several months to years, eosinophils continue to inÞltrate and release their cytoplasmic granules. Activated CD4+ T lymphocytes, macrophages, and mast cells in various states of degranulation are present as well. A prominent feature of chronic inßammation is the proliferation of Þbroblasts, with increased collagen secretion causing thickening of the airway walls. Increases in submucosal tissue, the adventitia, and smooth muscle are also apparent, followed by bronchial Þbrosis and scarring. Airway obstruction in asthma is considered reversible, but over time, chronic inßammation permanently reduces airway integrity and responsiveness. Smoothmuscle hypertrophy occurs in asthmatics who have even mild disease of long duration. In some patients, this condition results in irreversible airway obstruction and permanent reduction in baseline lung function, as demonstrated in long-term studies that followed asthmatics and age-matched, nonsmoking controls over a 15-year period (Lange P, 1998). Furthermore, observations in children with asthma suggest that preventing the progressive loss of lung function in childhood may require recognition and treatment of the disease during the Þrst Þve years of life (Martinez FD, 1995). A variety of mechanisms may contribute to the airway remodeling process, including endothelin-1 (a peptide released from inßammatory cells), growth factors, and mediators such as histamine. The roles that inßammatory cells (e.g., eosinophils, TH2 cells, mast cells) may play in causing structural airway changes are under investigation. A greater understanding of the pathophysiology and course of the remodeling process will aid researchers’ efforts to devise novel

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agents and therapeutic strategies that moderate the underlying chronic inßammatory response involved in asthma. CURRENT THERAPIES Current therapies used to manage asthma include prophylactic medications that treat the ongoing inßammation and bronchoconstriction that characterize asthma and rescue medications that relieve acute symptoms. Current treatment paradigms employ anti-inßammatory agents—most notably, inhaled corticosteroids—as Þrst-line prophylactic therapy for persistent asthma. For many patients with persistent asthma, long-acting bronchodilators are increasingly used in combination with prophylactic therapies. The treatment of asthma relies heavily on inhaled therapies that raise unique issues with regard to drug delivery and compliance for patients and prescribers. Most patients favor oral dosing over inhaled dosing. Compliance with inhaled drugs is generally low, but novel delivery devices are improving patient compliance. Combination inhalers deliver two drugs simultaneously, thereby simplifying dosing. User-friendly devices such as dry-powder inhalers and breath-operated aerosol inhalers have also improved patient compliance. Inhaler devices present unique challenges to drug developers. Countries that agreed to the terms of the Montreal Protocol on Substances that Deplete the Ozone Layer must eliminate chloroßuorocarbon (CFC) propellants from medical devices once suitable replacements become available. SigniÞcant progress has been made on this front, particularly in Europe and Japan, where CFC-free versions of most leading inhaled drugs are now available either as hydroßuoroalkane (HFA) formulations, which include an environmentally acceptable propellant, or as DPI formulations, which do not require a propellant. In the United States, the leading maintenance therapies are now available in HFA and/or DPI formulations, and rescue medications will make the transition in 2009. Table 2 summarizes the leading therapies available to treat asthma. Inhaled Corticosteroids Overview. Corticosteroids are the most effective pharmacotherapy for controlling the underlying inßammation of asthma. When administered continuously as prophylactic therapy, they improve lung function and reduce the frequency and severity of asthma exacerbations. Inhalation is the preferred and most common route of administration for these agents, although oral (systemic) formulations are sometimes required to provide more-potent effects and to control symptoms during severe exacerbations. Current treatment guidelines published by the Global Initiative for Asthma (GINA), the National Heart, Lung and Blood Institute (NHLBI), and the British Thoracic Society (BTS) recommend inhaled corticosteroids as Þrst-line therapy for the management of persistent asthma. In addition to alleviating the symptoms of the disease and the associated physiological abnormalities, regular use

TABLE 2. Current Therapies Used for Asthma Agent

Company/Brand

Inhaled corticosteroids Budesonide

AstraZeneca’s Pulmicort

Fluticasone propionate

GlaxoSmithKline’s Flovent/Flixotide

Mometasone Schering-Plough’s furoate Asmanex Long-acting beta2 agonists Salmeterol GlaxoSmithKline’s Serevent

Formoterol

Yamanouchi/Novartis’s Foradil, AstraZeneca’s Oxis

445

Corticosteroid/beta2 agonist combinations Fluticasone/ GlaxoSmithKline’s salmeterol Advair/Seretide Formoterol/ AstraZeneca’s Symbicort budesonide Leukotriene antagonists Montelukast Merck’s Singulair Anti-immunoglobulin E agents Omalizumab Genentech/Novartis/ Sankyo/Tanox Biosystems’ Xolair

[Daily] Dose DPI (Turbuhaler): 100, 200, 400 µg/inhalation; MDI: 50, 200 µg/inhalation—total 400–2400 µg, divided bid-qid. MDI: 88–880 µg, bid; highest dose is 880 µg, bid. DPI (Rotadisk): 100–1,000 µg, bid; highest dose is 1,000 µg, bid. DPI (Twisthaler):200–400 µg, qd.

Availability US, F, G, I, S, UK, J

US, F, G, I, S, UK, J

G, UK

MDI: 42 µg (2 inhalations), bid (morning and evening); for exercise-induced asthma: 42 µg, 30–60 min. before exercise. Diskus DPI: 50 µg (1 inhalation), bid (morning and evening); for exercise-induced asthma: 50 µg (1 inhalation), 30–60 min. before exercise. MDI: 12–24 µg, bid.

US, F, G, I, S, UK, J

DPI (Diskus): salmeterol/fluticasone: 50 µg/100 µg, 50 µg/250 µg, or 50 µg/500 µg, bid. DPI (Turbuhaler): budesonide/formoterol: 1–2 inhalations of 80 µg/4.5 µg or 160 µg/4.5 µg, bid.

US, F, G, I, S, UK

10 mg (oral tablet) qd (in the evening)

US, F, G, I, S, UK, J

SC injection: 150 mg to 375 mg administered every 2 to 4 weeks. Dose determined by total serum IgE level measured prior to administration and body weight.

US

US, F, G, I, S, UK, J

F, G, I, S, UK

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TABLE 2. (continued) Agent

Company/Brand

Short-acting beta2 agonists Albuterol Schering-Plough’s Proventil, GlaxoSmithKline’s Ventolin, generics Levalbuterol Sepracor’s Xopenex Oral corticosteroids Prednisone

Methylxanthines Theophylline (extended release) Cromolyns Cromolyn sodium

Anticholinergics Ipratropium bromide Ipratropium/ albuterol

[Daily] Dose

Availability

MDI: 180 µg (2 inhalations), q4–6h, PRN. Nebulized solution: 2.5 mg, tid-qid. Higher doses not recommended.

US, F, G, I, S, UK, J

Oral nebulization: initially 0.63 mg (1 inhalation), tid; up to 1.25 mg (1 inhalation), tid.

US

Multisource

Chronic asthma: ≤40 mg (oral tablets or liquid), every other day. Acute asthma: ≤50 mg/day for 5–14 days.

US, F, G, I, S, UK, J

AstraZeneca’s Theo-Dur, 3M’s Theolair, generics

Extended-release oral capsules: 400–1,600 mg/day, individualized to maintain peak serum theophylline concentration in the 10–20 µg/mL range. Initially dosed bid, often switched to qd.

US, F, G, I, S, UK, J

King Pharmaceuticals/ Sanofi-Aventis’ Intal

MDI: 1,600 µg (2 inhalations), qid. Maximum daily dose: 12.8 mg (4 inhalations), qid. For prevention, two inhalations, 10–60 minutes before exposure to inducing agent.

US, F, G, S, UK, J

Boehringer Ingelheim’s Atrovent

MDI: 36 µg (2 inhalations), qid; maximum dose: 12 inhalations. Oral nebulization: 500 µg (1 unit-dose vial), tid-qid. MDI: ipratropium 36 µg/albuterol 206 µg (two aerosol inhalations), qid; maximum dose: 12 inhalations. Oral nebulization: ipratropium 200 µg/albuterol 2.5 mg (1 unit-dose vial), tid-qid.

US, F, G, I, S, UK, J

Boehringer Ingelheim’s Combivent, Valeas’s Breva

US, F, I, S, UK

bid = Twice daily; DPI = Dry powder inhaler; MDI = Metered-dose inhaler; PRN = Pro re nata, or use as needed; qd = Once daily; bid = Two times daily; tid = Three times daily; qid = Four times daily; q4h = every four hours; SC = Subcutaneous. US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan.

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of inhaled corticosteroids—even at low doses—appears to substantially reduce asthma-related hospitalizations and deaths. In an epidemiological literature survey, researchers reported that eight independent studies strongly suggested that inhaled corticosteroids, taken regularly, can reduce the number of hospitalizations for asthma by as much as 80% (Suissa S, 2001). The evidence linking corticosteroid use to reductions in asthma-related mortality is less clear, but the same researchers supported recent Þndings that demonstrated a signiÞcant reduction in fatalities with the use of these agents. The following inhaled corticosteroids are listed in order of decreasing potency, based on in vitro studies (Johnson M, 1998): • • • •

Fluticasone propionate (GlaxoSmithKline’s [GSK’s] Flovent/Flixotide). Budesonide (AstraZeneca’s Pulmicort). Beclomethasone dipropionate (GSK’s Beclovent/Beclotide, ScheringPlough’s Vanceril/Vancenase). Flunisolide (Forest Laboratories’ Aerobid/Aerobid-M) and triamcinolone acetonide (Kos’ Azmacort)—approximately equipotent.

Researchers caution, however, that in vitro potency may not accurately reproduce clinical potency. For the vast majority of patients (who have mildto-moderate asthma), the effects of increasing potency may be minimal. Although corticosteroids are by far the most effective compounds for controlling asthma, they are associated with several potentially serious side effects—particularly when used at high doses for extended periods. In adult patients, the adverse effect of greatest concern is bone mineral loss; long-term treatment with oral corticosteroids increases the risk of bone loss and osteoporotic fracture (van Staa TP, 2000). However, because inhaled corticosteroids have been available for considerably less time than oral corticosteroids, similar long-term data are not yet available on patients treated with inhaled formulations. To date, treatment guidelines created by the GINA do not recommend prophylactic treatment for osteoporosis in patients treated with inhaled corticosteroids (GINA, 2004). The medical community has raised concerns about the possibility that stunted growth in the pediatric asthma population may result from treatment with inhaled corticosteroids, but a consensus appears to be emerging that patients treated with inhaled corticosteroids as children do reach full adult height (GINA, 2004). Recent clinical studies examining this issue have consistently found a lack of long-term effect on adult height, although the Childhood Asthma Management Program (CAMP) Research Group found slightly impaired growth during the Þrst year of treatment only (Agertoft L, 2000; CAMP Research Group, 2000). DeÞning the risk-beneÞt ratio in children is particularly important because early initiation of corticosteroid therapy in these patients may prevent irreversible remodeling of bronchial tissue. As a precaution, the FDA recommends that physicians monitor the growth of corticosteroid-treated children every three to six months with a sensitive device (e.g., a stadiometer).

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Although inhaled corticosteroids cause fewer serious systemic side effects than oral formulations, systemic side effects remain a problem because only a portion of the inhaled dose reaches the lungs. The remaining medication stays in the oropharynx or reaches the gastrointestinal (GI) tract, eventually being absorbed into the systemic circulation. Inhaled corticosteroids are also associated with local adverse effects, including oral candidiasis, hoarseness, dysphonia, and throat irritation. This section focuses on the inhaled corticosteroids budesonide (AstraZeneca’s Pulmicort) and ßuticasone propionate (GSK’s Flovent/Flixotide) because they are the top-selling inhaled corticosteroids across the major markets under study. It also discusses the most recently launched inhaled corticosteroid, mometasone furoate (Schering-Plough’s Asmanex). Additional agents in this class include beclomethasone dipropionate (GSK’s Beclovent/Beclotide, Schering-Plough’s Vanceril/Vancenase); ßunisolide (Forest Laboratories’ Aerobid/Aerobid-M); and triamcinolone acetonide (Kos’s Azmacort). Mechanism of Action. All corticosteroids exert their therapeutic effect by binding to intracellular glucocorticoid receptors, which are found in all cells and are abundant in the epithelium of bronchi. In this way, corticosteroids alter intracellular activities and reduce the transcription of genes that encode proteins involved in the inßammatory response, including the cytokines IL-1, -3, -4, -5, -6, and -8; tumor necrosis factor-alpha (TNF-α); granulocyte-macrophage colony-stimulating factor (GM-CSF); and RANTES (regulated upon activation, normal T cells expressed and secreted). By lowering the levels of these inßammation mediators, corticosteroid therapy reduces the activity of lymphocytes, eosinophils, macrophages, and mast cells in mucosal ßuids. In addition, corticosteroids increase the transcription of genes responsible for producing beta2 adrenoreceptors and lipocortin 1, a protein that inhibits phospholipase A2 (a proinßammatory enzyme). Other key mechanisms by which corticosteroids help alleviate the symptoms of asthma include inhibition of T-cell activation (and subsequent cytokine release), promotion of apoptosis of eosinophils, inhibition of nitric oxide (NO) synthase, and reduction in mucus secretion by submucosal gland cells. Budesonide. Budesonide (AstraZeneca’s Pulmicort) (Figure 2) is a secondgeneration corticosteroid. The drug is marketed in a dry-powder inhaler (Pulmicort Turbuhaler), approved for once-daily use in adults and in children aged six years or older with mild-to-moderate asthma, and in nebulizer form (Pulmicort Respules), approved for the treatment of children aged 12 months to 8 years. Although this drug has been available in Europe since the early 1980s, it was not introduced in the United States until June 1997. Budesonide has been available in Japan since December 2001. Like other corticosteroids, budesonide binds to the glucocorticoid receptor and suppresses the production of proinßammatory cytokines. Although budesonide has a shorter plasma half-life than other inhaled corticosteroids, its residence in the lung is extended by the formation of lipid conjugates with esterases

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OH O HO

O

H

O

O FIGURE 2. Structure of budesonide.

in the lung. Budesonide has greater oral bioavailability than ßuticasone propionate but lower oral bioavailability than older inhaled corticosteroids such as beclomethasone. The efÞcacy and safety of budesonide delivered by dry-powder inhaler (DPI) were assessed in a 12-week, double-blind, multicenter, parallel-group study of 473 adult patients with moderate-to-severe chronic asthma (Busse WW, 1998). Patients were treated with placebo or with 200 µg, 400 µg, 800 µg, or 1,600 µg total daily budesonide. The primary end points measured in the study were mean change from baseline forced expiratory volume at one second (FEV1 ) and morning peak expiratory ßow rate (PEFR). Patients treated with budesonide showed signiÞcant increases in both measures; patients treated with placebo showed declines in both measures. The mean change in morning PEFR was 12, 22, 27, and 30 L/min in the 200 µg, 400 µg, 800 µg, and 1,600 µg dose groups, respectively, compared with −27 L/min in the placebo group. Increases in FEV1 were approximately 0.14, 0.23, 0.23, and 0.28 L in the 200 µg, 400 µg, 800 µg, and 1,600 µg dose groups, respectively, compared with a 0.22 L decline in the placebo-treated group. Similar results were obtained in a study of 309 patients with mild-to-moderate asthma that was designed to test maintenance of stable asthma with lower doses of corticosteroid (McFadden ER, 1999). In this randomized, double-blind study, patients were treated with placebo or with 200 µg or 400 µg of budesonide delivered by dry-powder inhaler (Pulmicort Turbuhaler) once daily. Patients treated with 200 µg or 400 µg of budesonide showed FEV1 increases of 0.10 and 0.11 L, respectively, after 18 weeks of treatment, while patients treated with placebo showed a 0.09 L decline in FEV1 . Greater increases in FEV1 were observed in patients not previously treated with corticosteroids. The most commonly reported adverse effects were respiratory infection, headache, and bronchospasm; several cases of oral candidiasis were also reported in both the placebo and treatment groups. Fluticasone Propionate. Launched in the early 1990s, ßuticasone propionate (GSK’s Flovent/Flixotide) (Figure 3) quickly became one of the most widely used inhaled corticosteroids. It is sold in two DPIs (the Diskus inhaler and the Rotadisk inhaler) and in a metered-dose inhaler (MDI). The Diskus and Rotadisk

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F S H3C

HO H3C

CH3

O O CH3

H F

O

H

O F

FIGURE 3. Structure of fluticasone propionate.

formulations are approved for the treatment of asthma in patients four years of age or older; the MDI formulation is approved only for patients aged 12 years or older. It is available in all seven market under study. Like all corticosteroids, ßuticasone propionate binds to the glucocorticoid receptor and reduces the transcription of genes that encode proinßammatory cytokines. Fluticasone propionate has the highest relative afÞnity for the glucocorticoid receptor of all currently marketed inhaled corticosteroids—approximately twice that of budesonide (Allen DB, 2003)—and its long half-life extends residence time in the lung. Fluticasone propionate also has low oral bioavailability, an attribute associated with reduced risk of systemic exposure and adverse effects. This agent has demonstrated better efÞcacy than budesonide in the management of severe asthma (Georgitis JW, 1999). In addition, some studies suggest that a once-daily, higher-dose ßuticasone administration is efÞcacious in the treatment of persistent asthma (Berger WE, 2002). The efÞcacy of inhaled ßuticasone propionate powder in treating moderate persistent asthma was examined in a randomized, double-blind, placebo-controlled trial involving 342 adolescent and adult patients (Pearlman DS, 1997). Patients were treated with placebo or with 50 µg, 100 µg, or 250 µg of ßuticasone propionate delivered by two inhalations daily from the Diskhaler device. Patients in all three treatment groups experienced improvements in FEV1 of 17–18% above baseline, while patients in the placebo group experienced a decline in FEV1 . Patients treated with ßuticasone also demonstrated improved PEFR, less need for rescue medication, and fewer nighttime awakenings, compared with their placebo-treated counterparts. No serious adverse events occurred; the most common adverse events considered likely to be treatment-related were dysphonia, pharyngitis, headache, and oropharyngeal candidiasis, each occurring in no more than 6% of ßuticasone-treated patients. Of these events, only headache (3%) was reported in the placebo group. Similar results were obtained in studies of patients with mild-to-moderate persistent asthma (Sheffer AL, 1996; Chervinsky P, 1994). Mometasone Furoate. Mometasone furoate (Figure 4) is the Þrst inhaled corticosteroid approved for once-daily use and has been available since 1997 in most

CURRENT THERAPIES

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FIGURE 4. Structure of mometasone furoate.

markets as an aqueous nasal spray (Schering-Plough’s Asmanex) for treating allergic rhinitis. In January 2003, Schering-Plough announced that a dry-powderinhaler (DPI) formulation of mometasone, called Asmanex Twisthaler, was being launched in the United Kingdom for mild, moderate, and severe persistent asthma in patients aged 12 years or older. That February, it became available in Germany. The DPI formulation has been preregistered in the United States since 1998, but the drug did not receive FDA approval until March 2005. Like other inhaled corticosteroids, mometasone furoate binds to the glucocorticoid receptor and suppresses the production of cytokines involved in the inßammatory response. Mometasone is a potent inhibitor of anti-inßammatory cytokines but has low systemic availability (Affrime MB, 2000[a]). In a 12-week, randomized, placebo-controlled Phase III trial, 236 patients (aged 12 or older) with mild-to-moderate asthma were treated with once-daily DPI mometasone (200 or 400 µg) or with placebo (Nayak AS, 2000). Spirometric measurements indicated that patients in both the 200 µg and 400 µg mometasone-treated groups experienced a signiÞcant percentage increase in FEV 1 from baseline (14.8% and 14.2%, respectively) compared with the placebo group (2.5%). The percentage of subjects considered to be improved (based on an assessment of symptoms) by the end of the trial was higher in the 200 µg and 400 µg mometasone groups (64% and 66%, respectively) than in the placebo group (50%). Symptoms worsened in 5% of patients receiving 200 µg mometasone, in 8% receiving 400 µg mometasone, and in 21% of the patients receiving placebo. Treatment-related adverse events were distributed equally across the three groups; the most commonly reported events were oral candidiasis, headache, and pharyngitis. One patient receiving placebo, three patients receiving 200 µg mometasone, and one patient receiving 400 µg mometasone experienced severe adverse events. Mometasone may also beneÞt patients with severe persistent asthma by reducing the need for oral prednisone. A 12-week, double-blind, placebo-controlled trial compared the efÞcacy of 400 µg and 800 µg DPI mometasone administered twice daily in 132 severely ill asthmatics who required oral corticosteroid treatment (Fish JE, 2000). Results showed that treatment with 400 µg or 800 µg mometasone reduced daily oral corticosteroid requirements by 46% and 24%,

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respectively, compared with an increase of 164% in the placebo group. Furthermore, oral corticosteroids were completely eliminated in 40% and 37% of patients in the 400 µg and 800 µg treatment groups, respectively; corticosteroids were not eliminated in any patients in the placebo arm. Studies of the safety and tolerability of mometasone reveal that its systemic bioavailability is negligible. Plasma concentrations were below the limit of quantiÞcation (50 pg/mL) and were not detectable in 92% of healthy study participants, suggesting that it is extensively metabolized (Affrime MB, 2000[a]). Urinary cortisol excretion assays are often performed to determine whether a patient receiving corticosteroid therapy is adrenally suppressed, a condition commonly associated with chronic, high-dose corticosteroid therapy. Mometasone has been shown to have a minimal effect on cortisol suppression when delivered at high doses (800–1,200 µg daily total) (Affrime MB, 2000[b]). Although mometasone appears to have a favorable safety proÞle, long-term studies are needed to determine its potential for adverse effects. Long-Acting Beta2 Agonists Overview. Compared with short-acting beta2 agonists, long-acting beta2 agonists tend to have a slower onset of action and demonstrate prolonged receptor occupancy (Table 3). As a result, they are prescribed to provide partial, steadystate bronchodilation and longer-term beneÞcial effects. Formoterol (Yamanouchi/Novartis’ Foradil; AstraZeneca’s Oxis) has a relatively quick onset of action (within 5 minutes) compared with the rest of the agents in this class (salmeterol’s onset of action is between 8 and 20 minutes, for example); this agent may be prescribed for both acute and prophylactic therapy. Recent clinical studies provide support for this practice and suggest that formoterol may offer some advantages over short-acting beta2 agonists (Pauwels RA, 2003; TattersÞeld AE, 2001).

TABLE 3. Onset of Action of Short-Acting and Long-Acting Beta2 Agonists Onset of Action Source (Description)

Short-Acting Beta2 Agonist

Long-Acting Beta2 Agonist

Ullman A, 1992 (experimental model) Bensch G, 2001 (clinical trial) Anderson SD, 1991 (clinical trial of exercise-induced asthma)

Albuterol = 2 minutes

Salmeterol = 8 minutes

Albuterol = Within 5 minutes Albuterol was effective when administered within 30 minutes of exercise.

Formoterol = Within 5 minutes Salmeterol was as effective as albuterol when administered 30 minutes before exercise. It was more effective when administered 2.5, 4.5, and 6.5 hours prior to exercise.

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Ever since long-acting beta2 agonists were introduced into medical practice, some physicians have expressed concern about the potential development of tolerance to therapy. Although some publications suggest that long-term therapy is not associated with increased frequency or severity of attacks, other studies have indicated that partial tolerance to long-acting beta2 agonists develops in children when these agents are used as maintenance treatment (Bisgaard H, 2000). Mechanism of Action. Beta2 adrenergic receptors are found in smooth muscle in the airway and in the lungs, blood vessels, and other tissues throughout the body. In the airway, stimulation of these receptors by beta2 agonists relaxes bronchial smooth muscle and produces bronchodilation. Long-acting beta2 agonists provide protection against bronchoconstriction caused by allergens, exercise, histamine, and methacholine for at least 12 hours. Other effects of long-acting beta2 agonists that have been observed experimentally include the following: • • • •

Blocking of mast-cell release of leukotrienes and histamine in the lungs. Reduced microvascular permeability and mucus production. Increased mucociliary function. Possible inhibition of the activity of phospholipase A2, a key enzyme in the proinßammatory arachidonic acid pathway.

Salmeterol. Originally approved in 1994, salmeterol (GSK’s Serevent) (Figure 5) is indicated for long-term maintenance therapy for asthma and for the prevention of bronchospasm in patients older than 12 years who have demonstrably reversible obstructive airways. Salmeterol is also indicated for the prevention of exercise-induced bronchospasm in patients older than 12. The agent is marketed in both DPI and CFC-containing MDI formulations. An HFA-propelled MDI is in Phase III trials in the United States and preregistered in Europe. Several trials in asthmatics have demonstrated that salmeterol is superior to the short-acting beta2 agonist albuterol (GSK’s Ventolin; Schering-Plough’s Proventil, generics) in improving various outcomes (including asthma-speciÞc quality of life and lung function) and in reducing respiratory symptoms during the day and night. In a 12-week, randomized, double-blind, parallel-group, multicenter trial, 539 adult asthmatics were administered either salmeterol 42 µg twice daily (via MDI) or albuterol 180 µg four times daily (Wenzel SE, 1998). At 12 weeks, salmeterol resulted in signiÞcant changes from baseline quality-of-life (QOL)

OH

H N

HO

O HO

.CH3CO2H FIGURE 5. Structure of (S)-salmeterol.

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scores for asthma symptoms (1.0), emotional function (0.85), and environmental exposure (0.65), compared with albuterol (0.5, 0.4, and 0.45, respectively). Scores are based on QOL measurements over a two-week time period; each factor (including activity limitation, symptoms, and emotional function) is scored between 1 and 7, with higher scores indicating greater QOL. Average increases in PEFR over baseline for the morning and evening were signiÞcantly higher in the salmeterol group (39.3 L/min and 19.5 L/min, respectively) compared with the albuterol group (6.6 L/min and 8.4 L/min, respectively). In addition, average increases in FEV1 at weeks 4, 8, and 12 were signiÞcantly higher in the salmeterol group (0.32 L, 0.38 L, and 0.37 L, respectively) compared with the albuterol group (0.16 L, 0.21 L, and 0.20 L, respectively). Statistical significance was also observed in mean change from baseline of symptom-free day measurements (28.6% and 13.4% for salmeterol and albuterol, respectively). The frequency of adverse events between treatment groups was not signiÞcant, and the percentage of patients with a speciÞc drug-related adverse event was always below 1%. During the Þrst quarter of 2003, the Salmeterol Multicenter Asthma Research Trial (SMART), a safety study, was terminated as a result of a potential association between salmeterol and increased risk of life-threatening asthma episodes or asthma-related deaths. The study compared the effects of 28 weeks of salmeterol (42 µg twice daily) with placebo in patients with asthma. GSK added black box warnings to the package inserts of both salmeterol and the combination therapy Advair (ßuticasone/salmeterol) to alert physicians to the increase in asthma-related deaths that occurred (Pink Sheet; August 18, 2003). When the Þndings of the SMART trial were released, the FDA expressed interest in exploring the possibility of a class-based effect. However, no progress on this front has been reported to date. If investigations demonstrate that the risk is due to a class effect, similar label warnings could appear on formoterol or on formoterolcontaining products, such as Symbicort. The FDA has stated that patients should not stop taking medications containing salmeterol without consulting their physicians and that the beneÞts of treatment with salmeterol in patients with asthma (and chronic obstructive pulmonary disease [COPD]) continue to outweigh the potential risks when used according to product labeling. Formoterol. First marketed by Yamanouchi in Japan, formoterol (Yamanouchi/ Novartis’s Foradil; AstraZeneca’s Oxis) (Figure 6) was launched by Novartis in Europe and the United States in 2001. In the fourth quarter of 2002, one and one-half years after Foradil’s U.S. launch, Novartis licensed the U.S. rights to this agent to Schering-Plough for maintenance treatment of asthma and COPD and for the acute prevention of exercise-induced bronchospasm. In return, Novartis will receive royalties and milestone payments if sales exceed a certain level. Finally, AstraZeneca’s Oxis Turbuhaler has been approved in more than 60 countries and was approved for COPD in the European Union (EU) in December 2002. To evaluate the long-term efÞcacy and safety of formoterol in pediatric asthmatics, a 12-month, multicenter, randomized, double-blind study was conducted in 518 pediatric asthmatics (aged 5–12 years) who were still symptomatic despite

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OH H N

H N

OHC HO

CH3

OCH3

FIGURE 6. Structure of formoterol.

treatment with anti-inßammatory medication. Patients received placebo or formoterol 12 µg or 24 µg twice daily for 12 months (Bensch G, 2002). Morning premedication PEFR (averaged over the 12-month study) for the formoterol 12 µg and 24 µg groups increased 14.5% (+34 L/minute) and 16.3% (+40 L/minute), respectively, compared with an average increase in the placebo group of 8.6% (21 L/minute). Evening PEFR averages in patients treated with formoterol 12 µg and 24 µg increased 11.8% (+29 L/minute) and 13.7% (+35 L/minute), respectively, compared with an average increase of 7.5% (+19 L/minute) in the placebo group. The number of asthma patients who became symptomatic despite rescue treatment with albuterol during the study was not statistically different between the treatment groups, but signiÞcantly more serious asthma exacerbations (requiring hospitalization and/or discontinuation from the study) occurred in patients receiving formoterol, compared with patients receiving placebo (no serious asthma exacerbations occurred in the placebo group). The effectiveness of formoterol as relief medication is under investigation. An open-label study of 18,124 children and adults compared the safety and effectiveness of as-needed formoterol and as-needed albuterol (Pauwels RA, 2003). The primary efÞcacy variable was time to Þrst asthma exacerbation; the primary safety variables were asthma-related serious adverse events and adverse events resulting in participants’ discontinuing study participation. The frequencies of adverse events not related to asthma, asthma-related deaths, and cardiovascularrelated deaths were similar in both treatment groups. The incidence of asthma exacerbations assessed by clinical criteria was lower in the formoterol group. However, because the study evaluated exacerbations based on clinical criteria rather than spirometry and because the study design was not blinded, the possibility of investigator bias cannot be entirely ruled out. While acknowledging the study’s limitations, the investigators suggest that this study supports the prescription of formoterol to asthmatic patients on an as-needed basis. Corticosteroid/Beta2 Agonist Combinations Overview. The combination of inhaled corticosteroids and long-acting beta2 agonists produces superior outcomes compared with increased doses of singleagent inhaled corticosteroids. The combination of these classes of agents into one delivery system offers clinical beneÞts (such as increased patient compliance), as evidenced by the commercial success of marketed combination products. GSK’s Advair (Seretide in Europe) combines the long-acting beta2 agonist salmeterol and the anti-inßammatory corticosteroid ßuticasone. AstraZeneca’s Symbicort combines the beta2 agonist formoterol and the corticosteroid budesonide.

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Mechanism of Action. All corticosteroids exert their therapeutic effect by binding to glucocorticoid receptors, which are found in all cells and are abundant in the epithelium of bronchi. In this way, corticosteroids alter intracellular activities and reduce the transcription of genes that encode proteins involved in the inßammatory response, including the cytokines IL-1, -3, -4, -5, -6, and -8; TNF-α; GM-CSF; and RANTES. By lowering the levels of these inßammation mediators, corticosteroid therapy reduces levels of lymphocytes, eosinophils, macrophages, and mast cells in mucosal ßuids. In addition, corticosteroids increase the transcription of genes responsible for producing beta2 adrenoreceptors and lipocortin 1, a protein that inhibits phospholipase A2, a proinßammatory enzyme. Other key mechanisms by which corticosteroids help alleviate the symptoms of asthma include inhibition of T-cell activation (and subsequent cytokine release), promotion of apoptosis of eosinophils, inhibition of NO synthase, and reduction in mucus secretion by submucosal gland cells. Evidence also suggests these drugs help restore damaged epithelium. Long-acting beta2 agonists act on beta2 adrenergic receptors found in smooth muscle in the airway and in the lungs, blood vessels, and other tissues throughout the body. In the airway, stimulation of these receptors by beta2 agonists relaxes bronchial smooth muscle and produces bronchodilation. Long-acting beta2 agonists provide protection against bronchoconstriction caused by allergens, exercise, histamine, and methacholine for at least 12 hours. Other effects of long-acting beta2 agonists that have been observed experimentally include the following: • • • •

Blocking of mast-cell release of leukotrienes and histamine in the lungs. Reduced microvascular permeability and mucus production. Increased mucociliary function. Possible inhibition of the activity of phospholipase A2, a key enzyme in the proinßammatory arachidonic acid pathway.

In addition to the individual actions of each agent, several lines of evidence suggest interactions between corticosteroids and beta2 agonists. Salmeterol potentiates glucocorticoid receptor responsiveness by enhancing receptor translocation, binding to target sites, and phosphorylation of the receptor (Adcock IM, 2002). Corticosteroids inßuence the activity of beta2 agonists by increasing the number of beta2 adrenergic receptors. Furthermore, corticosteroids may reverse the downregulation of beta2 adrenergic receptors that occurs with long-term administration of long-acting beta2 agonists. Fluticasone/Salmeterol. Fluticasone/salmeterol (GSK’s Advair/Seretide) was launched in 1998 in the European Union and in 2001 in the United States, where it is indicated for twice-daily maintenance treatment of asthma in patients aged 4 years or older. Fluticasone/salmeterol is licensed to Almirall-Prodesfarma and Alter for sales in Spain, and in August 2000, Schwarz Pharma acquired exclusive marketing rights to ßuticasone/salmeterol in Germany. In Japan, GSK submitted a new drug application for the ßuticasone/salmeterol Diskus in April 2004.

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The individual components of ßuticasone/salmeterol provide anti-inßammatory and bronchodilating effects, respectively. Salmeterol stimulates beta2 adrenergic receptors and relaxes smooth muscle; ßuticasone suppresses the production of pro-inßammatory cytokines. Salmeterol is relatively dose-unresponsive at high concentrations; consequently, all available strengths of ßuticasone/salmeterol contain the same dose of salmeterol, but the dose of ßuticasone varies. Clinical studies were conducted to compare the efÞcacy of ßuticasone/ salmeterol in treating asthma with the efÞcacy of its constituent agents given independently. In a randomized, double-blind, placebo-controlled, parallel-group study, 349 asthmatics previously treated with medium doses of inhaled corticosteroids were given either twice-daily salmeterol 50 µg and ßuticasone 250 µg (combined in one inhaler), salmeterol 50 µg, ßuticasone 250 µg, or placebo (Shapiro G, 2000). The primary end point of mean change in FEV1 after 12 weeks was signiÞcantly greater with Advair (0.48 L) than with salmeterol (0.05 L), ßuticasone (0.25 L), or placebo (−0.11 L); these changes in volume represent percentage improvements from baseline of 23%, 4%, 13%, and 5%, respectively. Morning PEFR measurements at trial endpoint were signiÞcantly higher in participants who received combination therapy (53.5 L/min) than in those who were given salmeterol (−11.6 L/min), ßuticasone (15.2 L/min), or placebo (−14 L/min). A signiÞcantly greater number of patients remained in the study from the Advair group (84%) than from the salmeterol group (48%), ßuticasone group (73%), or placebo group (29%). The most common minor adverse events included oral candidiasis (2–4% of patients), headache (2–5%), tremors (2%), dizziness (2%), throat irritation (1–3%), and cough (1–3%). The recent addition of blackbox warnings to salmeterol package inserts, after the occurrence of asthma-related deaths in the SMART study, is likely to contribute to the shift from single inhalers to combination inhalers (Pink Sheet; February 24, 2003). Formoterol/Budesonide. The combination of formoterol and budesonide (AstraZeneca’s Symbicort), delivered by a DPI device (Turbuhaler), is marketed throughout Europe. In September 2005, AstraZeneca Þled a New Drug Application (NDA) for a formulation of formoterol/budesonide delivered by MDI in the United States. Development in Japan has not been reported. The pMDI (pressurized metered-dose inhaler) version of Symbicort is also in development in Europe. The individual components of budesonide/formoterol control inßammation by suppressing the production of proinßammatory cytokines and relax smooth muscle by stimulating beta2 adrenergic receptors. Because formoterol is doseresponsive across a wider range of doses than other long-acting beta2 agonists, physicians can increase or reduce the dose of the drug with the same inhaler by directing patients to take more or fewer puffs in response to variable symptoms. Formoterol/budesonide is approved in Europe for both Þxed-dose and adjustabledosing regimens.

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AstraZeneca is seeking regulatory approval in Europe to market budesonide/ formoterol for both maintenance and rescue therapy, allowing patients to use a single inhaler for maintenance and acute dosing. This strategy would eliminate the need for an additional inhaler containing a short-acting beta2 agonist. Because formoterol has a rapid onset of action, it may be able to provide relief of acute symptoms comparable to that of short-acting beta2 agonists. Therefore, the company is seeking to market the budesonide/formoterol combination inhaler as an agent that can also be administered in response to acute symptoms. In November 2003, AstraZeneca submitted an application to the European Medicines Agency (EMEA) for the new labeling. In November 2004, AstraZeneca withdrew the application and announced that a resubmission with additional clinical data is expected at a later date. Although corticosteroids are the most effective agent for treating moderateto-severe asthma, it is fairly common for patients to remain symptomatic and require additional treatment. To evaluate the efÞcacy of combined formoterol and budesonide, 852 asthmatics (who were being treated with glucocorticoids) were enrolled in a one-year, double-blind, placebo-controlled trial. Twice daily, patients received either 100 µg budesonide/placebo, 100 µg budesonide/12 µg formoterol, 400 µg budesonide/placebo, or 400 µg budesonide/12 µg formoterol (Pauwels RA, 1997). By the addition of formoterol to 100 µg budesonide, rates of severe and mild exacerbations were reduced by 26% and 40%, respectively; by the addition of formoterol to 400 µg budesonide, rates were reduced by 63% and 62%, respectively. The percentage of patients free of exacerbations during the study was 80.8% in the 400 µg budesonide/12 µg formoterol group and 61.4% in the group receiving budesonide alone. All treatment combinations were well tolerated, and the proportion of adverse events within each treatment group was similar. Eleven patients were hospitalized as a result of asthma exacerbation, and 29 patients withdrew because of adverse events. Seven of the withdrawals stemmed from pharmacologically predictable adverse events (headache, tremor, tachycardia, and oral candidiasis). Leukotriene Antagonists Overview. Derived from arachidonic acid, leukotrienes (LTs) are lipid mediators that are released from most of the inßammatory cells present in (or recruited to) the airways. They are logical targets for antagonism because they directly mediate bronchoconstriction and have proinßammatory effects. In addition, the leukotriene antagonists appear to lack the adverse effects on growth, bone mineralization, and the adrenal axis that have been associated with long-term corticosteroid therapy. Montelukast (Merck’s Singulair) is the class leader in the asthma market. Other available oral leukotriene antagonists include pranlukast (Ono’s Onon), zaÞrlukast (AstraZeneca’s Accolate), and zileuton (Critical Therapeutics’s Zyßo). Mechanism of Action. Leukotrienes are derivatives of arachidonic acid, an unsaturated fatty acid produced from membrane phospholipids. Mast cells and

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basophils generate a mixture of leukotrienes, which stimulate the production of airway secretions, enhance the migration of eosinophils, and cause microvascular leakage. Two modes of leukotriene inhibition have been developed: •

•

Leukotriene synthesis inhibitors. The leukotriene synthesis inhibitors include inhibitors of 5-lipoxygenase [5-LO] and 5-LO-activating protein [FLAP], which are components of the 5-LO pathway. This pathway produces several leukotrienes (e.g., leukotriene B4 (LTB4), cysteinyl leukotrienes) that can cause edema, eosinophil migration, and airway secretion—physiological processes that contribute to asthma and its associated airway remodeling (Drazen JM, 1999). Theoretically, inhibitors of this pathway might reduce inßammation due to asthma. LTD4 (leukotriene D4) receptor antagonists. The LTD4 receptor antagonists are also known as cysteinyl leukotriene-1 [cysLT1] receptor antagonists. Cysteinyl leukotrienes exert their biological effect by binding to receptors located on airway smooth muscles (Drazen JM, 1999). Therefore, blocking this interaction with LTD4 receptor antagonists might inhibit the inßammatory effects.

Montelukast. In 1998, montelukast (Merck’s Singulair) (Figure 7) became the Þrst once-daily treatment for asthma and the second LTD4 receptor antagonist (after zaÞrlukast) to win FDA approval in the United States. The agent was approved that same year in Europe; in Japan, it was approved in June 2001 and licensed to Banyu for marketing. It is the Þrst leukotriene receptor antagonist intended for use both in adults and in children as young as 12 months. Compared with inhaled antiasthma medications, montelukast enjoys a higher degree of patient acceptability thanks to its once-daily, oral administration and relative lack of side effects. Leukotriene antagonists were recently included in the GINA guidelines, but their position in current treatment algorithms is somewhat ill-deÞned. The most recent version of the guidelines cautions that discontinuing therapy with inhaled corticosteroids in favor of treatment with leukotriene antagonists could result in worsening asthma (GINA, 2004). The guidelines acknowledge that leukotriene antagonists can provide some beneÞt when used

FIGURE 7. Structure of montelukast sodium.

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in combination with inhaled corticosteroids, but most clinical trial data available to date indicate that long-acting beta2 agonists have superior efÞcacy as add-on therapy. Montelukast is an LTD4 receptor antagonist and exerts its biological effect by binding to receptors located on airway smooth muscles and preventing the binding of LTD4 (Drazen JM, 1999). Montelukast-treated patients show reductions in circulating eosinophils, sputum eosinophils, and exhaled nitric oxide. Montelukast is sometimes prescribed as monotherapy in patients with mild disease who might otherwise receive inhaled corticosteroids. A multicenter, randomized, double-blind, parallel-group, 24-week study compared the long-term effects of low-dose inhaled ßuticasone (88 µg; 2 puffs of 44 µg) with oral montelukast (10 mg/day) (Meltzer EO, 2002). The study involved 522 patients older than 15 years who had persistent asthma. The primary measures of efÞcacy were change from baseline (mean percentage) in morning and premedication FEV1 readings. The changes were signiÞcantly higher in the ßuticasone group (22%) than in the montelukast group (14%). Measurements of asthma symptom score and absolute change from baseline in morning and evening PEFR were significantly improved in the ßuticasone group as well: −53.7%, 63.7 L/min, and 52.7 L/min, respectively, compared with −32.9%, 37.6 L/min, and 27.2 L/min, respectively, in the montelukast group. SigniÞcantly more patients were satisÞed with ßuticasone therapy (83%) than with montelukast therapy (66%). Adverse events attributable to ßuticasone and montelukast treatment were not signiÞcantly different (7% and 8% of each group, respectively); the most common side effects reported were headache, common cold, and upper respiratory tract infection. Montelukast has been shown to attenuate LTD4-induced bronchoconstriction and exercise-induced bronchoconstriction. To compare the agent’s ability to reduce asthma exacerbations with that of salmeterol (in the presence of ßuticasone), researchers conducted a 52-week, randomized, multicenter, parallel-group, double-blind study involving 1,490 patients with chronic asthma aged 15–72 years (Bjermer L, 2003). A four-week run-in period during which all patients received ßuticasone 100 µg twice daily was followed by 48 weeks during which patients continued to receive ßuticasone 100 µg twice daily plus either montelukast (10 mg once daily) or salmeterol (50 µg twice daily). The trial’s primary end point was the percentage of patients with at least one asthma exacerbation (deÞned as worsening asthma requiring an unscheduled visit to a physician, emergency room, or hospital and/or exacerbations requiring treatment with oral, intravenous, or intramuscular corticosteroids). The difference in the number of asthma exacerbations between the treatment groups was insigniÞcant: 20.1% (150/747) of patients receiving montelukast/ßuticasone had an asthma exacerbation and 19.1% (142/743) of the patients receiving salmeterol/ßuticasone. The types of exacerbations and their distribution across each group were also insigniÞcant. The investigators reported that patients receiving salmeterol/ßuticasone experienced a signiÞcantly higher number of drug-related adverse events than patients receiving montelukast/ßuticasone during the trial—10% and 6.3%, respectively.

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The same was true for serious adverse events—7.4% and 4.6%, respectively (Bjermer L, 2003). Anti-immunoglobulin E Agents Overview. The anti-IgE monoclonal antibody omalizumab (also known as rhuMAb-E25; Genentech/Novartis/Sankyo/Tanox Biosystems’ Xolair) launched in the United States in 2003. Because IgE plays a central role in allergic respiratory diseases, the debut of omalizumab focused increased attention on the relationship between allergies and asthma. In the United States, the only country where omalizumab has received marketing approval to date, omalizumab is indicated for the treatment of moderate-to-severe persistent asthma in patients who have demonstrated reactivity to a perennial, airborne antigen and fail to respond to inhaled corticosteroids. Mechanism of Action. IgE plays a major role in allergic disease by binding cell surface IgE receptors, thereby forming receptor-IgE complexes, which (upon cross-linking with neighboring complexes) results in the release of histamine and other inßammatory mediators from mast cells and basophils. Omalizumab forms complexes with free, unbound IgE (the agent binds the same Fc portion of the IgE molecule as the high-afÞnity IgE receptor), blocking binding of IgE to mast cells and basophils and consequently inhibiting the release of inßammatory mediators (Figure 8). In addition, omalizumab does not bind to or recognize IgG molecules. Omalizumab. Omalizumab (also known as rhuMAb-E25; Genentech/Novartis/ Sankyo/Tanox Biosystems’ Xolair) is a recombinant, humanized antiimmunoglobulin E (IgE) monoclonal antibody that directly binds and neutralizes excess antibody in circulation. Omalizumab was approved in the United States Omalizumab

Anti-lgE binds free lgE and facilitates its removal through the reticuloendothelial system

lgE effector cells (mast cells and basophils) with high- (or low-) affinity lgE receptors Anti-lgE does not bind IgE that is already bound to high- (or low-) affinity lgE receptors

Result: Downregulation of lgE receptors lgE-mediated activation of effector cells is prevented lgE = Immunogloblin E.

FIGURE 8. Omalizumab’s mechanism of action.

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in June 2003 for the treatment of moderate-to-severe persistent asthma in adults and adolescents. In July 2005, the drug received marketing approval in Europe for the treatment of severe persistent allergic asthma. Omalizumab is in Phase III trials in Japan for allergic asthma and rhinitis. Tanox has marketing rights to the drug in some Asian markets, and Novartis and Genentech have marketing rights in the United States. In January 2003, Sankyo entered into an agreement with Novartis to codevelop and comarket omalizumab in Japan. Clinical data presented for omalizumab demonstrate that it has corticosteroidsparing effects. Investigators tested omalizumab in a randomized, double-blind, placebo-controlled trial involving 546 allergic asthmatics with moderate-to-severe asthma (Soler M, 2001). Ranging from 12 to 76 years of age, the subjects exhibited asthma symptoms despite inhaled corticosteroid use. Omalizumab-treated patients received subcutaneous injections (0.016 mg/kg) once every two or four weeks (depending on the weight of the patient) for seven months. During the Þrst four months, omalizumab-treated patients also received the dose of inhaled steroid at which they were stable. During the next three months (the steroid-reduction phase), the steroid dose was tapered by 25% every two weeks. Compared with placebo, omalizumab-treated patients exhibited 58% fewer exacerbations per patient during the stable phase and 52% fewer in the reduction phase. In addition, omalizumab reduced the required dose for inhaled corticosteroids; the average daily beclomethasone dose required in omalizumab-treated patients was 100 µg, compared with 300 µg in the placebo arm. Furthermore, 79% of patients treated with omalizumab, compared with 55% of patients on placebo, reduced their corticosteroid dose by 50%. By the end of the reduction phase, 43% of patients in the treatment group were able to withdraw from corticosteroids completely, compared with 19% in the placebo group. Local injection-site symptoms (bruising, redness, warmth, and mild itching), the most common adverse events that were likely drug-related, were observed in 4% (placebo-adjusted) of omalizumab-treated patients. Drug-related headaches occurred in 1% of patients, and fatigue and paresthesia occurred in 1%. No patient developed anti-omalizumab antibodies. Short-Acting Beta2 Agonists Overview. Because of their rapid onset of action and ability to relieve symptoms, short-acting beta2 agonists are used to treat acute asthma attacks. Typically, these agents are used on an as-needed basis because continuous administration is thought to reduce their effectiveness. Their effect may last for as long as eight hours (Lemanske RF, 1990), but the duration of action varies among the different agents in the class. This section discusses albuterol (the class leader in majormarket sales), also known as salbutamol (GSK’s Ventolin; Schering-Plough’s Proventil), and levalbuterol (Sepracor’s Xopenex), a recently launched isomer of racemic albuterol that offers greater potency without increased side effects. The market for short-acting beta2 agonists is currently dominated by low-cost generic agents in most countries, but environmental regulations banning the use of CFC propellants are forcing the removal of many generic agents. In the United

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States, the market for short-acting beta2 agonists is dominated by inexpensive, CFC-propelled generic albuterol, but in March 2005, the FDA announced that CFC-propelled albuterol will be withdrawn from the U.S. market as of January 1, 2009. A limited number of inhalers containing HFA are currently marketed in the United States; these agents are patent-protected, branded drugs. Therefore, the cost of rescue medication in the United States will increase considerably when CFC-MDIs are withdrawn. Mechanism of Action. Beta2 adrenergic receptors are found in smooth muscle in the airways, blood vessels, and other tissues throughout the body. In the airways, stimulation of these receptors by beta2 agonists relaxes bronchial smooth muscle and produces bronchodilation. Short-acting beta2 agonists have also been shown to have anti-inßammatory properties, including blocking the release of leukotrienes and inhibiting phospholipase A2, a key enzyme in the proinßammatory arachidonic acid pathway. Albuterol. Albuterol (GSK’s Ventolin; Schering-Plough’s Proventil, generics) (Figure 9) was approved for asthma in 1982. Known as salbutamol in Europe, this agent has more recently been recognized as effective in preventing symptoms of exercise-induced asthma when administered just before physical activities. Albuterol is also on the market as part of a combination therapy with the anticholinergic ipratropium bromide (Boehringer Ingelheim’s Combivent, Valeas’ Breva). Although albuterol has been on the market for decades, some details of the agent’s mechanism of action remain unclear. In a study designed to determine whether albuterol reverses bronchoconstriction by stabilizing mast cells, 16 mild asthmatics received 400 µg albuterol or placebo 30 minutes before being challenged with adenosine monophosphate ([AMP], which produces bronchoconstriction through the release of mediators from mast cells) or histamine (which induces direct smooth-muscle contraction). Prior to histamine challenge, albuterol and placebo inhalation increased FEV1 readings by 11% and 1.6%, respectively. Prior to AMP challenge, albuterol and placebo inhalation increased FEV1 by 16.4% and 0.3%, respectively. Albuterol provided protection against induced bronchoconstriction, as measured by the log of provocative concentrations (PC), causing a 20% or greater decline in FEV1 (PC20). The log PC20 for histamine after placebo and albuterol was -0.24 and 0.91, respectively. The log PC20 for AMP after placebo and albuterol was 0.69 and 2.22, respectively. Because these data suggest that OH HO

CH2

CH

NHR CH2

HO FIGURE 9. Structure of albuterol (R = C(CH3 )3 ).

464

ASTHMA

albuterol’s protective effect against AMP is signiÞcantly greater than the corresponding protection against histamine, the authors speculate that albuterol’s effects on lung function act through the stabilization of mast cells. Adverse events were not reported. Levalbuterol. Levalbuterol (Sepracor’s Xopenex), also known as levosalbutamol, launched in the United States in March 2002 in nebulized form for both adult and pediatric (6–11 years of age) asthmatic patients. An HFA-propelled MDI formulation of albuterol received FDA approval in March 2005. The MDI formulation launched in 2005, and became one of four HFA formulations of short-acting beta2 agonists available in the United States. Abbott Laboratories is copromoting levalbuterol in the United States with Sepracor. With the exception of levalbuterol (the [R]-isomer of albuterol), currently marketed beta2 agonists are racemic mixtures composed of both (R)- and (S)-isomers. The drive to develop levalbuterol began when the (R)-isomer of albuterol was discovered to be the active bronchodilator and the (S)-isomer was found to promote smooth-muscle contraction and oppose bronchodilation by increasing intracellular calcium levels in the airways (Yamaguchi H, 1996). A retrospective review of patients who had been hospitalized with asthma compared the efÞcacy of nebulized albuterol with that of nebulized levalbuterol. Thirty-Þve patients were nebulized with 2.5 mg albuterol every four hours, and 19 patients were nebulized with 1.25 mg levalbuterol every eight hours as medically necessary. Although FEV1 and forced vital capacity (FVC) values for patients in both groups did not differ signiÞcantly between hospital admission and discharge, the asthmatics receiving levalbuterol required 53% fewer nebulizer treatments (average of 14 treatments) than did patients in the albuterol group (average of 30 treatments). In addition, asthmatics treated with levalbuterol had a 27% shorter hospital stay (average of 3.3 days) than patients receiving albuterol (average of 4.5 days). Although fewer patients in the levalbuterol group required rescue nebulizations, the difference was not signiÞcant. This study did not Þnd a significant difference between levalbuterol and albuterol in the ability to improve lung function as measured by FEV1 , but previous studies involving outpatient analysis have reported signiÞcantly greater increases in FEV1 in asthmatics receiving levalbuterol (Milgrom H, 2001). Oral Corticosteroids Overview. Although oral corticosteroids are used less frequently than inhaled corticosteroids because of their high rate of side effects, short-term administration of oral corticosteroids is effective in patients with moderate-to-severe asthma in preventing progression of exacerbation, reversing inßammation, speeding recovery, and reducing rate of relapse. Long-term use of oral corticosteroids may prove efÞcacious for patients with severe asthma who do not respond to typical treatments. Side effects stemming from long-term use may include high blood pressure, osteoporosis, bone fractures, cataracts, muscle weakness, and slower

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growth in children. Prednisone (multisource) is the oral corticosteroid most commonly used for the treatment of asthma. Mechanism of Action. All corticosteroids exert their therapeutic effect by binding to glucocorticoid receptors, which are found in all cells and are abundant in the epithelium of bronchi. In this way, corticosteroids alter intracellular activities and reduce the transcription of genes that encode proteins involved in the inßammatory response, including the cytokines interleukin (IL)-1, -3, 4, -5, -6, and -8; TNF-α; GM-CSF; and RANTES. By lowering the levels of these inßammation mediators, corticosteroid therapy reduces levels of lymphocytes, eosinophils, macrophages, and mast cells in mucosal ßuids. In addition, corticosteroids increase the transcription of genes responsible for producing beta2 adrenoreceptors and lipocortin 1, a protein that inhibits phospholipase A2 (a proinßammatory enzyme). Other key mechanisms by which corticosteroids help alleviate the symptoms of asthma include inhibition of T-cell activation (and subsequent cytokine release), promotion of apoptosis of eosinophils, inhibition of NO synthase, and reduction in mucus secretion by submucosal gland cells. Evidence also suggests that these drugs help restore damaged epithelium. Prednisone. The oral, synthetic corticosteroid prednisone (multisource) (Figure 10)—used most often for its potent anti-inßammatory effects—has proved efÞcacious not only for asthma but also for arthritis, colitis, bronchitis, certain skin rashes, and allergic or inßammatory conditions of the nose and eyes. Prednisone is inactive in the body; to be effective, it must Þrst be converted to prednisolone by enzymes in the liver. Administering a short course of oral corticosteroids to asthmatics who are being discharged from a hospital setting is a common practice intended to reduce relapse. To compare the ability of Þve days of oral prednisone therapy (n = 261) to prevent relapse in children with acute asthma with the ability of two days of oral dexamethasone (n = 272), investigators conducted a prospective randomized trial with the primary outcome of relapse within ten days (Qureshi F, 2001). Patients were aged 2–18 years of age and had presented to a pediatric emergency room (ER) with acute asthma exacerbations. Prednisone and dexamethasone were

CH2OH C O

H3C

O OH

H3C

O FIGURE 10. Structure of prednisone.

466

ASTHMA

found to have similar outcomes when the following were analyzed: relapse rates (6.9% and 7.4%, respectively); hospitalization rates from the ER (12% and 11%, respectively) or after relapse (17% and 20%, respectively); and symptom persistence at ten days’ post-treatment (21% and 22%, respectively). Prednisone did, however, result in signiÞcant increases in the following (compared with dexamethasone): exclusion due to vomiting in the ER (3% and 0.3%, respectively); parental noncompliance (4% and 0.4%, respectively); and missing at least two days of school (19.5% and 13.2%, respectively). This trial suggests that although efÞcacy measures are similar, dexamethasone may offer some beneÞts in compliance and side-effect parameters in children with acute asthma. Methylxanthines Overview. Methylxanthines have been used as bronchodilators for treating asthma for the past 50 years. In this class, theophylline (AstraZeneca’s TheoDur, 3M Pharmaceuticals’ Theolair, generics) is the most frequently used agent for asthma treatment. Theophylline was once a Þrst-line therapy for asthma, but with the emergence of newer, more efÞcacious agents, it has been relegated to third- or fourth-line status in patients whose disease is not adequately controlled by corticosteroids and long-acting beta2 agonists. Mechanism of Action. Methylxanthines probably have several mechanisms of action, but questions remain as to which are responsible for their bronchodilating effect. The intracellular cyclic nucleotides cyclic adenosine monophosphate (cAMP) and cyclic guanine monophosphate (cGMP) regulate bronchodilation. By inhibiting phosphodiesterase (PDE) isoenzymes, particularly PDE types 3 and 4, which degrade cAMP and cGMP, methylxanthines relax smooth muscle in the pulmonary arteries and airways. Methylxanthines are also nonselective antagonists of adenosine receptors found on the surface of some cells. Although not important in mediating bronchodilation, adenosine-receptor antagonism by methylxanthines increases ventilation during hypoxia and reduces respiratory muscle fatigue. Theophylline. Theophylline (AstraZeneca’s Theo-Dur, 3M’s Theolair, generics) (Figure 11) is one of the oldest and most widely available drugs used for the treatment of asthma, but its use has declined in the past decade, and it is no longer a preferred pharmaceutical in asthma therapy. Given the adverse events associated with theophylline use, the agent is typically reserved for patients with refractory disease (i.e., asthmatics whose symptoms are not well controlled by typical asthma agents). In a trial designed to compare the efÞcacy and safety of doxofylline with that of theophylline, 346 patients with chronic, reversible bronchial asthma received oral administration of doxofylline 200 µg or 400 µg, theophylline 250 µg, or placebo for 12 weeks (Goldstein MF, 2002). The primary efÞcacy measurements were FEV1 readings. Statistically signiÞcant mean percentage increases in FEV1 two hours post-treatment were observed in patients receiving 400 mg doxofylline and

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250 mg theophylline, compared with patients receiving placebo (mean percent increases in FEV1 ranged from 10% to 15% above increases with placebo). Beginning at week 2 of the study, both the doxofylline 400 mg and theophylline 250 mg groups experienced a signiÞcant decline (six- to eightfold) in the average number of asthmatic attacks per day. This reduction in asthma attacks was found to accompany signiÞcant declines in albuterol use, particularly in patients in the doxofylline 400 mg and theophylline 250 mg groups, where between 1 and 1.5 fewer mean puffs/day were observed. The percentage of patients dropping out because of an adverse event was signiÞcantly lower in the doxofylline 400 mg group (11.4%) than in the theophylline 250 mg group (31.4%). The lower-dose doxofylline (200 mg) group and the placebo group experienced percentage dropout rates of 3.6% and 3.4%, respectively. The most common adverse events included headache, nausea, nervousness, and insomnia. The investigators claim that for every Þve patients treated with theophylline 250 mg, one adverse event was spared by doxofylline 400 mg. Both oral and intravenous administration of methylxanthines cause systemic side effects. Therapeutic serum concentrations fall within the range of 10–20 mg/mL, but such side effects as nausea, vomiting, tremors, and mild central nervous system symptoms may occur with only slightly higher concentrations, in the range of 20–30 mg/mL. Serum concentrations greater than 30 mg/mL are associated with a small risk of seizure or myocardial infarction. Because of methylxanthines’ narrow therapeutic index, patient serum concentrations must be monitored upon initiation of therapy and at least twice yearly thereafter. Use of sustained-release theophylline formulations, the most commonly used formulations, reduces the wide variations in serum concentration that characterize the immediate-release formulations. Cromolyns Overview. Cromolyn sodium (King Pharmaceuticals/SanoÞ-Aventis’ Intal) and necrodomil sodium (King Pharmaceuticals/SanoÞ-Aventis’ Tilade) have different chemical structures, but both are inhaled, nonsteroidal anti-inßammatory drugs (NSAIDs) for the treatment of asthma. When administered on a longterm basis, given four times daily, they reduce the frequency and severity of early- and late-phase allergen-induced reactions, exercise-induced asthma, and

FIGURE 11. Structure of theophylline.

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airway hyper-responsiveness. Cromolyns are often used to reduce seasonal hyperresponsiveness in asthmatics who have sensitivity to grass pollen. Because these drugs are not easily absorbed through mucous membranes, high concentrations of cromolyns are not found in the systemic circulation, and side effects are minimal. Cromolyns are extremely safe and are among the least toxic drugs used in asthma management. Therefore, these agents are more frequently prescribed for children than for adults. In recent years, however, physicians’ worries about inhaled steroids have eased somewhat, and the use of cromolyns is declining in the pediatric population as well. Mechanism of Action. Although cromolyns’ precise mechanism of action is unclear, they appear to inhibit the degranulation of sensitized and nonsensitized mast cells that occurs after exposure to speciÞc antigens. Cromolyn sodium is known to inhibit the release of histamine and SRS-A (the slow-reacting substance of anaphylaxis, a leukotriene) from the mast cell, resulting in membrane stabilization and inhibition of the early asthmatic response. Cromolyn Sodium. Cromolyn sodium (King Pharmaceuticals/SanoÞ-Aventis’ Intal) (Figure 12) is indicated by the Expert Panel II report (Second Expert Panel Report on Guidelines for the Diagnosis and Management of Asthma. NIH publication No. 97-4051; July 1997) as a component of therapy in the treatment of mild persistent and moderate persistent asthma, but it offers little beneÞt during an acute exacerbation of asthma. The agent has very few side effects and demonstrates better efÞcacy in asthmatic children than in asthmatic adults. Cromolyn administered as a dry powder can cause minor throat irritation and transient cough in some children; the following study was designed to assess the ability of a cromolyn solution, administered via nebulizer, to reduce symptoms and attacks and reduce the need for concomitant bronchodilators in children (Prenner BM, 1982). Cromolyn 20 mg was nebulized four times a day for eight weeks in two- to seven-year-old asthmatics. In addition to a physician’s overall assessment of asthma severity, symptom scores for dyspnea, tightness, wheeze, and cough were taken at baseline and in the Þnal two weeks of treatment. Scores taken near the end of the trial were signiÞcantly reduced (between threefold and sixfold) over baseline. Five of the 19 children were able to discontinue theophylline use, and 13 children had reduced theophylline use by at least 85% by the last two weeks of the trial. Fourteen children required emergency care at

O

O

O

O

OH NaOOC

O

O

FIGURE 12. Structure of cromolyn sodium.

COONa

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least once during the trial, and the incidence of these events did not change over the eight-week treatment period. Measures of the number of times the children were awakened by asthma and the number of days on which asthma interfered with normal activity did not reach signiÞcance. The investigators note that these numbers were already relatively low at baseline. This trial was not placebocontrolled, which adds to the difÞculty in interpreting these data. Anticholinergics Overview. Anticholinergics are the Þrst bronchodilators used to treat asthma. They act at cholinergic muscarinic receptors to reduce bronchial smooth-muscle tone and to block the neurogenic bronchoconstriction response triggered by inhaled irritants. Despite these favorable characteristics, marketed anticholinergics such as ipratropium bromide (Boehringer Ingelheim’s Atrovent) and oxitropium bromide (Boehringer Ingelheim’s Oxivent) are not ideal asthma agents. In addition to binding the M1 and M3 muscarinic receptors responsible for bronchoconstriction, these agents also bind to M2 receptors, thereby blocking the favorable bronchodilating effects of this receptor subtype. As discussed in “Emerging Therapies,” companies are developing novel, more-selective anticholinergics that inhibit only the M1 and M3 receptors. Mechanism of Action. Anticholinergic drugs block the effects of acetylcholine, a parasympathetic nervous system neurotransmitter that promotes bronchoconstriction. Acetylcholine, released from branches of the vagus nerve that run along the airways, binds to the M1 and M3 muscarinic receptors located in the smooth muscle and submucosal glands in the airways. Acetylcholine binding activates the receptors, stimulating both the contraction of smooth muscle (leading to bronchoconstriction) and the secretion of mucus from the submucosal glands. By binding to the M1 and M3 muscarinic receptors, anticholinergic drugs block the access of acetylcholine, thus reducing the number of activated receptors. As a result, smooth-muscle tone in the airways declines, thereby reducing bronchoconstriction and mucus secretion. Ipratropium Bromide. Ipratropium bromide (Boehringer Ingelheim’s Atrovent) (Figure 13) is the best studied of the approved anticholinergics. Boehringer Ingelheim has also developed oxytropium (Oxivent) and an ipratropium-containing combination therapy (Combivent), which is discussed in the next section. Ipratropium bromide alleviates bronchoconstriction by blocking the effects of acetylcholine. In addition, it is thought to be a useful adjunctive agent in the treatment of asthma and has shown increased efÞcacy when combined with a beta2 agonist (see “Ipratropium/Albuterol” section). Unlike adrenergic bronchodilators (such as albuterol), ipratropium’s spirometric beneÞts do not decline over time. In addition, because the agent is poorly absorbed and does not readily enter the central nervous system, high doses of ipratropium bromide can be administered to asthmatics.

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ASTHMA

In a trial designed to compare the bronchodilating effects of ipratropium bromide delivered by different inhalers, 70 patients (children and adolescents) with stable bronchial asthma were initially administered 40 µg of the agent via DPI (Ulrik CS, 1992). Twenty-one percent of the patients responded (deÞned as patients who experienced at least a 15% increase in FEV1 30 minutes after inhalation) and were then enrolled in the study’s investigational phase (a double-blind, randomized, crossover trial). Responders received 40 µg ipratropium bromide, administered by both an Ingelheim powder device system (IPI) and an MDI on two separate days less than a week apart. FEV1 measurements were taken after 15, 30, 60, 120, 180, 240, 300, and 360 minutes. Although FEV1 (% of predicted) values were lower for the agent when inhaled via MDI (5.1% and 11.2% at 15 and 30 minutes, respectively) versus IPI (12.2% and 16.2% at 15 and 30 minutes, respectively), this difference did not reach signiÞcance; both groups receiving ipratropium bromide experienced signiÞcant improvement in lung function compared with placebo. For both groups, the maximum FEV1 values (% of predicted) occurred 30 minutes post-administration, and maximal response lasted 90 minutes. No adverse events due to inhalation of ipratropium bromide were reported, and no reduction in patient FEV1 measures was observed. Anticholinergic agents can induce local and systemic side effects (e.g., blurred vision, cardiac stimulation). Uptake of anticholinergics has steadily declined since the introduction of the more speciÞc and efÞcacious beta2 agonists. Ipratropium/Albuterol. Ipratropium/albuterol (Boehringer Ingelheim’s Combivent, Valeas’ Breva) combines anticholinergic and beta2 agonist agents in a single inhaler. Anticholinergic agents promote bronchodilation primarily in the large airways, where cholinergic nerves and muscarinic receptors predominate. Beta2 agonists dilate mainly the bronchioles, where the beta-adrenergic receptors abound. Because these two drug classes act on different airways, combining an anticholinergic with a beta2 agonist should logically produce greater bronchodilation than either agent alone. Whether anticholinergics and beta2 agonists truly have additive or synergistic effects is uncertain. In general, however, lower doses of two agents are preferable to higher doses of a single agent because the risk of side effects is reduced. In particular, combination therapy may limit the dangers

FIGURE 13. Structure of ipratropium bromide.

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associated with the overuse of short-acting beta2 agonists. Combining anticholinergic and beta2 agonist agents in a single inhaler also reduces the cost of therapy and improves patient compliance. A double-blind, randomized, prospective trial involving 180 asthmatics hospitalized for exacerbation was designed to compare the combination of ipratropium and albuterol with albuterol and placebo (Rodrigo GJ, 2000). Primary outcome measures were improvement of pulmonary function (as measured by FEV1 and PEFR) and hospital admission rates. Agents were administered via MDI/spacer at ten-minute intervals for three hours (24 puffs = 2, 880 µg of albuterol and 504 µg of ipratropium). Patients receiving ipratropium/albuterol experienced signiÞcant average increases in FEV1 (48.1%) and PEFR (20.5%) compared with the control group. Post-administration, 39% of the patients in the albuterol/placebo group and 20% of the patients in the albuterol/ipratropium group were admitted, a 49% reduction in risk of admission for the ipratropium/albuterol group. Further analysis demonstrated that patients with more severe pulmonary obstruction (FEV1 ≤ 30% of predicted) were more likely to beneÞt from the addition of higher doses of ipratropium to beta2 agonist therapy than were patients with nonsevere pulmonary obstruction. With the exception of dry mouth (experienced by 50% of the ipratropium group and 22% of the control group), adverse events due to therapy did not differ signiÞcantly between treatment groups. The most common adverse events experienced by both groups were tremors and headache. EMERGING THERAPIES Current agents used to treat asthma exert their therapeutic effect by reversing bronchoconstriction, reducing inßammation, or combining these two actions. Although many emerging therapies exploit the same strategies, others are directed at novel targets or are targeted more speciÞcally than their predecessors. Table 4 summarizes the drug therapies in development for asthma. Several new agents that combine long-acting beta2 agonists and inhaled corticosteroids are in early-stage development. Noteworthy combinations now in Phase II clinical trials include Altana/SanoÞ-Aventis’s ciclesonide/formoterol combination; GlaxoSmithKline’s GSK-685698 and GSK-159797; ScheringPlough/Novartis’s mometasone (Schering-Plough’s Asmanex) and formoterol; and Skye Pharma’s combination of ßuticasone and formoterol (Flutiform). Because trial data on these new combination agents are scarce, they are not discussed in detail here. Inhaled Corticosteroids Overview. Novel corticosteroids are in various stages of development worldwide. The most commercially promising agents are those that have fewer side effects than their predecessors already on the market. The potential for side effects with long-term treatment is an ongoing concern for patients (especially children) with persistent asthma and contributes to noncompliance. Inhaled corticosteroids

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TABLE 4. Emerging Therapies in Development for Asthma Compound

Development Phase

Marketing Company

Inhaled corticosteroids Ciclesonide (Alvesco) United States Europe Japan

PR R PR

Altana/Sanofi-Aventis Altana Teijin

Long-acting beta2 agonists Arformoterol United States Europe Japan

IIIa — —

Sepracor — —

Phosphodiesterase 4 inhibitors Roflumilast United States Europe Japan

III PR II

Altana Altana Altana/Tanabe

Anticytokine agents Daclizumab (Zenapax) United States Europe Japan

II — —

Protein Design Labs/Roche — —

Mepolizumab United States Europe Japan

III — —

GlaxoSmithKline — —

Anticholinergics Tiotropium bromide United States Europe Japan

— III II

— Boehringer Ingelheim/Pfizer Boehringer Ingelheim

Cell adhesion inhibitors R-411 United States Europe Japan

II II —

Roche Roche —

IVL-745 United States Europe Japan

— II —

— Sanofi-Aventis —

a Phase III development for chronic obstructive pulmonary disease.

PR = Preregistered; R = Registered.

currently in development are designed to enable less-frequent dosing with fewer adverse effects. Altana/SanoÞ-Aventis/Teijin’s ciclesonide is preregistered for the treatment of asthma in the United States; the Þrst European launch took place in the United Kingdom in January 2005. Several other corticosteroids are in earlyphase trials for asthma, including GW-685698 (GlaxoSmithKline), NCX-1020

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(NicOx), and NS-126 (SS Pharmaceutical). However, owing to the scarcity of published clinical trial data, only ciclesonide will be discussed in detail here. Mechanism of Action. All corticosteroids exert their therapeutic effect by binding to glucocorticoid receptors, which are found in the immune cells that play a role in asthma and are abundant in the epithelium of bronchi. Corticosteroids alter intracellular activities and reduce the transcription of genes that encode proteins involved in the inßammatory response, including the cytokines interleukin (IL)-1, -3, -4, -5, -6, and -8; tumor necrosis factor-alpha (TNF-α); granulocytemacrophage colony-stimulating factor (GM-CSF); and regulated upon activation, normal T cells expressed and secreted (RANTES). By lowering the levels of these inßammation mediators, corticosteroid therapy reduces the activity of lymphocytes, eosinophils, macrophages, and mast cells in mucosal ßuids. Other key mechanisms by which corticosteroids help alleviate the symptoms of asthma include inhibition of T-cell activation (and subsequent cytokine release), promotion of apoptosis of eosinophils, inhibition of nitric oxide (NO) synthase, and reduction in mucus secretion by submucosal gland cells. Evidence also suggests that these drugs help restore damaged epithelium. Ciclesonide. In an attempt to reduce systemic absorption and side effects, researchers are developing corticosteroid therapies that act locally via activated metabolites. The most advanced of these corticosteroid pro-drugs (activated in the lungs) is ciclesonide, a once-daily, metered-dose inhaler (MDI) that is being codeveloped by Altana (formerly Byk Gulden), SanoÞ-Aventis, and Teijin. In 2001, Altana entered into an agreement with Aventis for the joint development and marketing of ciclesonide in the United States, where a new drug application (NDA) was submitted to the FDA in late December 2003. In October 2004, Altana received an approvable letter from the FDA for the treatment of persistent asthma in adults and in children at least four years old. Altana stated in a press release dated October 26, 2004, that the company is working with the FDA to resolve speciÞc clinical issues outlined in the letter. Altana markets ciclesonide in Europe, where the drug recently received approval for the treatment of persistent asthma in adults through the European Mutual Recognition Procedure. In January 2005, Altana announced ciclesonide’s launch in the United Kingdom. In January 2004, Teijin, Altana’s partner in Japan, submitted an approval application in Japan for the use of inhaled ciclesonide in asthma. Altana and SanoÞ-Aventis are also developing a combination inhaler that joins ciclesonide and formoterol in SanoÞ-Aventis’ Ultrahaler device; this combination product is in Phase II clinical trials. Ciclesonide is unique among inhaled corticosteroids because it is designed to be active primarily in the lung. Inhaled ciclesonide is the inactive parent compound of the active metabolite desisobutyryl-ciclesonide (des-CIC). Ciclesonide is converted to des-CIC by esterases that are expressed predominantly in the lung but also, at much lower levels, at most other sites in the body (Reynolds NA, 2004). Therefore, the portion of ciclesonide that is not delivered to the lung but is deposited orally in the pharynx and swallowed is far less likely to be converted

474

ASTHMA

to the active form than the portion delivered to the lung. Consequently, systemic exposure to the drug is minimized. Two 12-week, placebo-controlled trials of ciclesonide were presented at the 2002 Annual Meeting of the American Thoracic Society (ATS). In one trial, 360 patients were treated once daily in the morning, either with placebo, 100 µg ciclesonide, or 400 µg ciclesonide (Engelst¨atter R, 2002). Primary study end points were increases over baseline in morning peak expiratory ßow (PEF) and forced expiratory volume at one second (FEV1 ). Patients treated with placebo experienced a reduction in FEV1 of 28 mL; patients treated with 100 µg and 400 µg ciclesonide experienced increases of 129 mL and 192 mL, respectively. Morning PEF increased by 2 L/min and 3 L/min in the 100 µg and 400 µg ciclesonide groups, respectively; in the placebo group, PEF fell by 18 L/min. In another trial, 329 patients were treated either with placebo or with 200 µg or 800 µg ciclesonide (Chapman KR, 2002). In this trial, patients treated with placebo experienced a signiÞcant decline in both FEV1 (144 mL) and PEF (28 L/min), while FEV1 and PEF remained stable in patients treated with either dose of ciclesonide. Head-to-head trials indicate that ciclesonide has efÞcacy comparable to that of other low-dose corticosteroids. In a 12-week, randomized trial, 554 asthmatic patients received either 80 µg or 320 µg ciclesonide (once daily in the morning) or 200 µg budesonide twice daily (Engelst¨atter R, 2003). Both FEV1 and forced vital capacity (FVC) were reported to be signiÞcantly increased in the three treatment groups (data not available). SigniÞcant reductions were observed in asthma symptoms and use of rescue medications in all treatment groups (data not available). Importantly, both of the once-daily doses of ciclesonide were as effective as two daily doses of budesonide in improving morning and evening PEF, reducing the use of rescue medications, and increasing FEV1 and FVC (data not available). In addition, treatment with budesonide was associated with cortisol suppression in trial participants, while treatment with ciclesonide had no signiÞcant effect on urinary cortisol excretion. (Urinary cortisol excretion assays are often performed to determine whether a patient receiving corticosteroid therapy is adrenally suppressed, a condition commonly associated with chronic, high-dose corticosteroid therapy.) Long-Acting Beta2 Agonists Overview. Long-acting beta2 agonists demonstrate prolonged receptor occupancy, compared with short-acting beta2 agonists; accordingly, they are prescribed to provide partial, steady-state bronchodilation. When prescribed in conjunction with inhaled corticosteroids, beta2 agonists may also prevent airway remodeling. For this reason, therapeutic guidelines recommend that long-acting beta2 agonists be prescribed as adjuncts to corticosteroid therapy in cases of moderate persistent and severe persistent asthma. Several long-acting beta2 agonists are in development for asthma. Of particular note is a group of compounds originally developed by Theravance. In

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January 2003, GlaxoSmithKline (GSK) and Theravance announced an agreement to develop up to eight long-acting beta2 agonists using GSK’s drug delivery technology. GSK secured worldwide marketing rights to these agents as part of the agreement; among them are several candidates in development with a goal of once-daily dosing for asthma, including GSK-159797 and GSK-597901. Clinical trial data on these agents are not yet available, so they are not discussed in detail here. Mechanism of Action. Beta2 adrenergic receptors are found in smooth muscle in the airway and in the lungs, blood vessels, and other tissues throughout the body. In the airway, stimulation of these receptors by beta2 agonists activates adenylate cyclase, increasing the production of cyclic adenosine monophosphate (cAMP) and causing bronchodilation by relaxing smooth muscle. Other effects of beta2 agonists that have been observed experimentally include the following: • • • •

Blocking of mast-cell release of leukotrienes and histamine in the lungs. Reduced microvascular permeability and mucus production. Increased mucociliary function. Possible inhibition of the activity of phospholipase A2, an enzyme in the proinßammatory arachidonic acid pathway.

Arformoterol. Arformoterol, Sepracor’s single-isomer R,R-formoterol, is in Phase III trials in the United States for asthma and chronic obstructive pulmonary disease (COPD). In a press release dated October 26, 2004, Sepracor announced completion of two pivotal Phase III studies of arformoterol in COPD and plans to discuss submission of the arformoterol NDA for the COPD indication. The development status of arformoterol for the treatment of asthma is uncertain because no clinical data have been released for some time; the company’s web site lists COPD as the only indication for which the agent is currently in development (www.sepracor.com/products/pipeline.html. Accessed February 25, 2005). Arformoterol is a single-isomer version of its parent compound, formoterol, a racemic mixture of R,R- and S,S-formoterol marketed by Yamanouchi and Novartis. Sepracor is developing arformoterol based on evidence that the R,Renantiomer is 1,000 times more potent than the S,S-enantiomer. In vitro assays indicate that the S,S enantiomer may act as a beta2 receptor antagonist and impair bronchodilation (Moore RH, 2001; Mhanna MJ, 2003). In a randomized, double-blind, single-dose, six-way crossover study, 31 patients with asthma and FEV1 readings at 40–70% of predicted value were given arformoterol (12, 24, 48, or 72 µg), salbutamol (2.5 mg), or placebo (Sepracor, media release, 2001). Peak percentage changes in FEV 1 readings for arformoterol were signiÞcantly higher than for placebo (arformoterol generated changes of 45–52%, while placebo generated a 22% increase) but not signiÞcantly different from salbutamol peak percentage changes (55.5%). Although onset of action of the two agents was similar, arformoterol doses of 24 µg and 72 µg resulted in signiÞcantly higher percentage changes in FEV1 at the 24-hour postdose time point (data not available). The number of asthma exacerbations and adverse events was

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lower in arformoterol recipients than in placebo and salbutamol recipients (data not available). In a media release dated September 20, 2001, Sepracor revealed partial data from a separate Phase II study, in which 340 patients with asthma demonstrated signiÞcant improvement in lung function when treated with a range of doses of arformoterol. The subsequent increase in FEV1 readings, relative to baseline, was in the range of 24–27%. The duration of action of the drug was up to 24 hours. Data on adverse events were not available. Phosphodiesterase 4 Inhibitors Overview. NonspeciÞc phosphodiesterase (PDE) inhibitors, such as the currently available theophylline, are often relegated to second- or third-line status owing to their relatively poor side-effect proÞles. As a result, PDE inhibition research is now focused on targeting individual PDE enzymes (each with distinct activities and tissue distributions) in hopes of improving side-effect proÞles and efÞcacy. Inhibition of PDE4 is of particular interest because of its preferential expression in inßammatory cells thought to participate in the pathogenesis of asthma. Unfortunately, Þrst-generation PDE4 inhibitors demonstrated undesirable side effects such as headache, nausea, and emesis. Because PDE4 inhibitors represent a more speciÞc therapeutic approach in the treatment of airßow limitation (compared with theophylline), companies are trying to improve the sideeffect proÞles of these compounds. Roßumilast (Altana/Tanabe’s Daxas) is the most clinically advanced PDE4 inhibitor in development. Also in development are GSK/Elbion’s 842470 (AWD-12281), Almirall-Prodesfarma’s arofylline, and Kyowa Hakko Kogyo’s KW-4490, but the lack of published trial data precludes in-depth analysis of these drugs. GSK’s cilomilast was in development for asthma, but development for this indication has been discontinued. Mechanism of Action. Inßammatory cells thought to participate in the pathogenesis of asthma (and COPD) are known to express PDE4. PDE4 breaks down cAMP in inßammatory cells, including eosinophils, and in tracheal smoothmuscle cells. PDE4 inhibition elevates cellular levels of cAMP, which in turn suppresses the inßammatory response and causes bronchodilation. Therefore, PDE4 has become a molecular target of interest in the development of novel therapies for asthma and COPD. Roflumilast. Roßumilast, the most clinically advanced PDE4 inhibitor, is being developed by Altana/Tanabe as an orally administered, once-daily treatment for asthma and COPD. Altana entered into a worldwide codevelopment and comarketing agreement with Pharmacia (now PÞzer) in April 2002. Roßumilast is in development for asthma in the United States (Phase III), Europe (Phase III), and Japan (Phase II trials under way by Tanabe). However, in July 2005 PÞzer returned all rights to the drug to Altana. The NDA Þling in the United States was expected in 2005, but in October 2004, Altana announced that because of the challenge of recruiting adequate numbers of patients for clinical trials, the Þling would be delayed until 2006.

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Several abstracts from the 99th Annual Meeting of the ATS in May 2003 (Seattle, Washington) suggest that roßumilast is an effective and well-tolerated treatment for asthma. In a 12-week, randomized, double-blind study, 499 asthmatics received either a once-daily oral dose of 500 µg roßumilast or 400 µg inhaled beclomethasone dipropionate (BDP) administered as two daily doses of 200 µg each (Bousquet J, 2003). Both agents signiÞcantly improved lung function. Mean changes compared with baseline for FEV1 were 0.30 L for roßumilast and 0.37 L for BDP; for FVC, 0.30 L and 0.36 L, respectively; and for morning PEFR, 21.59 L and 26.94 L, respectively. Both agents also signiÞcantly reduced the need for rescue medication and lowered asthma symptom scores to a similar degree. Although the abstract does not provide data on adverse events, the authors claim that 500 µg roßumilast (oral, once-daily) is comparable to inhaled, twice-daily BDP (400 µg). A randomized, double-blind, parallel-group, dose-ranging study of 690 patients examined the dose-dependent effects of roßumilast on FEV1 (Leichtl S, 2002). At baseline, the mean FEV1 was 73%. Patients were treated with 100 µg, 250 µg, or 500 µg of roßumilast once daily. FEV1 increased by 11% (260 mL) in the 100 µg treatment group, by 13% (320 mL) in the 250 µg treatment group, and by 16% (400 mL) in the 500 µg treatment group. All three treatment groups also demonstrated improvements in morning PEF (10 L/min, 12 L/min, and 20 L/min, respectively, for the 100 µg, 250 µg, and 500 µg treatment groups) and in evening PEF. For all measures examined, the improvement in patients treated with the highest dose was greater than in patients treated with the lowest dose. It should be noted that this study lacked a placebo control and that details of adverse events were not provided in the published abstract. Data from clinical trials for the COPD indication provide some insight into the likelihood and nature of adverse events observed in patients treated with roßumilast. In a randomized, placebo-controlled trial of 1,411 patients with moderate-to-severe COPD, the most frequently reported adverse events included diarrhea, nausea, and abdominal pain (Bateman ED, 2004). Most adverse events were described as being mild to moderate in intensity. Anticytokine Agents Overview. The 2003 launch of the biological therapy omalizumab (also known as rhuMAb-E25; Genentech/Novartis/Sankyo/Tanox Biosystems’ Xolair) has directed much attention toward biologics-based approaches to asthma therapy. GSK’s mepolizumab, an IL-5 antagonist, is in Phase II clinical trials. Several other anticytokine agents—including Genaera/Medimmune’s anti-IL-9 monoclonal antibody and Bayer’s IL-4 antagonist pitrakinra—are in development for asthma, but are not analyzed here because of a lack of sufÞcient published trial data. Several companies are developing anticytokine agents that disrupt multiple targets. Regeneron’s dual IL-4/IL-13 antagonist is in Phase I clinical trials, and few data on its potential efÞcacy are available. Therefore, it is not discussed in detail here.

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Mechanism of Action. To date, most anticytokine research for asthma has focused on agents that inhibit IL-4, IL-5, and more recently, IL-13 and IL-2. Although the functions of IL-4 and IL-13 appear to overlap, a distinct role is emerging for IL-13 in allergic (and parasitic) responses, with potential implications for drug development. IL-4 mediates key proinßammatory events, including induction of IgE isotype switching, increased expression of cell adhesion molecules, and increased mucus production. The cytokine IL-5 is essential for the maturation of eosinophils and their release from the bone marrow. It is well documented that IL-5 is highly expressed in the bronchial mucosa of asthmatics. An association also exists between the number of activated eosinophils in the airways and the clinical severity of asthma. Expression of both IL-2 and the IL-2 receptor are elevated in asthmatic patients, and IL-2 has been implicated in the development of corticosteroid-resistant asthma. Daclizumab. Daclizumab (Protein Design Labs/Roche’s Zenapax) is a humanized monoclonal antibody directed against the interleukin-2 (IL-2) receptor. Daclizumab is currently marketed for the prevention of kidney transplant rejection but is in development for several other indications, including the treatment of persistent asthma. In September 2004, Roche and Protein Design Labs agreed to jointly develop and market daclizumab worldwide for the treatment of persistent asthma. Daclizumab inhibits IL-2-dependent activation of T cells by blocking the alpha subunit of the human high-afÞnity IL-2 receptor. Peripheral blood lymphocyte cultures from patients with asthma have disproportionate numbers of IL-2-positive T cells and elevated levels of IL-2. Preclinical data indicate that daclizumab inhibits production of several cytokines produced by activated T cells and implicated in the pathogenesis of asthma, including interferon-gamma (IFN-γ ), tumor necrosis factor-alpha (TNF-α), IL-10, IL-5, and IL-4 (McClellan M, 2002). Preliminary results from an ongoing Phase II trial of daclizumab were presented at the American Academy of Allergy, Asthma, and Immunology meeting on March 23, 2004, in San Francisco (Busse WW, 2004). The double-blind, placebo-controlled study enrolled 116 patients with chronic persistent asthma not adequately controlled by high-dose inhaled corticosteroids. The primary end point measured in the study was the change in FEV1 from baseline to day 84 of the study. Patients received an IV infusion of daclizumab or placebo every two weeks; all patients continued treatment with inhaled triamcinolone acetonide throughout the 12-week study period. Patients treated with daclizumab showed a mean increase in FEV1 of 4.4%, compared with a decline of 1.52% in patients treated with placebo. Patients treated with daclizumab also showed prolonged time to asthma exacerbations requiring corticosteroid rescue. Mepolizumab. GSK’s anti-IL-5 MAb mepolizumab (SB-240563) is in Phase III clinical trials in the United States for the treatment of severe asthma and atopic dermatitis. GSK holds a worldwide, exclusive license for Protein Design Labs’ patents covering humanized antibodies directed against IL-5. In turn, GSK

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has agreed to pay Protein Design Labs a licensing fee, maintenance fees, and royalties on any future sales. In a Phase II, multicenter, double-blind, placebo-controlled trial, three intravenous infusions (750 mg/dose) of mepolizumab or placebo were administered to 24 patients with atopic asthma over a 20-week period (Flood-Page PT, 2003). Although mepolizumab reduced blood eosinophils to negligible levels, the effect on bone marrow and airway eosinophils compared with placebo was less pronounced, producing a median decline from baseline of 52% and 55%, respectively. Subsequent experiments demonstrated that eosinophil numbers returned to baseline values 4–20 weeks after administration of the Þnal dose of mepolizumab. Clinical measurements of lung function, such as FEV1 and morning PEFR, were not signiÞcantly different between the mepolizumab and placebo groups. Anticholinergics Overview. Anticholinergics, the Þrst bronchodilators used to treat asthma, act at cholinergic muscarinic receptors to reduce bronchial smooth-muscle tone and block the neurogenic bronchoconstriction response triggered by inhaled irritants. Although they are less potent bronchodilators than the beta2 agonists, anticholinergics are still prescribed for some asthma patients. In the most recently updated treatment guidelines of the Global Initiative on Asthma (GINA), anticholinergics are recommended as alternative rescue medications for patients who experience adverse effects (such as tachycardia, arrhythmia, and tremor) from short-acting beta2 agonists. Some physicians prescribe them as third- or fourth-line maintenance therapy for patients whose disease is not adequately controlled by inhaled corticosteroids and long-acting beta2 agonists. Tiotropium bromide (Boehringer Ingelheim/PÞzer’s Spiriva) is a reÞnement of an existing therapeutic approach that should improve lung function and provide greater symptom relief and greater convenience than existing anticholinergics. Mechanism of Action. Anticholinergic drugs block the effects of acetylcholine, a parasympathetic neurotransmitter that promotes bronchoconstriction. Released from branches of the vagus nerve that runs along the airways, acetylcholine binds to the M1 and M3 muscarinic receptors located in the smooth muscle and the submucosal glands of the airways. Acetylcholine binding activates the receptors, stimulating both the contraction of smooth muscle (leading to bronchoconstriction) and the secretion of mucus from the submucosal glands. By binding to the M1 and M3 muscarinic receptors, anticholinergic drugs block the access of acetylcholine, reduce the number of activated receptors, and as a result, reduce mucus secretion and smooth-muscle tone in the airways. However, currently marketed anticholinergics (such as ipratropium [Boehringer Ingelheim’s Atrovent]) are not ideal because they also interfere with M2 receptors, which facilitate bronchodilation. Tiotropium Bromide. Tiotropium bromide (Boehringer Ingelheim/PÞzer’s Spiriva) is marketed in the United States and Europe for COPD; in Japan, the drug is registered for COPD. Phase III trials for asthma were reported in Germany

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in 1997, and Phase II trials were reported in the United Kingdom and Japan in the same year, but the drug’s development status for asthma is not clear. Although the agent is not approved for the treatment of asthma, physician audit data indicate that physicians are prescribing the drug to asthmatic patients. Currently marketed anticholinergics bind to and block activation of M1 and M3 receptors, which promote bronchoconstriction, but they also interfere with M2 receptors, which facilitate bronchodilation. Although tiotropium bromide binds all three receptor types, the agent dissociates faster from the M2 receptor than from the M1 and M3 receptors, thereby retaining the beneÞcial bronchodilating effect—a distinct advantage in this class of agents. Tiotropium bromide is approximately ten times more potent than ipratropium and has a longer duration of action. To analyze tiotropium bromide’s duration of bronchodilation, a double-blind, placebo-controlled, single-dose crossover study was conducted in 12 atopic patients with methacholine-induced bronchoconstriction (O’Conner BJ, 1996). Patients received either placebo or tiotropium bromide at 10, 40, or 80 µg, followed by methacholine PC20 challenge. Tiotropium bromide caused bronchodilation that was sustained for 24 hours at all doses. The increase in FEV1 relative to placebo for 10, 40, and 80 µg tiotropium bromide at each time point was 6.7%, 8.2%, and 10%, respectively, at 2 hours; 10.4%, 5.5%, and 11.1% at 12 hours; and 7.3%, 7.1%, and 9.4% at 24 hours. FEV1 readings did not differ signiÞcantly from placebo at 36 and 48 hours post-drug administration. Each dose of tiotropium bromide resulted in signiÞcant protection against methacholine challenge in all patients at all time points. Peak protection occurred at the two-hour time point, when the protective effect was 5, 7.1, and 7.9 doubling doses for 10, 40, and 80 µg of drug, respectively. Tiotropium bromide was well tolerated; no adverse events occurred at any dosage. Cell Adhesion Inhibitors Overview. Reducing the inÞltration of cells into asthmatic airways by blocking cell adhesion pathways is a novel approach to asthma therapy. Monoclonal antibodies speciÞc for adhesion molecules expressed on endothelial cells, epithelial cells, and eosinophils have been successful in blocking key steps of the inßammatory response in animal models of asthma. This section analyzes two cell adhesion inhibitors, R-411 (Roche) and IVL-745 (SanoÞ-Aventis), both of which target the very-late-antigen 4 (VLA-4) integrin receptor present on inßammatory cells such as eosinophils (R-411 also targets α4β7). Additional cell adhesion inhibitors in development that are not covered here (owing to lack of data) include Encysive (formerly Texas Biotechnology)/Revotar’s bimosiamose and SanoÞ-Aventis’ 1031. In February 2005, Biogen Idec and Elan announced a voluntary marketing suspension of the VLA-4 antagonist natalizumab (Tysabri) following two reports of progressive multifocal leukoencephalopathy (PML) in multiple sclerosis patients taking a combination of natalizumab and interferon beta-1a (Biogen Idec’s Avonex). In March 2005, the FDA announced that it was suspending clinical trials of other drugs in the same class, including GSK’s experimental multiple

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sclerosis drug 683–699. Roche has stated that it will continue to develop three VLA-4 antagonists, citing mechanistic differences between its compounds and those currently coming under scrutiny. Mechanism of Action. Integrin receptors promote cell adhesion and communication, leading to increases in migration, activation, and survival of inßammatory cells. The integrin receptors α4β1 (VLA-4) and α4β7 are known to be expressed on the surfaces of mast cells, eosinophils, and Th2 cells, all key players in asthma pathogenesis. In fact, VLA-4 and α4β7 have been shown to be more highly expressed in asthmatic patients, a Þnding that has made these receptors key targets for developing inhibitory drugs that would prevent inßammation by blocking the integrin-adhesion molecule connection. VLA-4 binds speciÞcally to vascularcell adhesion molecule 1 (VCAM-1), while α4β7 binds to both VCAM-1 and mucosal adressin cell adhesion molecule 1 (MAdCAM-1). R-411. Roche’s R-411 is a VLA-4 and α4β7 “dual” antagonist under investigation as a once-daily oral therapy for asthma. Phase II trials began in April 2002 in Europe; they include a dose-ranging study in patients not treated with inhaled corticosteroids, a study investigating whether R-411 can act as replacement therapy for inhaled corticosteroids, and a pediatric safety and pharmacokinetics study for chronic asthma. Roche expects its Þrst Þling for approval of R-411 to take place in 2007. R-411 interferes with the interaction between VLA-4 and VCAM-1, preventing eosinophil migration, which is observed in the late asthmatic response. VLA-4 is expressed by particular leukocytes and interacts with VCAM-1 on the endothelium to mediate leukocyte extravasation (migration of leukocytes from a vessel into surrounding tissue) to lung tissue. Early-stage trial data from a small study that assessed the pharmacokinetics and tolerability of R-411 in 50 healthy people were presented at the 2004 meeting of the ATS (Abbas R, 2004). Adverse events occurred at similar rates in people treated with R-411 and in people treated with placebo. No serious adverse events were reported. The study determined that R-411’s half-life was compatible with once-daily dosing. IVL-745. Prior to the acquisition of Aventis by SanoÞ-Synth´elabo, IVL-745 was one in a series of VLA-4 antagonists held by Aventis. Aventis announced that Phase IIa trials began in February 2002, but results from these trials have not yet been released, and no news on the drug’s development status has been released by SanoÞ-Aventis. In vitro studies suggest that IVL-745 is a powerful inhibitor of eosinophil adhesion to VCAM-1 (Cairns JA, 2000). Research in ovalbumin-challenged allergic rat models demonstrated that 3 mg/kg and 10 mg/kg IVL-745 were able to reduce ovalbumin-induced lung pathology in both the epithelial and bronchiolar regions of the lung (Sargent CA, 2000). In addition, these doses signiÞcantly reduced the numbers of eosinophil and lymphocyte cell populations in the mucosal/submucosal region of the bronchioles. Duration of action analysis demonstrated that 10 mg/kg IVL-745 inhibited bronchial inßammation when

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administered 6 hours, 4 hours, and 30 minutes before and 4 hours after ovalbumin challenge. REFERENCES Abbas R, et al. Safety, tolerability, and pharmacokinetics of R-411, a dual α4β1 and α4β7 integrin antagonist after multiple ascending doses in healthy subjects. 100th Annual International Meeting of the American Thoracic Society; May 22–26, 2004; Orlando, Florida. Abstract D31/Poster C4. Adams PF, et al. Current estimates from the National Health Interview Survey, 1996. National Center for Health Statistics Vital Health Statistics. 1999;10:83–84. Adcock IM, et al. Molecular interactions between glucocorticoids and long-acting beta 2 agonists. Journal of Allergy and Clinical Immunology. 2002;110(6 Suppl):S261–8. Affrime MB, et al. Bioavailability and metabolism of mometasone furoate following administration by metered-dose and dry-powder inhalers in healthy human volunteers. Journal of Clinical Pharmacology. 2000;40(11):1227–1236. [a] Affrime MB, et al. Mometasone furoate has minimal effects on the hypothalamic-pituitaryadrenal axis when delivered at high doses. Chest. 2000;118(6):1538–1546. [b] Agertoft L, et al. Effect of long-term treatment with inhaled budesonide on adult height in children with asthma. New England Journal of Medicine. 2000;343(15):1064–1069. Akinbami LJ, Schoendorf KC. Trends in childhood asthma: prevalence, health care utilization, and mortality. Pediatrics. 2002;110:315–322. Akinbami LJ, et al. U.S. childhood asthma prevalence estimates: the impact of the 1997 National Health Interview Survey Redesign. American Journal of Epidemiology. 2003;158:99–104. Akiyama K. Review of epidemiological studies on adult bronchial asthma in Japan. Nihon Kyobu Shikkan Gakkai Zasshi. 1994;32(suppl):200–210. Allen DB. Inhaled corticosteroids: past lessons and future issues. Journal of Allergy and Clinical Immunology. 2003;112(3 suppl):S1–S40. Altemeier WA, Gavin GR. How are allergic rhinitis and sinusitis connected with asthma? Pediatric Annals. 2000;29:391–398. American Lung Association (ALA). Trends in asthma morbidity and mortality. American Lung Association. Epidemiology and Statistics Unit. Best Practices and Program Services. April 2004. Available at www.lungusa.org. Accessed November 1, 2004. Anderson HR. Trends in prevalence and severity of childhood asthma. British Journal of General Practice. 1994;308:1600–1604. Anderson HR, et al. Trends in prevalence of symptoms of asthma, hay fever, and eczema in 12–14 year olds in the British Isles, 1995–2002: questionnaire survey. British Medical Journal. 2004;328:1052–1053. Anderson SD, et al. Duration of protection by inhaled salmeterol in exercise-induced asthma. Chest. 1991;100:1254–1260. Apter AJ, Szeßer SJ. Advances in adult and pediatric asthma. Journal of Allergy and Clinical Immunology. 2004;113:407–414. Arif AA, et al. Prevalence and risk factors of asthma and wheezing among U.S. adults: an analysis of the NHANES III data. European Respiratory Journal. 2003;21:827–833. Asthma in America Survey (AIA). 2000. Available at www.asthmainamerica.com.

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Braun-Fahrlander C, et al. No further increase in asthma, hay fever, and atopic sensitization in adolescents living in Switzerland. European Respiratory Journal. 2004;23: 407–413. Brown PJ, et al. Asthma and irreversible airßow obstruction. Thorax. 1984;39(2):131–136. Burney P. The changing prevalence of asthma? Thorax. 2002;57(suppl II):ii36–ii39. Burrows B, et al. Association of asthma with serum IgE levels and skin-test reactivity to allergens. New England Journal of Medicine. 1989;320:271–277. Busse WW, et al. Budesonide delivered by the Turbuhaler is effective in a dose-dependent fashion when used in the treatment of adult patients with chronic asthma. Journal of Allergy and Clinical Immunology. 1998;101(4 pt 1):457–463. Busse WW, et al. Asthma. New England Journal of Medicine. 2001;344:350–362. Busse WW, et al. Preliminary safety and efÞcacy of daclizumab in the treatment of patients with moderate to severe chronic persistent asthma. 60th Annual Meeting of the American Academy of Allergy, Asthma, and Immunology; May 19–23, 2004; San Francisco, CA. Poster 1043. Cairns JA, et al. The VLA-4 antagonist IVL-745 is a novel and potent inhibitor of eosinophil adhesion to VCAM-1. American Journal of Respiratory and Critical Care Medicine. 2000;161(suppl pt 2):202. Celedon J. Asthma, rhinitis, and skin test reactivity to aeroallergens in families of asthmatic subjects in Anqing, China. American Journal of Respiratory and Critical Care Medicine. 2001;163:1108–1112. Centers for Disease Control and Prevention (CDC). Asthma prevalence and control characteristics by race/ethnicity—United States, 2002. Morbidity and Mortality Weekly Report. 2004;53:145–148. Centers for Disease Control and Prevention (CDC). Reducing childhood asthma through community-based service delivery. Morbidity and Mortality Weekly Report. 2005; 54:11–14. Chapman KR, et al. Effects of ciclesonide versus placebo on lung function after 12 weeks of treatment in patients with asthma. 98th Annual International Meeting of the American Thoracic Society; May 17–22, 2002; Atlanta, GA. Abstract. Charpin D, et al. Epidemiologic identiÞcation of allergic rhinitis. Allergy. 1996;51: 293–298. Chervinsky P, et al. Fluticasone propionate aerosol for the treatment of adults with mild to moderate asthma. The Fluticasone Propionate Asthma Study Group. Journal of Allergy and Clinical Immunology. 1994;94:676–683. Chetta A, et al. Airway remodeling is a distinctive feature of asthma and is related to severity of disease. Chest. 1997;111:852–857. Childhood Asthma Management Program (CAMP) Research Group. Long-term effects of budesonide or necrodomil in children with asthma. New England Journal of Medicine. 2000;343:1054–1063. Chinn S, et al. Increase in diagnosed asthma but not in symptoms in the European Community Respiratory Health Survey. Thorax. 2004;59:646–651. Clarke JR, et al. Evidence for genetic associations between asthma, atopy, and bronchial hyperresponsiveness: a study of 8- to 18-yr-old twins. American Journal of Respiratory and Critical Care Medicine. 2000;162(6):2188–2193.

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Crohn’s Disease

ETIOLOGY AND PATHOPHYSIOLOGY Introduction Crohn’s disease (CD) is a chronic, relapsing/remitting inßammatory disease of the gastrointestinal (GI) tract. The regions of the GI tract most often affected by CD are the small intestine and the large intestine, or colon, including the rectum. Unlike ulcerative colitis (UC)—a related inßammatory bowel disease (IBD) in which inßammation and ulceration are limited to the mucosal and submucosal layers of the large intestine and rectum—the inßammation and ulceration of CD can extend through all layers of the intestinal wall in both the small and large intestines. Common symptoms of CD include diarrhea, abdominal pain, rectal bleeding, and weight loss, and complications such as intestinal abscesses, Þstulas, and intestinal obstructions (discussed in the section “Local Complications”) are common in CD patients. The course of CD is intermittent, with disease exacerbations followed by periods of remission. Because of similarities in their pathogenesis and clinical manifestations, CD and UC are often collectively referred to as IBD and are frequently examined together in research studies. Whenever possible, Þndings speciÞc for CD are reported here, and comments are made on IBD results when no deÞnitive differentiation can be made between the two diseases. Currently, the etiology and pathophysiology of CD are not well understood, but three major factors have been implicated in the disease: Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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First, the intestinal damage of CD is known to be mediated by chronic inßammation. As a result, many of the therapies currently used or in development for CD are immunosuppressants. Inhibitors of tumor necrosis factor-alpha (TNFα) and other inßammatory cytokines appear to be particularly promising for the treatment of CD. Second, the aberrant immune response in CD is thought to be initiated and/or perpetuated by intestinal microbes, including the normal intestinal ßora. Therefore, antibiotics are used to treat CD, and researchers are exploring ways to enhance the innate immune response against microbes as a treatment strategy for CD. Third, several genetic factors predispose people to develop CD, and many of these factors appear to be involved in immunity—for example, the nucleotide oligomerization-binding domain 2 (NOD2) gene. Research into how these genes cause CD may reveal future drug targets.

Anatomy Figure 1 illustrates the major constituent organs of the GI tract. As mentioned previously, the small and large intestine, including the rectum, are the areas of the GI tract most often affected by CD. In a small minority of patients, the disease can affect the mouth, the tongue, the esophagus, or the stomach. Food enters the small intestine from the stomach through the pyloric sphincter. The small intestine consists of three segments—the duodenum, the jejunum, and the ileum—and is responsible for most digestion of food and absorption of nutrients. Remaining material from the small intestine then enters the large intestine through the ileocecal valve, which allows only one-way transit of material, blocking colonic bacteria from entering the small intestine. The large intestine comprises the cecum—a blind pouch that is adjacent to the ileocecal valve—and Þve other segments: the ascending colon, the transverse colon, the descending colon, the sigmoid colon, and the rectum. The colonic phase of digestion is responsible for the Þnal reabsorption of water and electrolytes. The rectum—the distal portion of the large intestine and the Þnal portion of the GI tract—begins at the sigmoid colon and ends at the anus. Peristalsis (strong wavelike contractions) of the colon forces fecal material into the sigmoid colon and rectum, and distention of the rectal walls elicits a reßex that initiates the defecation process. The walls of the small intestine and large intestine comprise four layers: the mucosa, submucosa, muscularis, and serosa, illustrated in Figure 2. The inßammation and ulceration associated with CD are transmural (they can affect all layers of the intestinal wall), and diseased segments are often ßanked by regions of normal mucosa, thus resulting in a characteristic pattern of “skip lesions” (Figure 3). Etiology The Immune Response.. The human GI tract is routinely exposed to a wide variety of antigens, including those that are ingested from food and ßuids and those produced by the natural bacterial ßora that reside in the colon and participate

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FIGURE 1. The major constituent organs of the gastrointestinal tract.

in the digestive process. In healthy people, there is continuous, clinically undetected immune stimulation in response to these antigens. In CD, scientists believe, this antigenic stimulation results in an aberrant immune response that is both prolonged and ampliÞed. Neither the precise cause of the aberrant response nor a speciÞc antigenic trigger has been identiÞed. One leading theory proposes that CD results from a genetic predisposition affecting the immune system combined with an environmental trigger, such as exposure to endogenous or exogenous intestinal antigens. Therefore, in an effort to inhibit this characteristic aberrant immune response, immunosuppressive agents are used to treat CD patients; these agents include aminosalicylates, corticosteroids, and immunosuppressants (e.g., azathioprine, 6-mercaptopurine, methotrexate, and cyclosporine), all of which are discussed in greater detail in “Current Therapies.” Many immune cells—including neutrophils, macrophages, B and T lymphocytes, and mast cells—are present in the mucosal layer of healthy intestines.

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FIGURE 2. The four layers of the intestinal wall.

The intact epithelium lining the mucosa prevents these immune cells from being overstimulated by the large antigenic load to which the GI tract is exposed on a daily basis. According to one theory of how the disease is initiated, CD patients have increased intestinal permeability (perhaps due to a genetic predisposition) that exposes the immune cells to a multitude of antigens. In a study involving 11 CD patients, 32 healthy relatives of the patients, and 17 healthy volunteers who served as controls, researchers found that the CD patients and their relatives had a nearly twofold increase in intestinal permeability (demonstrated by enhanced absorption of polyethylene glycol-400 ingested with a meal), compared with the control participants (Hollander D, 1986). Animal studies have also demonstrated that compromises in mucosal integrity can cause CD-like lesions (Sanders DS, 2005). Thus, enhanced intestinal permeability could cause overexposure of immune cells to intestinal antigens, thereby prompting an aberrant immune response and subsequent tissue damage. Researchers believe that a cell-mediated immune response predominates in CD (Figure 4). The inßammatory response associated with CD is believed to be driven by activation of T-helper 1 (TH1) cells, as evidenced by the increased levels of interleukin (IL)-2 and interferon-gamma (IFN-γ ) in the intestinal mucosa of CD patients (Stenson WF, 1999). In contrast, UC is thought to be associated primarily with a TH2 response. The Inflammatory Cascade. CD exhibits classic characteristics of chronic inßammation: tissue inÞltration by large numbers of lymphocytes and

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FIGURE 3. Anatomical distribution patterns of Crohn’s disease.

macrophages and high levels of proinßammatory cytokines in the intestinal mucosa. During active disease, increased numbers of neutrophils and monocytes migrate from the bloodstream into the intestinal mucosa and submucosa, where they secrete large quantities of proinßammatory molecules, such as cytokines and tissue-damaging free radicals. Figure 5 illustrates key immune and inßammatory responses associated with CD. Investigation of these proinßammatory molecules has provided new therapeutic targets for CD and other immune-related diseases,

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FIGURE 4. A TH 1 response predominates in Crohn’s disease.

and many current and emerging CD pharmacotherapies target these proinßammatory molecules. Table 1 summarizes major inßammatory mediators involved in CD pathogenesis; the following sections discuss some of them. Cytokines. Specimens of inßamed mucosa taken from patients with CD have been shown to contain increased quantities of the cytokines IL-1β, IL-2, IL-6, IL8, IL-12, IL-18, TNF-α, and IFN-γ ; these molecules activate the immune system and promote tissue inßammation. Long-term overexpression of these molecules can lead to tissue damage, as occurs in CD. Conversely, blockage of cytokine overexpression may reduce inßammation and subsequent tissue damage. Furthermore, TNF-α has been shown to induce epithelial cell apoptosis, alter the expression of adhesion molecules, and affect tight junction functionality, thereby increasing intestinal permeability (Sanders DS, 2005). As mentioned previously, increased intestinal permeability is thought to be important in the initiation of CD. These cytokines, therefore, are potential drug targets in the treatment of CD. Currently, one TNF-α inhibitor, inßiximab (Centocor [a Johnson & Johnson subsidiary]/Schering-Plough/Tanabe Seiyaku’s Remicade), is approved for CD and is highly efÞcacious in treatment of the disease (discussed in “Current Therapies”). Additionally, two novel TNF-α inhibitors, several IL-12 inhibitors, an IL-6 inhibitor, and an IFN-γ inhibitor are in clinical development for CD (discussed in “Emerging Therapies”). Lipid Mediators. Prostaglandins and leukotrienes, commonly referred to as eicosanoids, are lipid inßammatory mediators. Mucosal tissues of CD patients contain markedly elevated levels of prostaglandin E2 (PGE2) and leukotriene B4

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TABLE 1. Major Inflammatory Mediators in Crohn’s Disease Mediator Cytokines Interferon-gamma (IFN-γ )

Interleukin-1 beta (IL-1β)

Interleukin-2 (IL-2) Interleukin-6 (IL-6)

Interleukin-8 (IL-8) Interleukin-10 (IL-10)

Interleukin-12 (IL-12)

Interleukin-18 (IL-18)

Transforming growth factor (TGF) Tumor necrosis factor-alpha (TNF-α)

Lipid mediators Leukotriene B4 (LTB4)

Primary Sources

Biological Activities

T cells, natural killer cells

Enhances immune activation through upregulation of major histocompatability complex (MHC) class I and II molecules and Fc receptors; potentiates production of tumor necrosis factor-alpha (TNF-α) by macrophages; promotes tissue infiltration by neutrophils and monocytes by up-regulating expression of intracellular adhesion molecule-1 (ICAM-1) on endothelial cells. Macrophages, B cells, Increases production of other inflammatory dendritic cells, mediators; stimulates phagocytes and neutrophils, T cells, lymphocytes; promotes release of endothelial cells, acute-phase proteins from liver; induces epithelial cells fever. Stimulates proliferation of T cells and natural Activated T helper 1 killer cells. (TH 1) cells Macrophages, T and B Produces acute-phase proteins; promotes cells, fibroblasts, differentiation of B cells; promotes antibody endothelial cells production. T cells, monocytes Attracts neutrophils and additional T cells. Monocytes, Induces differentiation of TH cells into TH 2 cell subtype; suppresses TH 1 functions; macrophages, downregulates IFN-γ , IL-1β, IL-2, IL-6, IL-8, subsets of T and B and TNF-α production. cells Monocytes, natural Induces differentiation of TH cells into TH 1 cell subtype; suppresses TH 2 functions; induces killer cells IFN-γ , IL-1β, IL-2, IL-6, IL-8, and TNF-α production; activates natural killer cells. Macrophages, Induces IFN-γ , IL-1β, IL-8, and TNF-α dendritic cells, production; activates endothelial cells and intestinal epithelial neutrophils; upregulates expression of cell cells adhesion molecules; attracts neutrophils and T cells. Activated Inhibits proliferation of T and B cells, macrophages, T cells macrophage activation, and the production of other inflammatory mediators; stimulates growth of connective tissue. Activated Activates endothelial cells, macrophages, and macrophages, some neutrophils; upregulates expression of cell lymphocytes adhesion molecules; upregulates the synthesis of other cytokines; induces fever and cachexia. Activated neutrophils, macrophages, mast cells

Vasoconstriction; increases vascular permeability; increases leukocyte adhesion to vascular endothelium and subsequent tissue infiltration; stimulates enzyme secretion by leukocytes.

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TABLE 1. (continued) Mediator Prostacyclin (PGI2 ) Prostaglandin D2 (PGD2 ) Prostaglandin E2 (PGE2 )

Thromboxane A2 (TXA2 ) Platelet-activating factor (PAF)

Other mediators Complement fragments (e.g., C5a)

Primary Sources Vascular endothelium, macrophages Mast cells, many other tissues White blood cells, many other tissues

Platelets, leukocytes Neutrophils, macrophages, mast cells, endothelial cells Result of immune response

Biological Activities Acute inflammatory response; vasodilation; inhibits platelet aggregation. Vasodilation. Acute inflammatory response; vasodilation; increases vascular permeability; induces fever and hyperalgesia; inhibits gastric acid secretion; inhibits renal tubular reabsorption. Vasoconstriction; increases vascular permeability; stimulates platelet aggregation. Activates neutrophils; increases vascular permeability; promotes vasoconstriction and platelet aggregation.

Vasodilation; increases vascular permeability; attracts neutrophils.

(LTB4). Although both PGE2 and LTB4 induce vasodilation and increase vascular permeability (actions that enhance tissue inÞltration by leukocytes) clinical studies suggest that these molecules play distinct roles. LTB4 is also a potent chemotactic factor that can attract neutrophils to the mucosa, whereas PGE2 is involved in wound healing. PGE2 and LTB4 are metabolic products of arachidonic acid, a lipid released from cell membranes. Typically, cells contain only limited amounts of arachidonic acid, but the increased macrophage activity in CD results in the release of larger amounts. Macrophages, epithelial cells, and Þbroblasts generate PGE2 through the cyclooxygenase (COX-1 and COX-2) pathway; neutrophils, macrophages, and mast cells produce LTB4 via the 5-lipooxygenase (5-LO) pathway. Aminosalicylates and corticosteroids, both currently used in CD treatment, inhibit the production of leukotrienes and prostaglandins (discussed in “Current Therapies”). Cell Adhesion Molecules. Leukocyte migration is mediated by cell adhesion molecules (CAMs), which include selectins (P-, E-, and L-selectin), integrins (e.g., β2 integrin), and the immunoglobulin superfamily of adhesion molecules (e.g., intracellular adhesion molecule-1 [ICAM-1]). These molecules are expressed on leukocytes and on endothelial cells, which line blood vessels. CAM overexpression correlates with several inßammatory disorders, including CD. Currently, one CAM inhibitor—Elan/Biogen Idec’s natalizumab (Tysabri)—is in late-stage clinical development for CD (discussed in “Emerging Therapies”). Mitogen Activated Protein (MAP) Kinases. The MAP kinase signal transduction pathway—which comprises the p38, JNK, and ERK pathways—stimulates

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FIGURE 5. Key immune and inflammatory responses in Crohn’s disease.

the synthesis of proinßammatory cytokines. This pathway can also be activated by the binding of these cytokines to their respective receptors and thus helps to regulate cellular responses to proinßammatory cytokines. Therefore, inhibition of the MAP kinase pathway has the potential to inhibit the feedback loop common to inßammatory conditions involving proinßammatory cytokines; several MAP kinase inhibitors are in development for CD (discussed in “Emerging Therapies”). Microbial Involvement. For decades, investigators have postulated that an infectious agent is involved in CD pathogenesis, but no single agent has been consistently recovered from CD patients. One hypothesis of the etiology of CD holds that normally nonpathogenic bacteria—potentially even bacteria of the

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normal intestinal ßora—may cause CD in susceptible hosts. In fact, leukocytes present in active intestinal lesions from IBD patients do display a reduced tolerance to certain normal intestinal ßora (Duchmann R, 1995). Diversion of the fecal stream can also prevent CD relapses (Rutgeerts P, 1991). Antibodies directed against the cell-wall phosphopeptides—speciÞcally the sugar moiety of the yeast Saccharomyces cerevisiae (anti-S. cerevisiae antibodies, or ASCAs)—have been detected in as many as 79% of CD patients (Barclay GR, 1992; Sendid B, 1996), which suggests a possible association between the organism and CD. Although several diagnostic kits are available to measure ASCA levels in CD patients, the presence or absence of ASCAs cannot be used to make a deÞnitive diagnosis of CD because ASCAs have been detected in the serum of healthy people and in the serum of patients with other forms of IBD, including UC and indeterminate colitis (Walker LJ, 2004). The precise clinical importance of this serological marker remains to be determined, but ASCA antibodies have been shown to be more highly associated with gastroduodenal and ileal CD, stricturing and penetrating CD, and CD requiring surgery than with colonic disease or CD with an inßammatory phenotype (Walker LJ, 2004). Mycobacterium paratuberculosis has also been implicated as a potential causative agent of CD because it triggers Johne’s disease, an intestinal disorder of cattle and sheep that displays pathological changes similar to those associated with CD, and because it has been isolated from a small number of CD surgical specimens. However, CD patients are rarely cured by antituberculosis drugs. M. paratuberculosis does not appear to play a major role in CD, although it may cause CD-like disease in a small number of people. Paramyxovirus-like particles have been isolated from affected tissues of some CD patients, leading some researchers to suspect the measles virus as a putative cause of CD. Researchers have also speculated that people born up to three months after a measles epidemic are at higher risk for developing CD before age 30 and that maternal measles infection during pregnancy may increase the fetus’s risk of developing this disorder (Ekbom A, 1990; Ekbom A, 1991; Ekbom A, 1994). Measles vaccination has been implicated as a risk factor for developing IBD. Such a fear could dissuade parents from immunizing their children. However, a meta-analysis of studies suggests that measles vaccination is not associated with increased risk of developing CD (Ghosh S, 2001). In addition to the association of CD with particular microbes, predisposition to develop CD has been linked to mutations in genes involved in the innate immune system—the body’s Þrst line of defense against many common microorganisms—again suggesting a possible microbial connection to CD. These genes include the NOD2 gene and toll-like receptors (TLRs) (Hugot JP, 1996; Hugot JP, 2001; Ogura Y, 2001) (discussed further in the “Risk Factors, Genetics” section). Risk Factors Genetics. Results from several family-based epidemiological studies suggest that genetics may play a role in the development of CD. The disease has been

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shown to aggregate in families, and relatives of CD patients are at increased risk for developing the disease. In a study involving 964 CD patients from the Mount Sinai Hospital IBD Research Unit, 10% were shown to have a family member afßicted with IBD; of these 96 patients, 78.1% had family members who were afßicted with CD only (McLeod RS, 1997). Despite these observations, no clear pattern of Mendelian inheritance has been associated with CD. Further evidence of the role of genetics in the etiology of CD is derived from studies of twins; these studies have found concordance rates for the development of CD in identical twins that range from 42% to 58% (Bonen DK, 2003). Nevertheless, the fact that concordance rates in identical twins are less than 100% supports the theory that other factors are involved in the etiology of the disease. Evidence that CD is a multigenic disorder comes from gene expression analysis (via DNA microarray) that utilized RNA extracted from resected bowel specimens from CD, UC, and control patients. Of the 7,306 genes analyzed, 170 were differentially regulated in CD, UC, or both (Lawrance IC, 2001). Numerous genetic linkage and association studies have identiÞed potential CD- and IBD-susceptibility loci on chromosomes 1, 3, 5, 6, 7, 12, 14, 16, and 19 (Bonen DK, 2003). Researchers have identiÞed numerous genes within these loci, as well as other genetic factors, that could potentially be involved in the pathogenesis of CD; many of these genes code for molecules that are involved in the immunoregulatory and inßammatory processes. Perhaps the most important genetic discovery in understanding CD etiology was the identiÞcation of an association of the NOD2 gene on chromosome 16 with CD (Hugot JP, 1996; Hugot JP, 2001; Ogura Y, 2001). The NOD2 protein, which is predominantly expressed by monocytes and by lipopolysaccharide [LPS]-stimulated macrophages, is an intracellular receptor for muramyl dipeptide, a peptidoglycan fragment found in Gram-positive and Gram-negative bacterial cell walls (Girardin SE, 2003; Inohara N, 2003; Philpott DJ, 2004). It belongs to the Apaf-1/Ced-4 superfamily of proteins involved in apoptosis regulation and has structural homology to disease-resistant proteins found in plants (Hugot JP, 2001; Ogura Y, 2001). Researchers observed CD-associated mutations in either of two amino-terminal caspase-recruitment domains (CARD) and in a leucine-rich repeat (LRR), the region responsible for activating the transcription factor NF-κB. NF-κB, in turn, activates expression of many genes, including pro-inßammatory cytokines, such as TNF-α. People who possess nonconservative amino-acid substitutions or frame-shift mutations in the LRR were found to be at increased risk of developing CD, with a relative risk of 3 for simple heterozygous mutation and 38 for homozygous mutation (Hugot JP, 2001). Mutations in the NOD2 gene are more commonly associated with early-onset disease and CD characterized by an ileal or stricturing (Þbrostenotic) phenotype, compared with colonic disease (Abreu MT, 2002; Ahmad T, 2002; Cuthbert AP, 2002; Lesage S, 2002). The physiological function of NOD2 is not yet clearly deÞned, but the protein appears to be important in the production of antimicrobial peptides, known as defensins, that are involved in innate mucosal immunity against bacteria, fungi, and viruses. Defensins are produced in the small intestine, mainly by Paneth

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cells, which are specialized secretory epithelial cells lining the intestinal walls that also express the NOD2 protein. In one study of 45 CD patients and 12 negative controls, CD patients with involvement of the small intestine had decreased expression of ileal defensins, compared with controls or CD patients without ileal involvement; this decrease was signiÞcantly more pronounced in patients with NOD2 mutations (Wehkamp J, 2004). Interestingly, CD conÞned to the colon also seems to be associated with impaired induction of colonic defensins, but this impairment is not associated with NOD2 mutations (Wehkamp J, 2005). Further evidence for the theory that mutations in NOD2 impair the intestinal immune response against microbes comes from animal studies: NOD2-deÞcient mice were unable to express a subgroup of murine intestinal microbial peptides and were susceptible to bacterial infection introduced orally (Kobayashi KS, 2005). However, controversy still lingers over the role of NOD2 mutations in CD. Some studies have suggested that defensins are overexpressed, not decreased, in the intestinal mucosa of CD patients, thereby contradicting the observations cited in the preceding paragraph (Lawrance IC, 2001; Cunliffe RN, 2002; Wehkamp J, 2002). Furthermore, NOD2 mutations in mice corresponding to mutations found in CD patients resulted in elevated NF-κB activation and IL-1β production. Unlike the NOD2-deÞcient mice that did not exhibit intestinal inßammation, these mice showed increased susceptibility to bacterial-induced intestinal inßammation (Maeda S, 2005). Therefore, these researchers believe that the NOD2 mutations associated with CD are gain-of-function mutations that result in overactivation of the innate mucosal immune system, which in turn leads to chronic intestinal inßammation. These researchers propose IL-1β as a good therapeutic target for the treatment of CD, but no speciÞc IL-1β inhibitors are currently in active development for CD. Additional research is needed to determine precisely the role that the NOD2 gene plays in CD. Geography. Evidence suggests that the incidence of CD displays a north-south gradient. The disease occurs more commonly in northern and western Europe than in southern and central Europe (Karlinger K, 2000). It is also more common in North America than in Africa, Asia, and Australia, but whether this Þnding is attributable to geographical location or ethnicity is not known. CD also occurs more frequently in urban areas than in rural regions. Race and Ethnicity. Epidemiological studies have demonstrated that CD is more common in Caucasians than in people of other races. Additionally, CD incidence is three to eight times higher in people of Jewish descent (Stenson WF, 1999). Interestingly, the incidence of CD is higher in U.S. and European Jews than in Israeli Jews. Ethnic and regional variations also occur in CD patients who carry NOD2 mutations (Aldhous MC, 2003). For example, studies have shown that the three most common NOD2 mutations found in Caucasian CD patients occur less frequently in African-American CD patients and were not present in a sample of 350 Japanese CD patients (Bonen DK, 2002; Inoue N, 2002).

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Smoking. Cigarette smoking appears to be a risk factor for developing CD. A meta-analysis of several studies suggests that smokers are two to four times more likely to develop CD than nonsmokers (Calkins BM, 1989). Smoking also appears to affect the frequency of disease recurrence in CD patients (Sutherland LR, 1990). Researchers examined the surgery recurrence rate in 174 patients who had previously undergone surgery for CD and found that patients who smoked had recurrence rates of 36% and 70% after Þve and ten years, respectively. The Þve- and ten-year recurrence rates were only 20% and 41%, respectively, for nonsmoking CD patients. The reason for the correlation between smoking and CD is not well understood, although nicotine has been shown to have an inhibitory effect on TH2 cell function (believed to predominate in UC) but no effect on TH1 cells (believed to predominate in CD) (Madretsma S, 1996). TH1 and TH2 cells are mutually inhibitory based on the cytokines each cell type produces, and IL-10 (produced by TH2 cells) blocks activation of TH1 cells (Monteleone G, 2000). Thus, nicotine’s inhibitory effect on TH2 cells may in turn promote TH1 cell differentiation. Diet. Diet affects digestive tract physiology and may inßuence CD etiology. In general, populations that consume non-Western diets (including Africans, Chinese, East Indians, and Japanese) have a low incidence of CD. In addition, researchers have proposed a direct relationship between CD and the consumption of reÞned sugar and between CD incidence and reduced fruit and vegetable consumption (Persson PG, 1987). In studies involving CD patients, consumption of wheat, dairy, corn, and yeast products was associated with IBD recurrence (Rhodes J, 1986; Riordan AM, 1993; Workman EM, 1984). However, additional studies are required to establish a deÞnitive association between diet and CD. Pathophysiology Location and Extent of GI Tract Involvement. Based on the location and extent of their inßammation, the majority of CD patients fall into one of three categories: inßammation of the small intestine only, inßammation of the large intestine only, or inßammation of both the small and large intestines. Figure 6 illustrates the regions of the GI tract within the small and large intestines that are most commonly affected by CD. Approximately 30% of CD patients present with disease limited to their small intestine (Stenson WF, 1999). Inßammation of the ileum—the region of the small intestine most often affected by CD—is referred to as ileitis. CD limited to the large intestine is called Crohn’s colitis and is found in approximately 25% of patients (Stenson WF, 1999); in these patients, the anorectal region is often affected as well. Approximately 40% of CD patients present with inßammation of both the small and large intestines simultaneously, which is referred to as ileocolitis (Stenson WF, 1999). Identifying disease localization is important in determining the best treatment regimen because therapies have specialized formulations to target release of the active drug in different areas throughout the small and large intestines. Crohn’s colitis is often treated

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FIGURE 6. Regions of the GI tract most commonly affected by Crohn’s disease.

similarly to UC, whereas CD that affects the small intestine can be treated quite differently with alternative drug formulations. CD may also affect the mouth, the tongue, the esophagus, the stomach, and the duodenum, but these clinical presentations of the disease are seen in only a very small percentage of patients. Complications of Crohn’s Disease Local Complications. Abscesses and Þstulas, two local complications of CD, arise when ulcers or Þssures extend through the entire intestinal wall into extraintestinal tissue. An abscess forms when intestinal contents leak from a Þssure and collect as a pocket in the peritoneal cavity. Abscesses, which occur in 15–20% of CD patients (Steinberg DM, 1973), are often accompanied by fever and abdominal pain. Resection of the affected portion of the intestine is often necessary to treat an abscess effectively because simple drainage of the collected ßuid may be inadequate and result in development of a Þstula. Fistulas develop when Þssures from one loop of intestine extend through to another epithelial surface, such as another loop of intestine, the bladder, the vagina, or even the skin (Figure 7). Fistulas develop in 20–40% of patients with CD (Steinberg DM, 1973). If a Þstula is small, pharmacological treatment alone may be sufÞcient to close it. Large or multiple Þstulas may require surgery if they are associated with intractable symptoms such as fever or pain.

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FIGURE 7. Fistulas in Crohn’s disease.

Other local complications found in CD patients are intestinal obstructions (especially in the small intestine) and toxic megacolon—a potentially lifethreatening, acute dilation of the colon. Acute inßammation of the intestinal layers, scarring from previous bouts of inßammation, adhesions in the small intestine, and impaction of digested material in a narrowed portion of the intestine can all lead to development of an intestinal obstruction. Patients with intestinal obstruction develop severe cramping pain in the midabdomen that is often accompanied by vomiting and abdominal distension. Physicians initially treat obstructions caused by inßammation with pharmacotherapy, but surgery is required if the obstruction is severe, resists pharmacotherapy, or recurs frequently. Surgery may also be required to treat toxic megacolon, which, if not rapidly improved by pharmacological treatment, may result in potentially fatal colon perforation (rupture). Systemic Complications. CD patients may suffer from several systemic complications, including fever, fatigue, malnutrition, and weight loss. Patients may also suffer from arthritis, dermatological conditions, inßammation of the eyes (typically uveitis), and liver dysfunction, but these complications are more common in CD patients with Crohn’s colitis than in patients whose disease is limited to the small intestine. The cause of these extracolonic complications is not known,

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but they may be linked to the abnormal immune system responses characteristic of CD. Most patients with CD experience weight loss, in part a consequence of diminished food intake and malabsorption of nutrients. Patients who suffer from CD may eat less because of diminished appetite or to avoid the abdominal pain that may follow a meal. Also, inßammation of the small intestine compromises patients’ ability to fully digest and absorb nutrients from food. In addition to weight loss, CD patients usually have moderate anemia (stemming from iron, vitamin B12, or folate deÞciencies) and gallstones and kidney stones (caused by bile salt malabsorption). Malnutrition may also put a CD patient at risk for loss of bone mass. CD-associated arthritis typically affects joints in the hands, elbows, ankles, knees, and lower spine, but unlike rheumatoid arthritis (which is more severe), CD-associated arthritis usually does not cause destructive changes to the affected joints. Ankylosing spondylitis—a rarer form of arthritis that affects the spine—is also seen in CD patients. Dermatological conditions, such as erythema nodosum (a disorder characterized by tender, red nodules), aphthous stomatitis (canker sores), and the more severe disorder, pyoderma gangrenosum (deep, chronic skin ulcers), also occur in CD patients. Other systemic complications associated with CD include inßammation of the liver and bile ducts (including pericholangitis and primary sclerosing cholangitis [PSC]). Except for spinal arthritis, ankylosing spondylitis, and PSC, all of these conditions tend to subside in parallel with the bowel disease activity. Risk of Colon Cancer. The risk of colon cancer in CD patients is less clearly understood than the well-established risk in UC patients, but studies suggest that CD patients with extensive Crohn’s colitis have an 18-fold higher risk of developing colon cancer than the general population (Gillen CD, 1994). The incidence of colon cancer in Crohn’s colitis patients increases with the amount of time that patients have suffered from the disease (Stenson WF, 1999). Colonic malignancies usually occur in areas of active inßammation, but they may also appear in regions of the large intestine that are unaffected by CD. CURRENT THERAPIES Because the etiology and pathophysiology of Crohn’s disease (CD) are not well elucidated, pharmacological therapies used to treat the disease tend to be general immunosuppressive drugs (e.g., aminosalicylates, corticosteroids, immunosuppressants) that inhibit the inßammatory process, but currently few agents with speciÞc targets exist. The exception is the tumor necrosis factor-alpha (TNF-α) inhibitor inßiximab (Centocor [a Johnson & Johnson subsidiary]/Schering-Plough/Tanabe Seiyaku’s Remicade)—the only biological agent currently approved for treatment of CD. Treatment of CD involves two main types of therapies: acute therapies designed to induce remission during ßares of disease activity and maintenance

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therapies designed to maintain long-term remission and prevent disease ßareups. In addition, therapies are used to induce the closure and healing of Þstulas, a manifestation of CD that is notoriously difÞcult to treat. Most current CD therapies are not actually approved for the treatment of CD; many of these drugs are indicated for ulcerative colitis (UC), a related inßammatory bowel disease, and are used off label for CD. Delivery of the active metabolite of many CD drugs (e.g., aminosalicylates, corticosteroids) to speciÞc regions of the small and large intestines is important to maximize their therapeutic beneÞt and minimize systemic side effects. Therefore, these agents are available in a variety of rectal formulations for local delivery and in special oral formulations associated with carrier molecules, semipermeable membranes, or pH-sensitive resins to help target the drug. Table 2 summarizes the leading therapies available to treat CD. In 2004, the top-selling CD products were inßiximab, the ethylcellulose-coated and enteric-coated formulations of mesalamine, and the immunosuppressants 6-mercaptopurine and azathioprine. TNF-α Inhibitors Overview. The most recent advance in the treatment of CD was the introduction of inhibitors of TNF-α, a potent inßammatory cytokine. TNF-α is currently the most promising target for treatment of CD, particularly in patients with Þstulizing disease. Despite the superior efÞcacy of these agents compared with other CD therapies, TNF-α inhibitors are typically reserved for CD patients with severe, refractory, or steroid-dependent disease because of their relatively high cost, risk of immunomodulatory side effects, and inconvenient parenteral administration. Not all TNF-α inhibitors have demonstrated efÞcacy in the treatment of CD, however. Etanercept (Amgen/Wyeth/Takeda’s Enbrel) failed to meet its primary efÞcacy endpoint (induction of clinical response or remission at week 4) in a study of 43 CD patients with moderate-to-severe active disease (Sandborn WJ, 2001). This failure probably resulted from etanercept’s inability to induce the apoptosis of intestinal T cells—a capability that appears to be critical for TNF-α inhibitor effectiveness in the treatment of CD (ten Hove T, 2002; van den Brande JMH, 2003; Shen C, 2004). Mechanism of Action. Receptors for TNF-α are found on the surface of most cells, including mononuclear cells in the gastrointestinal (GI) tract. Cleavage of the membrane-bound TNF receptors yields soluble TNF receptors that retain ligand-binding ability but cannot activate cells. TNF-α inhibitors reduce free, bioactive TNF-α by emulating the physiological role played by soluble TNF receptors. This action modulates the amount of circulating, bioactive TNF-α by binding to the cytokine before it can activate cell-surface receptors on mononuclear cells (Figure 8). Some TNF-α inhibitors can also induce the apoptosis of T cells in the intestinal mucosa, a capability that appears to be critical for efÞcacy in the treatment of CD (ten Hove T, 2002; van den Brande JMH, 2003; Shen C,

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TABLE 2. Current Therapies Used for Crohn’s Disease Agent TNF-α inhibitors Infliximab

Company/Brand Centocor/Schering-Plough/Tanabe Seiyaku’s Remicade

Oral aminosalicylates EthylcelluloseShire/Ferring/Nisshin/Kyorin Seiyaku’s coated Pentasa mesalamine Enteric-coated Procter & Gamble/Giuliani’s Asacol, mesalamine GlaxoSmithKline/Faes/Merckle’s Claversal, Falk/Provalis’ Salofalk, others Sulfasalazine Pfizer’s Azulfidine, Pfizer/Mitsubishi Pharma’s Salazopyrin Balsalazide Salix’s Colazal, Menarini’s Balzide, Shire’s Colazide Olsalazine Pfizer/UCB’s Dipentum Mesalamine

Solvay’s Rowasa

Immunosuppressants 6-mercaptopurine GlaxoSmithKline/Teva’s Purinethol, generics Azathioprine GlaxoSmithKline/Prometheus Product’s Imuran/Imurel/Imurek, Tanabe Seiyaku’s Azanin, generics Methotrexate Stada/Wyeth’s Rheumatrex, Santen Seiyaku’s Metolate, generics Cyclosporine Novartis’s Sandimmune/Neoral, generics Oral corticosteroids Budesonide Prednisone

Antibiotics Ciprofloxacin

AstraZeneca’s Entocort, generics Pfizer’s Deltasone, Sanofi-Aventis’s Cortancyl, Merck’s Decortin, generics

Bayer’s Cipro/Ciflox/Ciprobay/Ciproxin, generics Metronidazole Aventis/Shionogi’s Flagyl, Sandoz’s Metrolyl, generics Topical aminosalicylates Enteric-coated Procter & Gamble/Giuliani’s Asacol, mesalamine GlaxoSmithKline/Faes/Merckle’s Claversal, Falk/Provalis’ Salofalk, others EthylcelluloseFerring/Kyorin Seiyaku/Nisshin Pharma’s coated Pentasa mesalamine Mesalamine Solvay’s Rowasa, Axcan Scandipharm’s Canasa

Daily Dosea Acute: 5 mg/kg (one infusion at wks 0, 2, 6) Mt: 5 mg/kg (starting at wk 14, infusion every 8 wks) Acute: 3 g Mt: 1.5 g Acute: 3 g Mt: 1.5 g Acute: 4 g Mt: 2 g Acute: 6 g Mt: 3 g Acute: 2 g Mt: 1 g Acute: 3 g Mt: 1.5 g Mt: 50–100 mg Mt: 175 mg

Acute: 25 mg/wk Mt: 15–25 mg/wk Acute (IV): 250–300 mg Mt (oral): 50–100 mg Acute: 9 mg Acute: 1 mg/kg for 2 wks, then tapered by 10 mg/wk for 6 wks Acute: 500–1,000 mg Acute: 0.6–2 g

Acute: 1–4 g Mt: 1–4 g, 3 times per wk Acute: 1 g Mt: 1 g, 3 times per wk Acute: 1 g Mt: 1 g, 3 times per wk

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TABLE 2. (continued) Agent IV corticosteroids Hydrocortisone

Topical corticosteroids Hydrocortisone

Budesonide

Company/Brand

Daily Dosea

Pfizer’s SoluCortef, Sanofi-Aventis’s Hydrocort, Merck’s Hydrocortone, generics

Acute: 100 mg

Paddock’s Colocort, Schwarz Pharma’s Cortifoam, GlaxoSmithKline/Ferrer’s Colifoam, generics AstraZeneca’s Entocort

Acute: 100 mg

Acute: 2 mg

a Doses listed in the table are daily amounts unless otherwise specified.

IV = Intravenous; Mt = Maintenance; wk(s) = week(s).

2004). Because TNF-α plays an important role in Þghting infections and eradicating neoplastic cells, its suppression is not without hazard, particularly as a long-term strategy. Concerns have been raised as to whether chronic immunosuppression leads to opportunistic infections, malignancies, and other complications (Brown SL, 2002; Lee JH, 2002; Nahar IK, 2003). Infliximab. Inßiximab (Centocor [a Johnson & Johnson subsidiary]/ScheringPlough/Tanabe Seiyaku’s Remicade) is the Þrst biological agent approved for CD. It was Þrst approved in the United States in 1998 and then in Europe in 1999 to induce CD remission. The drug received orphan drug status in Japan early

FIGURE 8. Blockade of inflammation by TNF-α inhibitors (infliximab): proposed mechanism.

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in 2002 and launched later that year for CD remission induction. In June 2002, the FDA approved inßiximab for maintenance therapy in patients with moderateto-severe CD, and in April 2003, it approved the product for maintaining CD Þstula closure. In May 2003, inßiximab was approved for maintenance dosing in Europe in patients with severe active CD, and in October 2003, the European Commission approved inßiximab for maintenance dosing for sustaining clinical response in patients with Þstulizing CD. Phase III trials are ongoing in Japan to evaluate the drug’s potential as a maintenance therapy. Inßiximab is also marketed in the United States and Europe for rheumatoid arthritis (RA), psoriatic arthritis, and ankylosing spondylitis, and in Japan for RA. Inßiximab received FDA approval for UC in September 2005 and is in Phase III development for UC in Europe. In July 2003, Tanabe Seiyaku Þled a new drug application (NDA) in Japan seeking approval of inßiximab for the additional indication of Behcet disease (a chronic, recurrent disease characterized by ulceration of multiple sites, including the mouth, genitalia, eyes, and veins). Inßiximab is also in Phase III development for psoriasis in the United States and Europe. Inßiximab is a chimeric monoclonal antibody (MAb) composed of 75% human and 25% mouse protein. The drug exerts its therapeutic effect by binding soluble and membrane-bound TNF-α (Figure 8). The mouse portion of the antibody contains a TNF-α binding site sufÞcient for sequestering the cytokine, and the human portion is responsible for effector function, including immunoglobulin G (IgG)stimulated elimination of TNF-α-positive cells by both antibody-dependent cellular cytotoxicity and complement-dependent mechanisms (Feldmann M, 2001). Inßiximab is capable of inducing T-cell apoptosis, probably by cross-linking TNF-α on the surface of these cells (ten Hove T, 2002; van den Brande JMH, 2003). The ability to induce apoptosis appears to be critical for efÞcacy in the treatment of CD. Inßiximab may also inhibit the expression and activation of cell adhesion molecules (CAMs) on endothelial cells and leukocytes, thereby preventing leukocyte migration into the bowel mucosa. Inßiximab is administered as an intravenous infusion by a health care professional. The recommended dosing frequency for induction of remission is three infusions, administered in weeks 0, 2, and 6 and every eight weeks thereafter for maintenance of remission. The ability of inßiximab to induce a clinical response and remission in active CD was demonstrated in a double-blind, placebo-controlled trial of 108 patients with moderate-to-severe CD (Crohn’s Disease Activity Index [CDAI] between 220 and 400). Patients were randomized to receive a single infusion of placebo or inßiximab at 5, 10, or 20 mg/kg; stable doses of concomitant CD therapies (prednisone, 6-mercaptopurine [6-MP], azathioprine, or oral aminosalicylates) were permitted. The trial met its primary end point: a signiÞcantly greater number of inßiximab-treated patients than placebo patients achieving a clinical response as measured by a 70 or greater point reduction in the CDAI score at 4 weeks. Only 17% of the placebo group achieved a clinical response at week 4, but 81% of the 5 mg/kg group, 50% of the 10 mg/kg group, and 64% of the 20 mg/kg

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group met this end point. Additionally, at week 4, 33% of inßiximab-treated patients achieved remission, deÞned as a CDAI score of less than 150; only 4% of placebo-treated patients achieved remission. SigniÞcant differences in the response rates persisted at 12 weeks (41% of inßiximab patients versus 12% of placebo patients) but not in the remission rates (30% versus 18%) (Targan SR, 1997). Inßiximab’s ability to maintain CD remission was demonstrated in ACCENT I (A Crohn’s Disease Clinical Trial Evaluating Inßiximab in a New Long-Term Treatment Regimen). In this Centocor-sponsored trial, conducted in North America, Europe, and Israel, all 573 patients enrolled received a 5 mg/kg infusion at week 0. At week 2, responses were measured, and 335 responders (58%) were randomized to one of three treatment groups: • • •

Placebo injections at weeks 2 and 6 and then every eight weeks thereafter until week 46 (group 1). Inßiximab 5 mg/kg injections at weeks 2 and 6 and then every eight weeks thereafter until week 46 (group 2). Inßiximab 5 mg/kg injections at weeks 2 and 6 followed by 10 mg/kg injections every eight weeks until week 46 (group 3).

Patients were allowed to continue stable doses of concurrent CD therapies, including azathioprine, 6-MP, corticosteroids, and aminosalicylates, during the course of the trial. At week 30, 39% of group 2 and 45% of group 3 patients were in remission, compared with 21% in group 1 (placebo). The median time to relapse throughout the 46-week trial was longer for both inßiximab-treated groups—38 weeks and 54 weeks for groups 2 and 3, respectively, compared with 19 weeks for group 1 (Hanauer SB, 2002). Because Þstulizing CD is notoriously challenging to treat and control pharmacologically, inßiximab’s efÞcacy in reducing the number of draining Þstulas is a much-needed addition to the CD armamentarium. In a randomized, doubleblind, placebo-controlled study, 94 patients with either abdominal or perianal Þstulas that existed for the three months preceding the trial were randomized to inßiximab 5 mg/kg, inßiximab 10 mg/kg, or placebo. Each group received three injections during the Þrst six weeks of the study, and patients were followed for up to 26 weeks. Closure of 50% or more of Þstulas (the primary end point) was achieved in 68% and 56% of the 5 mg/kg and 10 mg/kg inßiximab groups, respectively, compared with 26% of the placebo group. Complete Þstula closure was reported in 55% and 38% of the 5 mg/kg and 10 mg/kg inßiximab groups, respectively, compared with 13% of the placebo group (Present DH, 1999). The ACCENT II study, sponsored by Centocor in North America, Europe, and Israel, found similar, albeit diminishing, results in the treatment of Þstulizing CD over 54 weeks. Induction regimens consisting of three infusions of inßiximab were given to the 306 patients enrolled, all of whom had Þstulizing CD. Of the 282 patients remaining in the study at week 14, 69% had responded with

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a 50% or greater reduction in the number of draining Þstulas and were randomized to maintenance therapy with inßiximab (5 mg/kg every eight weeks) or placebo. Patients were allowed to continue stable doses of concurrent CD therapies (e.g., azathioprine, 6-MP, corticosteroids, aminosalicylates) during the course of the trial. The 31% of patients who did not demonstrate a response at week 14 were also randomized to either inßiximab maintenance therapy or placebo; these patients were included in the overall safety analysis at the end of the trial but not in the primary efÞcacy analysis. Of the 195 patients who demonstrated an initial response at week 14, 46% of those randomized to inßiximab maintenance therapy demonstrated a response at week 54, compared with 23% of those randomized to placebo. A complete response, deÞned by the absence of draining Þstulas, was maintained in 35% of inßiximab-treated patients, compared with 19% in the placebo group (Sands BE, 2004). Further analysis of the 282 patients completing the 54-week ACCENT II trial demonstrated that maintenance therapy with inßiximab signiÞcantly reduced hospitalizations and surgeries in patients with Þstulizing CD, compared with placebo (Lichtenstein GR, 2005[a]). Of the 139 trial participants who had at least a 50% reduction from baseline in the number of draining Þstulas at both weeks 10 and 14 and were randomized to receive 5 mg/kg inßiximab every eight weeks thereafter through week 46, only 8.6% required hospitalization, compared with 18.9% of the placebo-treated group (27/143). The mean number of all surgeries and procedures for responders in the inßiximab-treated group was 65, compared with 126 in the placebo group. Inßiximab maintenance treatment also signiÞcantly reduced the mean number of inpatient surgeries and procedures, the mean number of major surgeries per 100 patients, the mean number of hospitalization days per patient, and the mean number of CD-related hospitalizations. The reduction in the number of hospitalizations was similar in the United States and in other countries; no bias was introduced by differences in national health care systems. These results suggest that the high price of inßiximab maintenance therapy might be offset by the lower cost of CD-related hospitalizations and surgeries, but formal cost-effectiveness studies are needed to determine whether these savings make up for the expense of the agent and its associated adverse events. Adverse events associated with inßiximab are usually mild and include headache, nausea, rash, coughing, sinusitis, dyspnea, urticaria, hypotension, and upper respiratory tract infection. However, inßiximab is associated with potentially serious infusion reactions. In the clinical studies discussed in the preceding paragraphs, the overall frequency of any adverse event or serious adverse event was not statistically different in the placebo and inßiximab-treated groups. In the induction study, 60% of CD patients receiving placebo and 75% of patients receiving a single infusion of inßiximab had at least one adverse event (Targan SR, 1997). In the ACCENT I study, serious adverse events occurred in 29% of placebo-treated patients and in 28% and 22% of patients in the two inßiximabtreated groups (Hanauer SB, 2002). Similarly, of the 282 patients included in the safety analysis in ACCENT II, 14% of patients treated with inßiximab maintenance therapy and 23% of patients treated with placebo experienced serious

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adverse events. Serious infections occurred in 3% of inßiximab-treated patients and 6% of placebo-treated patients (Sands BE, 2004). However, infusion reactions did appear to occur more frequently in the inßiximab-treated groups: 23% and 19% versus 9% for placebo in the ACCENT I study (Hanauer SB, 2002). Because inßiximab is a chimeric MAb that contains murine (mouse) components, patients may develop antibodies against the drug. In the ACCENT I and ACCENT II trials, 14% and 17% of patients, respectively, tested positive for antibodies to inßiximab. An additional 46% of patients in ACCENT I and 52% of patients in ACCENT II had inconclusive antibody tests, owing to the presence of detectable inßiximab concentrations, which could compete with the detection of antibodies in the immunoassay (Hanauer SB, 2002; Sands BE, 2004). In each of these trials, the incidence of infusion reactions was higher in patients who developed antibodies to inßiximab than in patients who tested negative for such antibodies. A separate study involving 125 inßiximab-treated CD patients found that the presence of anti-inßiximab antibodies in patients correlated with a shorter duration of response (Baert F, 2003). Research has shown that continuous administration of inßiximab every eight weeks (as opposed to unscheduled, episodic treatment) lowers the incidence of anti-inßiximab antibody formation and leads to better treatment outcomes (Rutgeerts P, 2004[c]; Hanauer SB, 2004[b]). In one retrospective observational study, 37 CD patients received scheduled inßiximab maintenance therapy every eight weeks and 32 received episodic inßiximab treatment; study results demonstrated signiÞcantly lower rates of hospitalization, surgery, and permanent disability in the group receiving scheduled treatment (Williams JB, 2005). Prior to the initiation of inßiximab treatment, the rates of hospitalization (51% versus 47%) and surgery (62% versus 59%) were the same in the two groups (scheduled versus episodic, respectively). However, after inßiximab maintenance therapy for more than one year, the rate of hospitalization in the scheduled treatment group was only 14%, versus 59% for patients receiving episodic treatment, and the rate of surgery was 14% versus 53%. Furthermore, the rate of permanent disability was 3% for the scheduled treatment group and 28% for the episodic treatment group. Scheduled maintenance therapy, therefore, has signiÞcantly better treatment outcomes than episodic therapy. In fact, episodic maintenance therapy with inßiximab did not signiÞcantly improve treatment outcomes in this study, as evidenced by hospitalization and surgery rates that were the same before and after episodic treatment. These results strongly suggest that inßiximab maintenance therapy should be administered on a scheduled basis. Studies also suggest that coadministration of inßiximab with an immunosuppressant (e.g., azathioprine, 6-MP, methotrexate) lowers the incidence of antiinßiximab antibody formation, leading to better treatment outcomes (Sandborn WJ, 2002[a]; Baert F, 2003). In a May 17, 2004, press release, ScheringPlough and Centocor announced their intention to study the potential beneÞts of combination therapy with inßiximab and azathioprine in the SONIC (Study of Immunomodulatory Na¨õve Patients in Crohn’s Disease) trial. SONIC will compare the efÞcacy of this combination with the efÞcacy of either drug alone in

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patients with moderately to severely active CD who have not previously taken an immunosuppressant or biological agent for their disease. Steroid-free remission will be the trial’s primary end point. No information on the number of study participants or the length of the trial was included in the press release. A recent study that compared induction therapy with the combination of inßiximab and azathioprine to induction therapy with corticosteroids was presented at Digestive Disease Week in May 2005; results of this study suggest that the combination therapy was superior to corticosteroids in inducing remission and mucosal healing (Hommes D, 2005). The study involved 129 CD patients with active disease (CDAI >220) who had been diagnosed within the past four years and had not previously received corticosteroids, immunosuppressants, or biological agents. The primary end points were remission rates at 6 and 12 months for two treatment strategies: •

•

Group 1: Top-down strategy. Induction therapy was initiated with 5 mg/kg inßiximab at weeks 0, 2, and 6 in combination with 2–2.5 mg/kg azathioprine daily. Recurrence of disease activity was treated with inßiximab and/or methotrexate. Group 2: Step-up/bottom-up strategy. Induction therapy was initiated with 9 mg budesonide enema daily or 40 mg prednisone daily, tapered over eight weeks. Recurrence of disease activity was treated with corticosteroids; patients failing repeated corticosteroid therapy were given azathioprine. Patients failing azathioprine were permitted inßiximab therapy.

At six months, 74.5% of patients in group 1 were in steroid-free remission compared with 48.1% of patients in group 2—a statistically signiÞcant difference. In the second group, 32.7% of patients still needed corticosteroid therapy at 6 months and 18.8% at 12 months. The majority of patients in group 2 (67%) required azathioprine to achieve remission; 15% needed to resort to inßiximab for remission induction. Endoscopic evidence of mucosal healing was signiÞcantly greater in group 1 at two years, with absence of ulceration in 75% of the group 1 patients examined versus 21.4% of the group 2 patients examined. Fistulas developed in two group 2 patients and none of the group 1 patients. The occurrence of serious adverse events was similar in both groups. This study suggests that corticosteroids are not necessary in treatment of moderate-to-severe CD and that initiating induction therapy with inßiximab plus azathioprine may be a better treatment strategy than induction with corticosteroid therapy. Inßiximab’s long-term safety remains a concern for gastroenterologists. The long-term safety of inßiximab in CD is being prospectively studied using the Crohn’s Therapy, Resource, Evaluation and Assessment Tool (TREAT), a registry of CD patients from community and academic practices. Results presented at Digestive Disease Week in May 2005 for the 6,290 patients enrolled in this registry through August 2004 (3,179 receiving inßiximab and 3,111 receiving other CD therapies), with a mean follow-up of 1.7 years, demonstrate that inßiximab’s safety was similar to that of conventional immunosuppressants. Mortality

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in patients who received inßiximab was slightly lower than in patients who did not receive inßiximab: 0.41 per 100 patient-years versus 0.50 (RR = 0.83). The incidence of neoplasms and lymphomas in the two groups was very similar. The incidence of serious infections appeared to be slightly higher in patients who received inßiximab than in patients who did not receive inßiximab: 1.33 per 100 patient-years versus 0.70 (RR = 1.93). However, this increased risk was attributable to confounding factors, because inßiximab was not an independent predictor of serious infection when analyzed using logistic regression. Interestingly, prednisone was found to be an independent predictor of increased mortality and serious infection, and narcotics and moderate-to-severe CD were found to be independent predictors of serious infection (Lichtenstein GR, 2005[b]). Studies using the TREAT registry are ongoing. Despite accumulating data that inßiximab is relatively safe for long-term use, experts are still wary of the potential risks, which will limit its use as a CD maintenance therapy. Inßiximab’s package insert includes a black box warning noting an increased risk of developing tuberculosis and other opportunistic infections, and the agent has been contraindicated in patients with moderate to severe heart failure. Additionally, the product label was updated in July 2004 to include a warning about hematologic and neurological events associated with inßiximab use. The warning cautions physicians that cases of leukopenia, thrombocytopenia, pancytopenia, and central nervous system (CNS)-manifested systemic vasculitis—some of which have been fatal—have been reported in postmarketing experience. The product label for inßiximab was further updated in October 2004 to include a warning about the increased incidence of lymphoma in patients treated with TNF-α inhibitors, including inßiximab. According to the revised product label, 3 patients (1 with RA and 2 with CD) among 2,410 patients treated with inßiximab during controlled and open-label portions of RA and CD clinical trials developed lymphomas; this rate is approximately sixfold higher than the rate expected for the general population. In December 2004, an additional warning was added to inßiximab’s label about severe hepatic reactions, including acute liver failure, jaundice, hepatitis, and cholestasis, which have been reported in postmarketing data in patients receiving inßiximab. Some of these cases were fatal or necessitated liver transplantation. Inßiximab’s European patents are not expected to expire until 2014. Regulatory procedures for biosimilars (the European term for generic equivalents of biological agents) are not yet in place and will likely require at least limited indication-speciÞc clinical trials to conÞrm pharmacoequivalence, efÞcacy, and safety, thereby delaying the entry of biosimilars in Europe at least a year beyond the end of patent protection. In the United States, inßiximab is protected by method of use patents for the treatment of CD, RA, and neurological conditions; these patents will expire in 2014. However, these patents do not prevent the development of generic equivalents for inßiximab’s other approved indications (ankylosing spondylitis, psoriatic arthritis, and UC), which could then be used off label in CD and RA. However, regulatory procedures for the approval of follow-on biological agents in the United States have been slow in coming; an

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approval process is not likely to be in place for several years. Like European regulatory agencies, the FDA is likely to require additional clinical trials for follow-on biologics that will further delay the entry of generic equivalents. In Japan, the lack of regulatory guidelines and an approval procedure for biosimilars, together with low generics penetration, will make the entry of a generic equivalent of inßiximab unlikely. Oral Aminosalicylates Overview. The aminosalicylate class consists of agents that contain 5-aminosalicylic acid (5-ASA), one of the oldest anti-inßammatory compounds used in the treatment of inßammatory bowel diseases. In high doses, these drugs can induce remission in acute attacks of CD. In long-term use, lower doses of 5-ASA may be prescribed as a CD maintenance therapy, but its use in that capacity has not proved to be as effective as it is for ulcerative colitis (UC) maintenance therapy. There is mounting evidence from studies in UC patients that continuous therapy with oral or topical aminosalicylates offers some protection against development of colorectal cancer, which occurs at a higher rate in both UC and CD patients than in the general population. In fact, it is estimated that colorectal cancer accounts for 15% of all deaths related to these diseases (Munkholm P, 2003). One early study that compared 102 UC patients with colorectal cancer to 196 matched UC controls demonstrated that pharmacological therapy, especially sulfasalazine, lasting at least three months, was signiÞcantly associated with a protective effect (Pinczowski D, 1994). Another case control study in UC patients showed that continuous mesalamine therapy at doses greater than 1.2 g per day reduced the risk of colorectal cancer by 81% (Eaden J, 2000). A retrospective study of 175 UC patients found that only 3% of patients who took continuous, long-term sulfasalazine therapy developed colorectal cancer, compared with 31% of patients who were noncompliant or had their treatment stopped by a physician (Moody GA, 1996). A large epidemiological study of 18,969 patients with UC or CD demonstrated that regular 5-ASA users (based on prescriptions Þlled) had a lower risk of colorectal cancer than irregular users (van Staa TP, 2005). Furthermore, a recent meta-analysis of nine observational studies involving 1,932 UC patients concluded that 5-ASAs protect against colorectal cancer (Velayos FS, 2005). Mechanism of Action. Although 5-ASA formulations have been researched and used in CD treatment for decades, their precise mechanism of action remains unclear. Potential mechanisms that may account for the efÞcacy of 5-ASA include the following: •

Inhibition of leukotriene production. 5-ASAs inhibit cyclooxygenase (COX) enzymes and lipooxygenase (LO), thereby inhibiting the conversion of arachidonic acid to prostaglandins (PGs) and leukotrienes (LTs), respectively. PG and LT levels are elevated in the sera and mucosa of CD patients.

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Scavenging reactive oxygen metabolites (free radicals). In CD, these metabolites are generated by neutrophils and monocytes in the diseased mucosa. Inhibition of IL-1β and IFN-γ production. Both are proinßammatory cytokines involved in disease pathogenesis (Mahida YR, 1991).

In addition to the effects of 5-ASAs in reducing symptoms of CD, these agents may offer some protection against the development of colon cancer. The aminosalicylates are thought to confer this protection by increasing apoptosis in the intestine and inhibiting the proliferation of the colorectal mucosa (Bus PJ, 1999; Reinacher-Schick A, 2000). Formulation Because oral 5-ASA is rapidly absorbed in the stomach and jejunum (proximal small intestine), reformulation is necessary to enable the drug to reach the site of CD in the distal small intestine or colon. Therefore, mesalamine (mesalazine in Europe and Japan), the term for any 5-ASA that is not bound to another compound, is often attached to a carrier molecule, such as sulfapyridine, to deliver 5-ASA to the colon. Typically, carrier molecules are linked to 5-ASA via an azo bond, which is cleaved by the enzyme azoreductase, which is present in many intestinal bacteria species. This separation of the carrier molecule from the active 5-ASA moiety delivers the latter to the intestines, where it is absorbed and acetylated by intestinal epithelial cells. Because many side effects are associated with the sulfa moiety, second- and third-generation aminosalicylates were developed in formulations that deliver 5-ASA to the affected areas of the intestine without the use of a sulfa carrier molecule. Instead, these newer formulations of mesalamine are coated with semipermeable membranes or pH-sensitive resins that target the 5-ASA to the small intestine or colon. Although unbound, uncoated mesalamine is used occasionally in the treatment of CD, the specially formulated versions of mesalamine predominate and are therefore the agents discussed in greater detail in the following sections. Ethylcellulose-Coated Mesalamine. Ethylcellulose-coated mesalamine (Shire/Ferring/Nisshin/Kyorin Seiyaku’s Pentasa) is indicated for the induction of UC remission and for the treatment of mildly to moderately active UC. It is used off label for CD in all major-market countries except Spain. This prolonged-release formulation of mesalamine consists of 5-ASA granules coated with a semipermeable membrane that gradually dissolves in the intestines. Much of the 5-ASA is delivered to the ileum, but some active drug is delivered to the proximal colon as well. The ability of ethylcellulose-coated mesalamine to induce remission of active CD has been demonstrated in three major clinical trials. In the Þrst study, involving 67 patients, 40% of patients receiving 1.5 g Pentasa daily achieved remission at 16 weeks, compared with 30% of the placebo group (Rasmussen SN, 1987).

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The second study, involving 40 patients, had similar results: 40% of patients receiving 1.5 g Pentasa daily achieved remission at 6 weeks, compared with 35% of the placebo group (Mahida YR, 1990). The third and most rigorous study was a double-blind, multicenter trial in which 310 patients with active CD were randomized to receive placebo or Pentasa at a dose of 1, 2, or 4 g/day for 16 weeks. Only the highest dose of Pentasa proved to be more effective than placebo; 64% of patients treated with 4 g/day Pentasa achieved remission, versus 40% of the placebo group (Singleton JW, 1993). The efÞcacy of 5-ASAs in maintaining CD remission is not well documented; their use as a maintenance therapy is under debate. In two trials of 161 and 220 CD patients, Pentasa (3 g/day or 2 g/day) was unable to signiÞcantly improve remission rates over placebo at 12–24 months (Bondesen S, 1991; Gendre JP, 1993). Because it is a sulfa-free 5-ASA, ethylcellulose-coated mesalamine is a common option for the majority of patients who are intolerant of sulfasalazine. However, ethylcellulose-coated mesalamine is not without side effects, the most common being diarrhea, headache, nausea, abdominal pain, dyspepsia, vomiting, and rash. The drug is contraindicated in patients with impaired hepatic or renal function. Ethylcellulose-coated mesalamine has already lost patent protection. The product is designed to deliver mesalamine—an agent that is also off patent—to a targeted site within the GI tract using a specially formulated, semi-permeable membrane coating. This delivery mechanism gives the agent a unique pharmacokinetic proÞle. Given the agent’s unique release proÞle and targeted site of action, regulatory authorities will likely require more rigorous clinical data (e.g., dissolution, bioequivalence, and controlled comparative clinical trials) before they deem a generic oral 5-ASA equivalent to the branded version. Enteric-Coated Mesalamine. Enteric-coated mesalamine formulations (Procter & Gamble/Giuliani’s Asacol, Norgine’s Fivasa, GlaxoSmithKline/Faes/Merckle’s Claversal, Falk/Provalis’s Salofalk, Astellas’ Asamax, Chiesi’s Asalex, Norvartis’ Mesalazine Sandoz/Mesalazina San, Sofar’s Pentacol, and ScheringPlough’s Lixacol) are indicated for treatment of mildly to moderately active UC and for maintenance of UC remission. They are used off label for CD in all major-market countries except Japan. These mesalamine formulations are coated with pH-sensitive acrylic resins. Mesalamine coated with resins that dissolve at pH 7 or higher (e.g., Asacol) delivers 5-ASA to the distal ileum and colon; mesalamine coated with resins that dissolve at pH 7 or lower (e.g., Claversal and Salofalk) delivers 5-ASA to the proximal ileum. Asacol has demonstrated the ability to induce remission in patients with mildly to moderately active CD in a placebo-controlled trial involving 38 patients. After 16 weeks of treatment, 60% of patients receiving 3.2 g/day Asacol were in remission, compared with 22% of the placebo group (Tremaine WJ, 1994). The ability of 5-ASAs to maintain CD remission is under debate, but Asacol has demonstrated signiÞcantly greater remission rates than placebo at 12 weeks

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COOH

N NHSO2

OH

N

N

FIGURE 9. Structure of sulfasalazine.

in clinical trials. In one study involving 83 patients, 66% of patient receiving Asacol (2.4 g/day) achieved remission, compared with 30% of patients receiving placebo (Caprilli R, 1994). Another study involving 125 patients showed similar remission rates: 66% for Asacol (2.4 g/day) versus 45% for placebo (Prantera C, 1992). Overall, the rate of adverse events in patients treated with enteric-coated mesalamine has been similar to the placebo rate in clinical trials. Reported side effects are typically mild: headache, pain, nausea, pharyngitis, dizziness, rash, dyspepsia, vomiting, and constipation. Caution is advised when using these drugs in patients with impaired renal function. Enteric-coated mesalamine has lost patent protection in the United Kingdom and is expected to lose patent protection in the United States in 2013 and in Italy in 2007. Like the specially formulated, ethylcellulose-coated mesalamine, the enteric-coated formulations of mesalamine have unique pharmacokinetic proÞles that will be difÞcult to reproduce in generic equivalents. Sulfasalazine. The Þrst 5-ASA developed and marketed, sulfasalazine (PÞzer’s AzulÞdine/Salozopyrine/Salazpyrina, PÞzer/Mitsubishi Pharma’s Salazopyrin, generics) (Figure 9), has been used to treat CD for decades. An enteric-coated, delayed-release version of sulfasalazine (PÞzer/Santen Seiyaku’s AzulÞdineEN/Salazopyrin-EN, SanoÞ-Aventis’s Colo-Pleon, generics) is also available. Although sulfasalazine is formally indicated for UC but not CD, the drug’s utility in treating CD was established in a trial during the 1970s (Anthonisen P, 1974). Sulfasalazine is more effective in CD patients with colonic and ileocecal disease than in patients with disease conÞned to the small intestine. It is modestly effective in reducing the recurrence rates of CD following surgery (Achkar JP, 2000). Sulfasalazine consists of the sulfonamide antibiotic sulfapyridine and 5-ASA joined via an azo bond. This bond is cleaved by azoreductases to deliver the active 5-ASA moiety primarily to the colon. Because sulfasalazine has been on the market for a relatively long time, many clinical trials examining its efÞcacy are nearly 30 years old. The Anthonisen study was the Þrst double-blind trial to demonstrate sulfasalazine’s efÞcacy in inducing remission in patients with active CD. The 17-week study of 31 patients found that in the subgroup of patients who had not undergone intestinal resection, response rates were higher in the sulfasalazine-treated group (53%) than in the placebo group (6%) (Anthonisen P, 1974). In the subgroup of patients who had

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undergone earlier intestinal resection, the difference in response rates between those treated with sulfasalazine and those treated with placebo was statistically insigniÞcant. Later studies of sulfasalazine found response rates ranging from 38% to 75%; rates for placebo ranged from 8% to 55% in all trials (Malchow H, 1984; Summers RW, 1979; Van Hees PA, 1981). Despite sulfasalazine’s demonstrated efÞcacy in the treatment of active CD, clinical trial data are contradictory about its utility in maintaining remission. Most recently, several large studies have found that the risks of relapse are comparable in patients treated with sulfasalazine and patients treated with placebo (Malchow H, 1984; Summers RW, 1979; Wenckert A, 1978). Sulfasalazine has a number of common side effects. According to the product labels for AzulÞdine and AzulÞdine-EN, the most common side effects, which occur in approximately one-third of patients, include headache, nausea, dyspepsia, and anorexia. Less common side effects are fever, rash, arthralgia, hemolysis, neutropenia, exacerbation of colitis, hypersensitivity reactions, and nephrotoxicity. CD patients with acetylation polymorphisms are more likely to suffer adverse events owing to the slower metabolism of the sulfapyridine moiety (Schroder H, 1972). Male infertility is a side effect that is fully reversible within three months of discontinuing therapy. Women who are pregnant or nursing may safely use sulfasalazine, but the importance of folic acid supplementation must be stressed because sulfasalazine lowers systemic levels of this vitamin, which is critical for fetal development. Balsalazide. Balsalazide (Salix’s Colazal, Menarini’s Balzide, Shire’s Colazide) is a relative newcomer in the 5-ASA drug class, launching for the treatment of UC in 1997 in the United Kingdom and in 2001 in the United States and Italy. Although prescription data suggest that balsalazide is being used to treat patients with CD, no available clinical data demonstrate balsalazide‘s efÞcacy in treating this disorder. Balsalazide consists of 5-ASA linked to the inert, nonabsorbable molecule 4-aminobenzoyl-β-alanine via an azo bond. As described previously, this bond is cleaved by azoreductases to deliver the active 5-ASA moiety in the distal ileum and proximal colon. Balsalazide’s side effects are similar to those associated with other oral 5-ASA products; they include headache, abdominal pain, nausea, diarrhea, and vomiting. Less common adverse events include respiratory infection, fever, rash, and itching. In rare cases, hepatotoxicity has been reported in patients taking balsalazide. Balsalazide is expected to lose patent protection in the United States, Italy, and the United Kingdom in 2006. Olsalazine. Olsalazine (PÞzer/UCB’s Dipentum) consists of two 5-ASA molecules joined via an azo bond. Like sulfasalazine and balsalazide, olsalazine’s mechanism of action is dependent on bacterial azoreductase present in the intestines to cleave the azo bond and release 5-ASA to the affected areas of the GI tract. Olsalazine launched in the United States in the early 1990s for the

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maintenance of remission in UC patients who are intolerant of sulfasalazine; it is used off label for the induction and maintenance of CD remission. Few data exist to justify the use of olsalazine in the acute treatment of active CD, and the drug’s efÞcacy in the maintenance of CD remission is controversial. Although earlier studies suggested that olsalazine may be useful in maintaining remission in CD, more recent studies suggest that this drug is not superior to placebo. In a 52-week, randomized, double-blind trial, 328 patients with quiescent CD (patients who had been in remission for at least one month prior to randomization) were treated with either olsalazine (2 g/day) or placebo. No statistically signiÞcant difference in the frequency of relapse was evident between olsalazinetreated patients (48.5%) and patients receiving placebo (45%) (Mahmud N, 2001). According to the product label for olsalazine, approximately 11% of patients taking olsalazine develop profuse, nonbloody diarrhea, which can be ameliorated by reducing the dosage. Other possible side effects include headache, rash, itching, fever, and nausea—effects similar to those of other oral 5-ASAs. The patent for olsalazine expired in Italy and the United Kingdom in 2001 and 2002, respectively. The drug is expected to lose patent protection in the United States, Germany, and France in 2004, 2004, and 2007, respectively. Immunosuppressants Overview. In recent years, immunosuppressants have increasingly become Þrstline maintenance therapy for moderate and severe cases of CD. For the most part, the immunosuppressants’ delayed onset of action makes them unsuitable for acute treatment of active disease, although they are typically initiated together with acute agents to give the immunosuppressants a chance to start working before the acute therapy is halted. None of the agents in this class is approved for CD, but their off-label use for this indication began following trial results that demonstrated their utility for the following purposes: • • • •

Achieving and maintaining remission in patients with refractory CD. Healing or reducing the drainage of perianal Þstulas. Reducing dependence on corticosteroids. Reducing the need for surgical treatment.

Mechanism of Action. Immunosuppressants target key immunological processes and molecules through a variety of mechanisms. Among these agents’ effects are blocking the activation of T lymphocytes, promoting the accumulation of anti-inßammatory molecules, and reducing the formation of antibodies. 6-Mercaptopurine. 6-MP (GlaxoSmithKline/Teva’s Purinethol, generics) (Figure 10) is a purine analogue that impairs nucleic acid synthesis and inhibits both humoral and cell-mediated immunity. The drug is indicated for remission induction and maintenance therapy for acute lymphatic (lymphocytic, lymphoblastic) leukemia but is used off label for CD. In July 2003, GlaxoSmithKline granted Teva rights to Purinethol in North America as part of a settlement between

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SH H N

N R

N

N

FIGURE 10. Structure of 6-mercaptopurine (R = H).

the two companies of patent litigation over the arthritis medication nabumetone (GlaxoSmithKline’s Relafen). GlaxoSmithKline and Teva will share gross proÞts from Purinethol sales; additional details of the agreement were not disclosed. Although its precise mechanism of action is unclear, 6-MP is known to inhibit T-helper lymphocyte function. Inactive by itself, 6-MP must be metabolized in the intestinal mucosa and liver into the active metabolites that are responsible for its cytotoxicity (antitumor activity) and immunosuppressant activity. The enzyme hypoxanthine guanine phosphoribosyl transferase (HGPRT) converts 6-MP into thiopurine nucleotides such as 6-thioguanine (6-TG), the active metabolite. Two other enzymes work competitively to produce inactive metabolites: xanthine oxidase converts 6-MP into 6-thiouric acid, while thiopurine methyl transferase (TPMT) converts 6-MP into 6-methylmercaptopurine (6-MMP). The efÞcacy of 6-MP versus placebo in CD was demonstrated more than two decades ago in a two-year, double-blind, crossover design trial involving 83 CD patients. Two-thirds of 6-MP-treated patients displayed a clinical response during the two years, compared with only 8% of patients on placebo. Fistula closure occurred in 31% of the 6-MP group compared with 6% of the placebo group (Present DH, 1980). Subsequent placebo-controlled and head-to-head trials have afÞrmed these results. Although it is a reasonably effective treatment for patients with refractory CD, response rates to 6-MP can vary greatly for several reasons. The drug has a slow onset of action and therefore requires several months of administration before its therapeutic effects become apparent. In the aforementioned study by D.H. Present, for instance, the mean time to response for 6-MP-treated patients was 3.1 months. Therefore, 6-MP is better suited as maintenance therapy for CD than for inducing remission in the acute setting. The efÞcacy and toxicity of 6-MP in some patients can also be inßuenced by the presence of a genetic polymorphism. Approximately 11% of the general population has low activity of the enzyme TPMT, which normally converts 6-MP into an inactive metabolite. In these patients, metabolism of 6-MP shifts toward accumulation of the active thiopurine nucleotide metabolites. Because these active metabolites are cytotoxic, increased levels put patients at higher risk of drug toxicity and leukopenia. According to the product label for Purinethol, coadministration of 6-MP with drugs that inhibit TPMT (e.g., aminosalicylates such as olsalazine, mesalazine, or sulfasalazine) may exacerbate this problem. Another 10% of the population has high TPMT activity, resulting in decreased levels of the active metabolites of 6-MP. These patients are less likely to suffer

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drug toxicities, but they are also receiving subtherapeutic doses of 6-TG (Cuffari CD, 2001; Kader HA, 2000). Approximately 15% of patients treated with 6-MP experience adverse reactions to the drug (Lamers CB, 1999). Side effects include fever, rash, nausea, and diarrhea, as well as more serious adverse events such as bone marrow toxicity (myelosuppression, leukopenia), infections, pancreatitis, and hepatitis. Therefore, patients taking 6-MP must be closely monitored, particularly in the weeks following initiation of therapy. Also, there is concern that long-term immunosuppression with 6-MP may be associated with an increased risk of malignancies; studies have yielded conßicting results thus far (Connell WR, 1994; Farrell RJ, 2000; Fraser AG, 2000). Although the patent for 6-MP expired more than three decades ago, the Þrst generic version of this drug became available in the United States only in March 2004. Azathioprine. Developed primarily to prevent tissue rejection in transplant surgery, azathioprine (GlaxoSmithKline/Prometheus Products’ Imuran/Imurel/Imurek, Tanabe Seiyaku’s Azanin, generics) (Figure 11) is a purine analogue that impairs nucleic acid synthesis and inhibits lymphocyte proliferation. It is approved for the management of severe, refractory RA and as an adjunctive therapy for the prevention of renal transplant rejection. Like 6-MP, azathioprine is used off label as a treatment for CD. Because of its long onset of action, azathioprine is better suited for maintenance therapy than for acute treatment to induce remission. Azathioprine is a pro-drug of 6-MP. After Þrst-pass metabolism, azathioprine is nonenzymatically converted into 6-MP (88%) and S-methyl-4-nitro-5thioimidazole (12%). From that point, the resulting 6-MP is metabolized via enzymatic processes described in the previous section. Azathioprine and 6-MP are used interchangeably; both agents reduce overall symptoms and steroid intake in patients with active CD. A randomized, placebo-controlled, double-blind study of azathioprine followed 63 patients with active CD. At baseline, all patients began a 12-week tapering dose of corticosteroids and were randomized to either azathioprine (2.5 mg/kg) or placebo. Remission rates at week 12 were equivalent, but after 15 months, there was a signiÞcantly greater remission rate for azathioprine (42%) than for placebo (7%) (Candy S, 1995). NO2 N S N N CH3 N

H N N

FIGURE 11. Structure of azathioprine.

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Azathioprine and 6-MP have been shown to signiÞcantly reduce the risk of relapse in CD patients for up to four years. In a long-term follow-up study, researchers followed 157 CD patients who were in clinical remission for at least six months and were treated with either azathioprine (n = 153) or 6-MP (n = 4) for more than six months (Bouhnik Y, 1996). The only concomitant CD medications allowed during the study were aminosalicylates. After one, three, and Þve years, the cumulative probability of relapse among the 157 patients was 11%, 22%, and 32%, respectively. In the study, 42 patients discontinued treatment with either azathioprine or 6-MP because of personal reasons, planned pregnancy, or side effects or because the treatment duration was deemed sufÞcient by a physician. In these patients, the cumulative risk of relapse was higher than in patients who continued azathioprine or 6-MP: 38%, 61%, and 75% at years one, three, and Þve, respectively. Results from this study also indicated that patients who continued azathioprine or 6-MP treatment were at a signiÞcantly lower risk of relapse during the Þrst four years of remission compared with patients who discontinued their study medication during this time. However, after four years of remission, there was no signiÞcant difference in relapse risk between patients continuing azathioprine or 6-MP treatment and patients who discontinued their medication. The study authors caution that because of the small number of study subjects, the lack of statistical difference after four years may result from insufÞcient statistical power. Azathioprine’s side effects are similar to those of 6-MP: fever, rash, nausea, and headache. Serious adverse events include leukopenia, pancreatitis, severe infections, and myelosuppression (bone marrow suppression). As with 6-MP, there is concern that long-term immunosuppression with azathioprine may be associated with an increased risk of malignancies, but clinical trial results are conßicting (Connell WR, 1994; Farrell RJ, 2000; Fraser AG, 2000). Methotrexate. Methotrexate (MTX; Stada/Wyeth’s Rheumatrex, Santen Seiyaku’s Metolate, generics) (Figure 12) is a folic acid analogue that interferes with DNA synthesis. The drug is indicated for several oncology indications and for severe, recalcitrant, disabling psoriasis and severe, active RA. Like other agents in this class, MTX is used off label for the treatment of CD, both in the acute and maintenance settings. Both orally administered and intramuscularly injected MTX are used in treating CD. Long-term, low-dose MTX therapy inhibits thymidylate, purine, and methionine production, resulting in accumulation of the potent anti-inßammatory

N COOH HOOCCH2CH2CHNHC O

N

N NCH2 CH3

NH2

N

NH2

FIGURE 12. Structure of methotrexate .

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molecule adenosine. MTX inhibits cellular proliferation and reduces the formation of antibodies and inßammatory mediators such as cytokines and eicosanoids. MTX has been shown to be effective in inducing remission in CD patients with chronically active, steroid-dependent disease. In a double-blind, placebocontrolled trial, 141 patients were randomized to receive MTX (25 mg/week intramuscularly) or placebo for 16 weeks. Patients received concomitant prednisone; the corticosteroid dose was tapered during the trial until the patient’s condition worsened. At the conclusion of the trial, 39% of patients in the MTX group were in clinical remission (deÞned as a Crohn’s Disease Activity Index [CDAI] score ≤ 150 points and the discontinuation of prednisone) compared with 19% of patients treated with placebo. Seventeen percent of MTX-treated patients withdrew from the study, compared with 2% in the placebo group. Side effects leading to withdrawal of MTX-treated patients included asymptomatic elevation of serum aminotransferase levels, nausea, skin rash, pneumonia, and optic neuritis (Feagan BG, 1995). MTX has also been shown to maintain remission over a period of 40 weeks in CD patients with chronically active disease. Seventy-six patients who previously entered remission following treatment with 25 mg intramuscular MTX weekly for 16–24 weeks were randomized to receive either MTX (15 mg/week intramuscularly) or placebo for 40 weeks. At the conclusion of the study, 65% of MTX recipients were in remission (CDAI score ≤ 150 points), compared with 39% of patients treated with placebo. During the course of the study, only 28% of patients treated with MTX required prednisone to treat a disease relapse (deÞned as an increase of ≥ 100 points over baseline in the CDAI), compared with 58% of placebo recipients. Of the MTX-treated patients, 40% experienced nausea and vomiting, 25% experienced symptoms of a cold, 18% experienced abdominal pain, and 18% experienced a headache. Corresponding adverse event rates for the placebo group were 25%, 28%, 25%, and 17%, respectively (Feagan BG, 2000). Although MTX is relatively well tolerated as a long-term therapy in CD, it is associated with some minor side effects, including nausea, vomiting, diarrhea, anorexia, stomatitis (inßammation of the oral mucosa), headache, skin rash, brittle nails, and alopecia. Major, less common side effects include pancytopenia, leukopenia, thrombocytopenia, irreversible and potentially lethal allergic pneumonitis, and liver Þbrosis. Product labeling, which includes a long list of black-box warnings of serious and potentially fatal side effects, recommends careful monitoring of hematology and renal and liver function every four to eight weeks in patients receiving long-term treatment. MTX is contraindicated in pregnant women. Cyclosporine. Cyclosporine (Novartis’ Sandimmune/Neoral, generics), a lipidsoluble polypeptide, is a strong immunosuppressant. Novartis reformulated the original cyclosporine product, Sandimmune, and developed Neoral, a microemulsion formulation that surpasses the original in bioavailability. Both Neoral and Sandimmune are indicated for prevention of organ rejection in patients

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receiving kidney, liver, or heart transplants; Neoral is also indicated for severe, active, methotrexate-refractory RA and for severe, recalcitrant plaque psoriasis in adult, non-immunocompromised patients. Although not approved for CD, both oral cyclosporine formulations and intravenous cyclosporine are used to treat patients with severe, steroid-refractory, and/or Þstulizing disease, typically as acute therapies to induce remission. A metabolite of the fungus Beauveria nivea, cyclosporine binds cyclophilin (an intracellular receptor) and selectively blocks the activation of T lymphocytes by inhibiting the calcium-dependent transcription of interleukin-2 (IL-2) and interferon-gamma (IFN-γ ) and, to a lesser degree, IL-3, IL-4, IL-5, TNF-α, and TNF-β. Oral and intravenous cyclosporine are generally as effective as other immunosuppressants for treating active CD, but oral cyclosporine’s long-term use in the maintenance of CD remission is not well supported by clinical trial data. Oneyear relapse rates were identical (20%) in the cyclosporine and placebo groups in one multicenter European trial involving 182 CD patients treated with concomitant corticosteroids (Stange EF, 1995). An 18-month Canadian study involving 305 patients found that CD symptoms worsened in more cyclosporine-treated patients (60%) than placebo-treated patients (52%) (Feagan BG, 1994). Like other immunosuppressant therapies, cyclosporine is associated with a number of side effects, some of them severe. The most commonly reported adverse events are paresthesias (abnormal sensations such as burning and tingling), hypertrichosis (hirsutism, or excessive hair growth), hypertension, tremor, renal insufÞciency, headache, and opportunistic infections. Both Neoral and Sandimmune’s labels contain black-box warnings cautioning physicians about the increased risk of infections and the potential risk of lymphoma. The microemulsion formulation of cyclosporine (Neoral) will lose its U.S. patent protection in 2009. Generic versions of Sandimmune are already available in the United States. Oral Corticosteroids Overview. Corticosteroids are among the most effective agents for inducing remission in acute CD attacks, but because they are associated with several signiÞcant side effects, their use as a CD maintenance therapy is typically reserved for steroid-dependent disease—severe cases in which patients cannot be weaned off steroids without resumption of disease activity. Prolonged therapy with corticosteroids can lead to adrenal atrophy, whereas abrupt cessation can cause adrenal insufÞciency, hypotension, and even death. Insomnia, night sweats, mood changes, and altered glucose metabolism may occur shortly after beginning corticosteroid use; prolonged use is associated with acne, abnormal fat deposition, and excessive hair growth. Some long-term side effects that most concern clinicians are osteoporosis, hypertension, and growth retardation, particularly in the case of pediatric CD patients. Corticosteroids can thwart healing of Þstulas and are therefore inappropriate for long-term therapy in patients with Þstulizing CD.

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Mechanism of Action. Corticosteroids exert their anti-inßammatory and immunosuppressive effects by regulating hormone-responsive gene expression. They inhibit the production of cytokines such as IL-1β, IL-2, and TNF-α, as well as leukotrienes. Corticosteroids also inhibit or reduce T-cell proliferation, monocyte and neutrophil migration into intestinal tissues, and total circulating lymphocyte levels. Formulation. Corticosteroids are available in many different formulations, so physicians can tailor corticosteroid therapy to the severity and extent of CD activity. The oral corticosteroids are the most convenient formulations and are most appropriate for CD activity located in the proximal and transverse colon. Budesonide. Budesonide (AstraZeneca’s Entocort) (Figure 13) is a secondgeneration corticosteroid that is available in an oral formulation and as an enema (discussed later in the section on topical corticosteroids). The oral formulation has a controlled-release mechanism that allows the agent to exert its effect when it reaches the ileum and ascending colon. Oral budesonide is approved for CD and is used in the acute setting to control active CD and induce remission. Because it has fewer side effects, budesonide can be used in CD patients who are intolerant of other corticosteroids. Comparative studies of budesonide and prednisolone have yielded conßicting results regarding budesonide’s comparative efÞcacy. In an eight-week study involving 176 participants, remission rates for patients treated with prednisolone and budesonide were 66% and 53%, respectively (Rutgeerts P, 1994). However, another study showed that over a 12-week period, budesonide resulted in a greater reduction in CDAI scores than prednisolone (Campieri M, 1997). Genetic studies have found that a human leukocyte antigen (HLA) gene is associated with budesonide treatment failure. Of 243 patients in one study, 17 expressed the HLA-DR8 genotype, and 13 of these 17 did not respond to budesonide therapy (Gelbmann CM, 2001). Further studies are needed to verify the link between genotype and budesonide treatment failure. Multiple trials have demonstrated that budesonide is associated with fewer adverse reactions than other corticosteroids (Campieri M, 1997; Rutgeerts P, 1994). Because budesonide is rapidly converted by the liver to inactive metabolites (primarily 16-α-hydroxyprednisolone and 6-β-hydroxybudesonide), only a OH O HO

O

H

O

O FIGURE 13. Structure of budesonide.

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low level (10–20%) of the active drug reaches the bloodstream, thus avoiding the many systemic side effects that occur when other corticosteroids circulate in high concentrations in the bloodstream. Replacement studies show that steroiddependent patients can taper prednisone and prednisolone therapy and incorporate budesonide treatment without increasing the risk of disease exacerbations (Cortot A, 2001). The patent that provided European protection for oral budesonide through 2010 was revoked in 2001 following opposition proceedings Þled by Falk and Ferring. Generic versions of the drug became available in Germany and the United Kingdom in 2004. Prednisone. Prednisone (PÞzer’s Deltasone, SanoÞ-Aventis’s Cortancyl, Merck’s Decortin, generics) (Figure 14) is one of the least expensive CD therapies and one of the most common oral corticosteroids prescribed for moderately acute CD, often for patients who do not respond to initial 5-ASA therapy. Patients typically receive two to four weeks of prednisone therapy and thereafter are tapered off the drug as quickly as possible without provoking a subsequent ßare-up. Because this class has been available for an extensive period of time, no highquality clinical studies pertaining to CD have been conducted in recent years. Therefore, this section does not cite speciÞc efÞcacy results from clinical studies for prednisone. It is well known that prolonged use of systemic corticosteroids—including prednisone—is associated with many serious side effects. (These serious side effects were discussed previously in the “Overview” section for the corticosteroid class.) Therefore, the beneÞts of prednisone must be weighed against the potential side effects when determining whether to use this agent to treat a CD patient on a long-term basis. Antibiotics Overview. Although not formally indicated for the treatment of CD, antibiotics have been used to treat CD patients for decades. Antibiotics are generally prescribed for CD with complicating abscesses or Þstulas and for postsurgical cases. Because long-term use of antibiotics can lead to development of resistance, antibiotics are typically prescribed only in the acute setting. The role of CH2OH C O

H3C

O OH

H3C

O FIGURE 14. Structure of prednisone.

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antibiotics in CD treatment is likely to increase; mounting evidence that bacteria play a role in the initiation and pathogenesis of CD is prompting physicians to prescribe antibiotics more frequently to “sterilize the gut” and thereby remove the inßammatory stimulus. (See “Etiology and Pathophysiology” for additional information on the possible role of bacteria in CD pathogenesis.) Mechanism of Action. Broad-spectrum antibiotic agents that are highly effective against anaerobic bacteria are most effective for CD, which suggests that they may sterilize microabscesses in inßamed tissue or alter gut ßora. However, not all broad-spectrum agents are equally effective, which suggests that an additional mechanism is at work in agents that are equally effective. In a study involving a murine model of CD, animals treated with a combination of ciproßoxacin and metronidazole (two antibiotics commonly used for CD; discussed in the following sections) displayed a reduced percentage of activated gut lymphocytes and reduced production of IFN-γ and TNF-α, compared with untreated control mice (Bamias G, 2002). Ciprofloxacin. One of the principal antibiotics used in the treatment of CD is ciproßoxacin (Bayer’s Cipro/Cißox/Ciprobay/Ciproxin, generics) (Figure 15). Ciproßoxacin is a broad-spectrum ßuoroquinolone approved for treatment of infections caused by susceptible bacteria. It is used off label to induce remission of active CD and in the treatment of Þstulizing disease. Despite the paucity of placebo-controlled data on ciproßoxacin’s efÞcacy in CD, comparative studies demonstrate that it is at least as effective as other antibiotic or anti-inßammatory therapies for inducing remission. In a retrospective Italian study, the remission rates achieved with ciproßoxacin alone, metronidazole alone, and combination therapy were similar: 69%, 73%, and 71%, respectively (Prantera C, 1998). A French trial that compared the efÞcacy of ciproßoxacin with oral ethylcellulose-coated mesalamine in 40 patients suggested that these agents are equivalent for treating mildly to moderately active CD. Remission rates at six weeks were 56% in patients receiving ciproßoxacin and 55% in patients receiving mesalamine (Colombel JF, 1999). For Þstulizing disease, ciproßoxacin in combination with inßiximab was shown to be more effective than inßiximab alone in one small, double-blind, placebo-controlled study. In this study, 22 CD patients with perianal Þstulas

FIGURE 15. Structure of ciprofloxacin.

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were randomized to receive 500 mg ciproßoxacin twice daily or placebo for 12 weeks. Inßiximab (5mg/kg) was administered to all patients at weeks 6, 8, and 12. Concomitant therapy (aminosalicylates, oral corticosteroids, azathioprine, or methotrexate) at stable doses was permitted. The primary end point was clinical response, deÞned as a 50% or greater reduction in the number of draining Þstulas at 18 weeks; 73% of ciproßoxacin-treated patients achieved this primary end point, compared with only 39% of the placebo group, and logistic regression analysis indicated better response in the ciproßoxacin group (West RL, 2004). Most side effects associated with ßuoroquinolone antibiotics are minimal, usually mild GI disturbances and CNS effects. Side effects associated with ciproßoxacin use in particular include nausea, vomiting, diarrhea, restlessness, rash, urticaria, painful oral mucosa, and, infrequently, tendon rupture. Ciproßoxacin has lost patent protection in the United States, Japan, and Europe, excluding Italy, where the ciproßoxacin patent will expire in 2009. Generic versions of Cipro were available in the United States, France, Germany, Spain, the United Kingdom, and Japan in 2004. Metronidazole. Metronidazole (Aventis/Shionogi’s Flagyl, Sandoz’s Metrolyl, generics) is a synthetic antiprotozoal and antibacterial agent indicated for infections in a wide variety of body systems. These dual actions have led to its use in healing CD Þstulas (in which anaerobes are suspected to play an important role) and in inducing remission in CD ßare-ups (of which bacterial activity is likely a component). More than two decades ago, metronidazole was shown to be effective in healing Þstulas in 10 of 18 (56%) patients with active CD (Bernstein LH, 1980). In a retrospective analysis of inpatient records for 233 Italian CD patients, researchers found 73% of metronidazole-treated patients achieved complete or partial remission (Prantera C, 1998). Metronidazole has also been shown to signiÞcantly improve CDAI scores, compared with placebo, in a 16-week study involving 99 CD patients (Sutherland L, 1991). However, only 56 patients completed the trial; 43 patients withdrew because of worsening disease, adverse events, or protocol violations. The most common side effect associated with metronidazole use is nausea, accompanied occasionally by headache, anorexia, and vomiting. Parasthesias are the major side effect of long-term therapy. Metallic taste, glossitis, stomatitis, urticaria, vaginal and urethral burning, dark urine, and reversible neutropenia may also occur. Additionally, the combination of metronidazole and alcohol can cause abdominal cramps, nausea, vomiting, headaches, and ßushing. Therefore, alcohol consumption should be avoided while taking metronidazole, and for at least one day following discontinuation of therapy. Topical Aminosalicylates Overview. As stated in the discussion of oral aminosalicylates, this class consists of agents that contain 5-ASA, one of the oldest anti-inßammatory compounds used in the treatment of inßammatory bowel diseases. Topical aminosalicylates, indicated for treatment of UC, do not appear to be as efÞcacious in CD

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as in UC. However, the agents are used in some cases of CD that are conÞned to the colon, often in combination with an oral aminosalicylate or other acute therapy. Because many patients consider topical formulations inconvenient, the topical 5-ASAs are used more frequently in the acute setting than in long-term maintenance of CD remission. Few data describe the efÞcacy and safety of topical aminosalicylates in CD patients. Studies have shown that rectal mesalamine formulations are moderately effective in CD; the response rate of these formulations in treating CD affecting the rectum is relatively slower than in treating UC, and healing is incomplete in 50% of the CD patients treated (Williams CN, 1990). Rectal mesalamine formulations can cause headaches as well as anal irritation and ßatulence in 5–10% of patients. Abrupt discontinuation of rectal mesalamine can occasionally trigger a severe relapse (rebound) response: development of pancolitis requiring subsequent colectomy (surgical removal of the colon). Patients who intend to stop rectal therapy are therefore counseled to begin oral 5-ASA therapy prior to step-down discontinuation of enema or suppository. Mechanism of Action. Although the 5-ASAs have been researched and used in CD treatment for decades, their precise mechanism of action remains unclear. They are thought to have numerous anti-inßammatory mechanisms, which are discussed in greater detail in the “Oral Aminosalicylates” section. As previously mentioned, these agents may offer some protection against the development of colon cancer, which occurs at a higher rate in patients with UC or CD. The aminosalicylates are thought to confer this protection by increasing apoptosis in the intestine and inhibiting the proliferation of the colorectal mucosa (Bus PJ, 1999; Reinacher-Schick A, 2000). Formulation. Topical 5-ASAs are available in many different rectal formulations, including enemas, suppositories, and rectal foams. These agents come as unbound, uncoated mesalamine, which can reach the proximal colon directly when administered rectally, or in formulations with special coatings that help target the release of the active 5-ASA to the site of disease. Enteric-Coated Mesalamine. Enteric-coated mesalamines are available as enemas (GlaxoSmithKline/Merckle’s Claversal, Giuliani’s Asacol, Chiesi’s Asalex, Astellas’s Asamax, Provalis’s Salofalk, So Se Pharm’s Mesaßor, generics), as suppositories (Procter & Gamble/Giuliani’s Asacol, Norgine’s Fivasa, GlaxoSmithKline/Faes/Merckle’s Claversal, Falk/Provalis’s Salofalk, Astellas’s Asamax, Chiesi’s Asalex, Stada’s Enterasin, generics), and as rectal foams (Procter & Gamble/Giuliani’s Asacol, Faes/Merckle’s Claversal, Sofar’s Pentacol, Chiesi’s Asalex, Provalis’s Salofalk). Rectal formulations of the entericcoated mesalamines are indicated for treatment of UC and used off label for CD. Although the foam preparations are easier for patients to use (and many patients prefer them), enemas and suppositories provide greater proximal penetration and are thought to be more effective.

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Ethylcellulose-Coated Mesalamine. Ethylcellulose-coated mesalamine is available as an enema (Ferring/Kyorin Seiyaku/Nisshin Pharma’s Pentasa) and as a suppository (Ferring’s Pentasa). Rectal formulations of ethylcellulose-coated mesalamine are indicated for treatment of UC and used off label for CD. Mesalamine. Uncoated mesalamine is available as an enema (Solvay’s Rowasa, generics) and as a suppository (Solvay’s Rowasa, Axcan Scandipharm’s Canasa). Rowasa is indicated for treatment of active, mild-to-moderate, distal UC, proctosigmoiditis, and proctitis (inßammation of the rectum). It is used off label for treatment of CD, particularly when the disease is conÞned to the distal colon. Uncoated mesalamine enemas will lose their patent protection in the United States and Europe in 2004 and 2007, respectively. Intravenous Corticosteroids Overview. As mentioned in the “Oral Corticosteroids” section, corticosteroids are among the most effective agents for inducing remission in acute CD attacks. However, their association with several signiÞcant side effects relegates their use primarily to short-term treatment of moderate-to-severe CD. Intravenous (IV) corticosteroids are typically employed in particularly severe, refractory cases that require hospitalization and are administered as a short course, generally ten days, to bring severe disease under control. Mechanism of Action. Corticosteroids exert their anti-inßammatory and immunosuppressive effects by regulating hormone-responsive gene expression. They inhibit the production of cytokines and leukotrienes and inhibit T-cell proliferation, monocyte and neutrophil migration into intestinal tissues, and total circulating lymphocyte levels. Formulation. Corticosteroids are available in many different formulations, so physicians can tailor corticosteroid therapy to the severity and extent of CD activity. IV formulations are used in patients with severe disease as a way to bypass the impaired GI absorption that these patients may exhibit. Hydrocortisone. IV hydrocortisone (PÞzer’s SoluCortef, SanoÞ-Aventis’ Hydrocort, Merck’s Hydrocortone, generics) is one of the most commonly used IV corticosteroids for patients with severe or refractory CD. Because hydrocortisone has been available for decades, no high-quality clinical studies pertaining to CD have been conducted in recent years; thus this section does not cite speciÞc efÞcacy results from clinical studies for hydrocortisone. The side effects of hydrocortisone are similar to side effects of other corticosteroids, discussed in the “Oral Corticosteroids” section. Topical Corticosteroids Overview. In an attempt to utilize the highly effective corticosteroids for CD treatment while minimizing their systemic side effects, rectally administered topical formulations of corticosteroids have been developed. These formulations are used for treatment of CD that is conÞned to the colon.

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Mechanism of Action. Corticosteroids exert their anti-inßammatory and immunosuppressive effects by regulating hormone-responsive gene expression. They inhibit the production of cytokines and leukotrienes and inhibit T-cell proliferation, monocyte and neutrophil migration into intestinal tissues, and total circulating lymphocyte levels. Formulation. Topical corticosteroids are available as suppositories, enemas, and rectal foams, which effectively treat CD activity in the rectosigmoid area of the colon while minimizing the doses needed and the side effects of the systemic corticosteroids. Hydrocortisone. Topical hydrocortisone is available as an enema (Paddock’s Colocort, generics) and as a rectal foam (Schwarz Pharma’s Cortifoam, GlaxoSmithKline/Ferrer’s Colifoam). These formulations of hydrocortisone are typically administered in short courses to induce remission in moderately active CD proctitis. Although the foam preparations of hydrocortisone and other corticosteroids are easier for patients to use, enemas provide greater proximal penetration and are more suitable for extensive CD activity. Because hydrocortisone has been available for a long time, no high-quality clinical studies pertaining to CD have been conducted in recent years. Therefore, this section does not cite speciÞc efÞcacy results from clinical studies for hydrocortisone. Budesonide. Budesonide (AstraZeneca’s Entocort) is a second-generation corticosteroid that is available as an enema and as an oral formulation (discussed in the “Oral Corticosteroids” section). Budesonide enema is approved for UC and is used off label in the acute setting to control active CD and induce CD remission. Because it has fewer side effects, budesonide can be used in CD patients who are intolerant of other corticosteroids. Studies suggest that the efÞcacy of budesonide is similar to that of other corticosteroids and have clearly demonstrated that budesonide is associated with fewer side effects (Rutgeerts P, 1994; Campieri M, 1997). Nonpharmacological Approaches Nonpharmacological approaches for CD include probiotics, nutritional therapies, and surgical procedures. Probiotics—dietary supplements containing live microorganisms thought to restore the beneÞcial bacteria normally present in the GI tract—are gaining popularity among gastroenterologists as an adjunct CD therapy. Nutritional therapies are generally reserved for the most severe cases, such as patients suffering from poor GI absorption and malnutrition, while surgery is appropriate for patients with acute, life-threatening complications of CD and/or patients with extensive CD activity that is refractory to all available drug therapies. Probiotics are an alternative to conventional pharmacotherapy for patients with mild CD who prefer to remain drug-free, and they are increasingly used as an adjunct therapy in the treatment of CD. One theory of the etiology of

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CD proposes that bacteria in the normal intestinal ßora trigger the aberrant and ampliÞed immune response seen in CD; therefore, probiotic products aim to alter the balance of “good” and “bad” bacteria in the intestines of CD patients. Probiotic foods, such as yogurt, and food supplements, often in powder and capsule form, are now sold in stores and health food markets. However, large-scale, highquality, randomized clinical trials are needed to clarify how probiotics work and to deÞnitively demonstrate the utility of these nonpharmacological agents in the induction and maintenance of CD remission. Nutrient absorption is generally normal in mild CD cases involving the terminal ileum, except for vitamin B12, which can be replaced via monthly injections. However, if more than two or three feet of the small intestine are diseased or surgically removed, malabsorption of fats, calcium, and vitamins A, E, and D can occur. In most markets under study, hospitalized patients who are severely malnourished and pediatric or adolescent CD patients who have failed to grow normally may receive one of the following nutritional therapies to replace otherwise lacking nutrients: •

•

•

Liquid elemental diets—formulated to meet all nutritional needs of CD patients—include concentrated nitrogen in the form of amino acids and are almost completely absorbed by the upper intestinal tract. Enteral nutrition (EN) is provided via a stomach feeding tube inserted through the nose. EN is absorbed directly by the GI tract and causes few side effects. Patients can remove the tube in the morning and resume normal daily activities. Total parenteral nutrition (TPN) provides nutrition intravenously through a superior vena cava catheter that cannot be removed and reinserted by the patient. TPN is generally administered for a short course (one month) or just prior to surgery. Some patients may receive long-term TPN, but 15% will experience hyperglycemia; copper, zinc, and fatty-acid deÞciencies; or serious IV line infections.

CD patients typically undergo emergency surgery because of complications, including excessive bleeding, bowel perforation, intestinal obstruction, and abscess formation. Elective surgery may be indicated for treatment-refractory patients who are experiencing severe pain, weight loss, fever, or extreme fatigue. Surgical procedures for CD are generally aimed at removing or bypassing the diseased areas of the small intestine, leaving the healthy portions intact: • •

Bowel resection involves removal of the diseased area of the small intestine, followed by anastomosis to connect the newly severed ends. An alternative to resection is bowel bypass surgery, which creates a diverted path between the healthy portions of the small intestine and the colon. Bypass operations, which are associated with complications in leftover diseased portions of intestine, are becoming less common, thanks to improvements in bowel resection techniques.

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•

537

A third type of surgery, total abdominal colectomy with ileorectal anastomosis, is used for patients with Crohn’s colitis. The entire colon is removed and the ileum is joined to the rectum. Because this procedure leaves the rectum intact (except when the rectum is diseased and unusable for anastomosis), CD patients do not generally require subsequent use of an ostomy bag. Another surgical procedure for CD is strictureplasty. In this procedure, surgeons use a balloon catheter to Þnd areas of stricture. The intestinal wall at that point is cut and resealed in a different orientation to allow free movement of intestinal contents. A newer version of strictureplasty uses an endoscope instead of a balloon catheter.

Because CD recurs postsurgically in as much as 55% of patients at 5 years and 76% at 15 years, regardless of procedure type, surgery is usually followed by aggressive pharmacological management with mesalamine, 6-MP, or azathioprine (Achkar JP, 2000). One possible complication of surgical CD treatments (particularly those that involve small bowel resection) is short bowel syndrome (SBS). Approximately half of the small intestine can be removed without ill effects for the patient, particularly when the ileum is left intact. Removal of the jejunum, for instance, results in lactose intolerance, but the ileum can compensate and absorb most important nutrients. However, patients with a severely diseased or resected ileum can suffer from SBS, which causes malabsorption of nutrients, particularly fats, bile salts, and some critical vitamins. Diarrhea, weight loss, fatigue, abdominal bloating, and fatty stools are common symptoms of SBS and can be ameliorated by use of a parenteral diet. EMERGING THERAPIES The etiology and pathophysiology of Crohn’s disease (CD) are not well understood. Theories about the cause of CD and its progression suggest that both immunodeÞciencies, particularly in the innate immune system, and an overactive adaptive immune response may be to blame. As a result, a wide range of therapies are in development for CD treatment whose mechanisms of action appear to be mutually exclusive—for example, immune suppressants and immune stimulators, cytokine inhibitors and recombinant cytokines, antibiotics and probiotics. However, all of these therapeutic approaches have demonstrated the potential to offer beneÞt in the treatment of the disease. Current research focuses primarily on the development of biological agents that target key inßammatory components, particularly tumor necrosis factor-alpha (TNF-α) and other proinßammatory cytokines. The only biological agent currently approved for treatment of CD is the TNF-α inhibitor inßiximab (Centocor [a Johnson & Johnson subsidiary]/Schering-Plough/Tanabe Seiyaku’s Remicade). Efforts are also under way to evaluate the potential of novel mechanisms of action in the treatment of CD, including immune stimulators, interleukin inhibitors, cell adhesion molecule (CAM) inhibitors, and inhibitors of the mitogen-activated protein (MAP) kinase signal transduction pathway. The most promising therapies in

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late-stage development are two TNF-α inhibitors, adalimumab (Abbott’s Humira) and CDP-870 (UCB’s Cimzia), which offer advantages over inßiximab, including decreased immunogenicity and a more convenient route of administration. These therapeutic approaches are discussed in detail in the following sections; Table 3 summarizes the drug therapies in development for CD. In addition to the therapies in development for CD discussed in the previous paragraph, Protein Design Labs (PDL) has been developing a humanized monoclonal antibody (MAb) against interferon-gamma (IFN-γ ), called fontolizumab or HuZAF. According to a PDL press release in March 2004, the drug failed to meet its primary end point of induction of a signiÞcant clinical response, compared with placebo, in two Phase II trials (HARMONY 1 and 2). In August 2005, Biogen Idec entered into collaboration with PDL to jointly develop, manufacture, and commercialize fontolizumab. It is unclear whether PDL and Biogen Idec will continue to pursue development of fontolizumab for CD after the disappointing efÞcacy results or seek instead to develop the drug for other autoimmune indications. This drug is not discussed in further detail here. TNF-α Inhibitors Overview. During active ßares of CD, neutrophils and monocytes migrate from the bloodstream into the intestinal mucosa and submucosa, where they secrete proinßammatory molecules, including cytokines and tissue-damaging free radicals. Inhibitors of TNF-α, a potent inßammatory cytokine, represent the most recent advance in the treatment of CD. Inßiximab, the Þrst anti-TNF-α agent approved for CD, was launched in 1998 in the United States, 1999 in Europe, and 2002 in Japan. Because of inßiximab’s relatively high cost, cumbersome route of delivery (IV), immunogenicity, and risk of immunomodulatory side effects, researchers have directed their attention toward the development of improved anti-TNF-α therapies for the treatment of CD. The TNF-α inhibitor etanercept (Amgen/Wyeth/Takeda’s Enbrel) has been examined in the treatment of CD but failed to meet its primary efÞcacy end point: induction of a clinical response or remission in CD patients with moderate-tosevere active disease (Sandborn WJ, 2001). This failure was probably the result of etanercept’s inability to induce the apoptosis of intestinal T cells—a capability that appears to be critical for TNF-α inhibitor effectiveness in the treatment of CD (ten Hove T, 2002; van den Brande JMH, 2003; Shen C, 2004). Phase II or Phase III trial results have been published for other TNF-α inhibitors, including Abbott’s adalimumab (Humira), UCB’s CDP-870 (Cimzia), and Serono’s onercept. Onercept is currently in Phase II development for CD in the United States and Europe. However, a randomized, placebo-controlled study of 207 patients presented at the United European Gastroenterology Week in September 2004 showed no signiÞcant beneÞt over placebo of the four onercept doses tested (Rutgeerts P, 2004[a]). Furthermore, development of onercept for psoriasis was discontinued because of two cases of sepsis, one of them fatal. The unfavorable Phase II efÞcacy results, questions about its safety, and the fact that the agent is no longer listed in the company’s pipeline make it unlikely that development

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TABLE 3. Emerging Therapies in Development for Crohn’s Disease Compound TNF-α inhibitors Adalimumab (Humira) United States Europe Japan

Development Phase

Marketing Company

III III —a

Abbott Laboratories Abbott Laboratories —

III III —

UCB UCB —

II II —

Abbott Laboratories Abbott Laboratories —

II — —

Synta Pharmaceuticals — —

Immune stimulators Sargramostim (Leukine) United States Europe Japan

III III —

Berlex Laboratories Berlex Laboratories —

Cell adhesion molecule inhibitors Natalizumab (Tysabri) United States Europe Japan

IIIb PRc —

Elan/Biogen Idec Elan/Biogen Idec —

MAP kinase inhibitors RDP-58 (delmitide) United States Europe Japan

— IId —

— Procter & Gamble —

II II —

Cytokine PharmaSciences Cytokine PharmaSciences —

CDP-870 (Cimzia) United States Europe Japan Interleukin inhibitors ABT-874 (formerly J-695) United States Europe Japan STA-5326 United States Europe Japan

Semapimod (formerly CNI-1493) United States Europe Japan

a Adalimumab is in Phase II development in Japan for the treatment of rheumatoid arthritis and psoriasis

under licensee Eisai Pharmaceuticals. b Natalizumab, which had been launched in the United States for multiple sclerosis (MS) in 2004, was

voluntarily withdrawn from U.S. markets and all clinical trials of the agent suspended by Biogen Idec and Elan in February 2005. Phase III U.S. clinical trials had been completed prior to this date. The decision to continue development of natalizumab for Crohn’s disease (CD) and to relaunch natalizumab for MS in the United States is pending. c The review of natalizumab for CD in Europe has been slowed because of the agent’s safety issues and is ongoing. d After disappointing Phase II clinical trial results for RDP-58 in CD, Procter & Gamble has decided to focus on development for ulcerative colitis; RDP-58’s continued development for CD remains uncertain. PR = Preregistered.

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of onercept for CD will continue. Therefore, only adalimumab and CDP-870 are discussed in detail in following sections. Mechanism of Action. Receptors for TNF-α are found on the surface of most cells, including mononuclear cells in the gastrointestinal (GI) tract. Cleavage of the membrane-bound TNF receptors yields soluble TNF receptors that retain ligand-binding ability but cannot activate cells. Agents in this class reduce free, bioactive TNF-α by emulating the physiological role played by soluble TNF receptors. This action modulates the amount of circulating, bioactive TNF-α by binding to the cytokine before it can activate cell-surface receptors on mononuclear cells. Some TNF-α inhibitors, such as inßiximab and adalimumab, can also induce the apoptosis of T cells in the intestinal mucosa, a capability that appears to be critical to TNF-α inhibitor effectiveness in the treatment of CD (ten Hove T, 2002; van den Brande JMH, 2003; Shen C, 2004). Adalimumab. The fully human anti-TNF-α monoclonal antibody (MAb) adalimumab (Humira) was discovered through a collaboration between Cambridge Antibody Technology (CAT) and Knoll Pharmaceuticals (acquired by Abbott Laboratories in 2001). Abbott owns exclusive worldwide rights to adalimumab and pays sales-based royalties to CAT. Abbott is developing adalimumab as a potential treatment for CD; Phase III trials are under way in the United States and Europe. Abbott launched adalimumab for the treatment of rheumatoid arthritis (RA) in the United States and Europe in 2003. Adalimumab also launched in 2005 in Europe for the treatment of psoriatic arthritis and is preregistered in the United States for this indication. In Japan, adalimumab is in Phase II development for RA and psoriasis with licensee Eisai Pharmaceuticals, but no development for CD has been reported yet. Adalimumab, whose mechanism of action is similar to that of inßiximab, blocks the activity of TNF-α and induces T-cell apoptosis (Shen C, 2004). Unlike inßiximab, which is a chimeric monoclonal antibody (MAb) composed of 75% human and 25% mouse (murine) protein, adalimumab is a fully human, antiTNF-α MAb. According to the inßiximab and adalimumab product labels, the frequency of development of neutralizing antibodies against these two agents is respectively 10% and 5%. Therefore, adalimumab is expected to be less immunogenic than inßiximab, which may result in a lower frequency of infusion reactions and fewer problems with loss of efÞcacy. Another important characteristic that differentiates adalimumab from inßiximab is the agent’s route of administration. Inßiximab must be administered by a health care professional as an IV infusion, whereas adalimumab can be self-administered by a patient as a subcutaneous (SC) injection every other week. Adalimumab’s ability to induce CD remission was demonstrated in the Phase III CLASSIC (Clinical Assessment of Adalimumab Safety and EfÞcacy Studied as an Induction Therapy in Crohn’s) trial presented at the 2004 Digestive Disease Week conference (Hanauer SB, 2004[a]). This double-blind study involved 299 patients with moderately to severely active CD (Crohn’s Disease Activity

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Index [CDAI] 220–450) who were naive to anti-TNF-α therapy. Patients were randomized to receive SC injections at weeks 0 and 2 of either placebo or one of three adalimumab regimens at the following loading and maintenance dosages: 160 mg/80 mg, 80 mg/40 mg, or 40 mg/20 mg. At week 4, 30% of patients in the two higher-dose adalimumab treatment groups achieved remission (deÞned as a CDAI score less than 150), compared with 12% of placebo recipients; this comparison, which served as the study’s primary end point, produced a statistically signiÞcant difference. The highest rate of remission induction (36%) was achieved by the 160 mg/80 mg adalimumab group. Patients in this group also achieved the highest rate of clinical response—49% achieved a 100-point or greater reduction in CDAI score and 58% achieved a 70-point or greater reduction, compared with 23% and 35%, respectively, of patients treated with placebo. These differences were statistically signiÞcant. Scant safety data were reported, but all doses of adalimumab were said to be well tolerated, and there was no difference in the incidence of adverse events between the four groups. Mild injection site reactions were the most common adverse event in this study. Only one adalimumab-treated patient tested positive for antibodies to the agent during the four weeks of the study. An uncontrolled, open-label extension of the CLASSIC trial, referred to as CLASSIC II, was presented at the 2005 Digestive Disease Week conference in Chicago, Illinois (Sandborn WJ, 2005[a]). Two hundred and twenty patients who were not in remission (deÞned as a CDAI score of less than 150) at either week 0 or 4 of the CLASSIC I study were given 40 mg adalimumab SC every other week. Dose escalation to 40 mg weekly was allowed for ßares or persistent nonresponse. After six months, 156 patients (71%) remained on adalimumab (40 mg every other week or weekly), and 63% of these patients did not require dose escalation. The rate of clinical remission at 24 weeks was 33% (73/220); clinical response rates were 78% (122/156) for achieving a 70-point or greater reduction in the CDAI score and 70% (109/156) for achieving a 100-point or greater reduction. Adverse events, not necessarily drug-related, occurred in approximately 40% of patients; 10% of patients experienced serious adverse events, and 10% withdrew from the study because of adverse events. Five infections were observed in four patients during the six-month study period. The 55 patients who were in remission (deÞned as a CDAI score of less than 150) at both week 0 and week 4 in the CLASSIC I trial are also being studied to assess adalimumab’s efÞcacy in maintenance of remission in CD (Sandborn WJ, 2005[a]). These patients were randomized to receive placebo, 40 mg adalimumab every other week, or 40 mg adalimumab weekly for up to one year. Results from this study have not yet been released. Adalimumab also appears to be efÞcacious and safe in CD patients previously treated with inßiximab who had either stopped responding or became intolerant of the drug. In a Phase III, open-label, uncontrolled study, a total of 24 patients were treated with an initial dose of 80 mg adalimumab at week 0 and then a 40 mg dose of adalimumab every other week for 12 weeks. Beginning at week 4, dosing could be increased to weekly 40 mg SC injections of adalimumab

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if necessary. At the conclusion of the 12-week treatment period, 59% of the 17 patients who had a baseline CDAI score of 220 or higher achieved a clinical response (deÞned by a 100-point or greater decline in CDAI score), while 29% achieved clinical remission (deÞned by a CDAI score of 150 points or lower). Of the nine patients who had draining Þstulas at baseline, Þve experienced a 50% or greater decline in the total number, and three patients experienced complete Þstula closure. Adalimumab was well tolerated in this study, and none of the study participants experienced hypersensitivity reactions to adalimumab, even among the 20 patients who had acute and delayed hypersensitivity reactions with inßiximab (Sandborn WJ, 2004[c]). As previously mentioned, adalimumab is marketed for RA, and its product label contains warnings similar to those in the inßiximab label. Adalimumab has a black box warning cautioning physicians and patients about the risk of tuberculosis infection. The warning recommends that patients be tested—and possibly treated—for latent tuberculosis infection prior to receiving adalimumab. The product label for adalimumab also reports that malignancies have been observed in recipients of TNF-α inhibitors, including adalimumab. According to the product label, 0.4% of the 2,468 RA patients treated with adalimumab for a median of 24 months during clinical studies developed lymphomas; this occurrence rate is higher than the rate in the general population, but not signiÞcantly higher than the rate in people with RA. In 2004, warnings were added to adalimumab’s label about hypersensitivity reactions, hematologic events, and use in patients with congestive heart failure. Adalimumab offers several potential advantages over inßiximab in the treatment of CD, including decreased immunogenicity and a more convenient subcutaneous route of administration. Currently available data suggest that the efÞcacy of adalimumab is similar to inßiximab’s efÞcacy in inducing a clinical response or remission in CD patients (see “Current Therapies” for additional information on inßiximab’s efÞcacy). This similarity bodes well for adalimumab’s potential in the CD market. However, no data have been published yet on adalimumab’s ability to maintain remission, and larger-scale, placebo-controlled trials over a longer study period are needed to verify adalimumab’s ability to treat Þstulizing disease. Adalimumab will likely be used initially in CD patients who are intolerant of or have failed inßiximab therapy, supported by the study showing adalimumab’s efÞcacy in patients who stopped responding to or were intolerant of inßiximab. Adalimumab may also challenge inßiximab as the Þrst-line TNF-α therapy for CD. CDP-870. Two Phase III trials of CDP-870 (certolizumab pegol; UCB’s Cimzia) for the induction and maintenance of a clinical response in CD have been completed in the United States and Europe. In an October 2005 press release, UCB announced its plans for regulatory submissions in both regions during the Þrst quarter of 2006. No development has been reported in Japan. This drug is also in Phase III development as a potential treatment for RA.

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CDP-870 was being developed by Celltech Group, but the company was acquired by UCB in 2004. Previously, Celltech had been codeveloping CDP-870 for RA with PÞzer. However, in December 2003, PÞzer terminated its rights to CDP-870 (prior to the completion of Phase III RA trials); PÞzer cited as its reason for termination of the agreement Celltech’s refusal to renegotiate the Þnancial terms originally established between Celltech and Pharmacia (now PÞzer) in March 2001. In May 2005, Lonza entered into an agreement with UCB to manufacture CDP-870. Using a proprietary technology, CDP-870 can be manufactured in Escherichia coli at a cost much lower than the cost of creating an antibody or receptor fusion product by using mammalian cell culture. Thus, CDP-870’s lower cost of production may translate into a price lower than inßiximab’s price. Such a cost advantage would be an important distinguishing feature for CDP-870, considering the high level of price sensitivity in the markets under study and the fact that CDP-870 will face stiff competition in the RA market, where there are already three well-established TNF-α inhibitors (etanercept, inßiximab, and adalimumab). CDP-870 is a humanized anti-TNF-α MAb fragment conjugated to two polyethylene glycol (PEG) subunits that add stability and increase the agent’s half-life. Because CDP-870 is humanized, it may be less immunogenic than the chimeric inßiximab. CDP-870 binds TNF-α with high afÞnity, thereby blocking its ability to activate the inßammatory cascade. CDP-870 lacks an Fc region, so this agent is unable to activate complement or to lyse cells through antibodydependent cell cytotoxicity, which inßiximab and adalimumab are able to do; it is not known whether those mechanisms of action are important for efÞcacy in CD. It is also not known whether CDP-870 is, like inßiximab and adalimumab, capable of inducing T-cell apoptosis, which is recognized as important in the treatment of CD. CDP-870 is formulated as a lyophilized powder for reconstitution, which can then be administered intravenously or as an SC injection using a 23-gauge needle, according to a company representative from Celltech (now UCB). Therefore, CDP-870 can be self-administered by the patient; if the patient is unwilling or unable to self-administer the SC injection, the drug can be administered intravenously by a health care professional. The ßexibility of CDP-870’s dosing and capability of being self-administered by SC injection gives it an advantage over inßiximab’s IV dosing. Furthermore, CDP-870’s monthly dosing schedule offers an advantage over the other subcutaneous TNF-α inhibitor in development for CD, adalimumab, which is administered every other week. UCB is conducting the Pegylated Antibody Fragment Evaluation in Crohn’s Disease: Safety and EfÞcacy (PRECiSE) program involving four studies to investigate CDP-870’s potential to treat CD. PRECiSE-1 and PRECiSE-2 are Phase III, double-blind, placebo-controlled studies in a total of 1,330 CD patients with moderately to severely active disease over a 26-week period. A July 2005 press release from UCB announced that the PRECiSE-1 trial met its primary efÞcacy end points with statistical signiÞcance, but analysis is ongoing and detailed results have yet

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to be released. Results from PRECiSE-2 demonstrating that CDP-870 met the primary end points with statistical signiÞcance were reported in an October 2005 press release from UCB and presented at the United European Gastroenterology Week in Copenhagen, Denmark (Schreiber S, 2005) (see detailed discussion of PRECiSE-2 results in the next paragraph). UCB stated that CDP-870 was well tolerated in these two studies, but the company will further assess the agent’s tolerability and long-term safety in the PRECiSE-3 and PRECiSE-4 trials. These studies are 24-month, open-label extensions of PRECiSE-1 and PRECiSE-2 and are currently ongoing. CDP-870’s ability to maintain a clinical response (100 point or more decrease in the CDAI score) in moderate-to-severe CD was demonstrated in the Phase III PRECiSE-2 trial (UCB press release, October 2005; Schreiber S, 2005). In the study, 668 patients were treated with 400 mg CDP-870 at weeks 0, 2, and 4; 428 patients (64%) achieved a clinical response after this induction regimen and were randomized to receive placebo or CDP-870 every four weeks for up to 24 weeks. At week 26, 62.8% of patients in the CDP-870 group maintained a clinical response compared with only 36.2% of the placebo group, a statistically signiÞcant difference. The percentage of patients in clinical remission at week 26 also reached statistical signiÞcance; 47.9% of CDP-870-treated patients versus 28.6% of placebo-treated patients. Based on Phase II trial results (discussed in the next two paragraphs), which found higher response rates in patients with elevated C-reactive protein (CRP; a marker for inßammation), PRECiSE-2 evaluated CDP870’s efÞcacy in patients based on CRP levels (Lorenzo A, 2004). However, the primary end points in the PRECiSE-2 trial were met regardless of the patients’ levels of CRP. Phase II trial results were reported in a Celltech company press release in February 2002 and presented at the 2003 Digestive Disease Week conference (Schreiber S, 2003). The dose-ranging, placebo-controlled study assessed the product’s efÞcacy and safety in 292 CD patients with an initial CDAI score of 220–450 points. Patients were treated with monthly SC injections of placebo or 100, 200, or 400 mg CDP-870 at weeks 0, 4, and 8. Clinical response was assessed every two weeks through week 12 using the CDAI; the primary end point was the percentage of patients achieving a 100-point or greater decline in CDAI score or a CDAI score of 150 points or lower (indicating remission) at week 12. The best response rate, 52.8%, was observed with the highest dose at week 10; this result was statistically signiÞcant compared with placebo (30.1%). However, the week 12 response rates did not reach statistical signiÞcance: 44.4% of patients receiving 400 mg CDP-870 compared with 35.6% of placebo recipients. Although it failed its primary efÞcacy end point, CDP-870 was well tolerated, and no difference in the incidence of adverse events was observed between the drugtreated and placebo groups. Additional analysis of the Phase II results revealed that signiÞcantly higher response rates to CDP-870 were found in patients who had elevated levels of CRP prior to being treated with the drug (Schreiber S, 2003). In the subset of 118 patients with a baseline CRP greater than or equal to 10 mg/L, statistically signiÞcant

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differences in response rates between placebo recipients and the 400 mg CDP-870 group were found at all time points. The highest response rates to CDP-870 were found for the highest dose at weeks 6 and 10: 67.7% and 64.5%, compared with 28.6% and 17.9% for placebo, respectively. An IV formulation of CDP-870 also demonstrated efÞcacy in CD patients (Winter TA, 2004). In a randomized, double-blind, placebo-controlled Phase II study, 92 patients with active CD were administered a single infusion of CDP870 (1.25, 5, 10, or 20 mg/kg) or placebo. The trial’s primary endpoint was the percentage of patients achieving a clinical response (deÞned as a 100-point or greater decline in CDAI score) or remission (deÞned as a CDAI score of 150 points or lower) at week 4, although efÞcacy was also measured at weeks 2, 8, and 12. Compared with placebo, treatment with CDP-870 was not associated with statistically signiÞcant differences in response rates. However, 47.1% of patients receiving 10 mg/kg CDP-870 achieved remission at week 2 compared with 16% of placebo recipients, a statistically signiÞcant difference. This difference did not persist at week 4. In total, 22 patients withdrew from the study as a result of disease progression, non-improvement, or adverse events. Adverse events, the majority of them mild to moderate in nature, were reported by 43 patients receiving CDP-870 and 15 placebo-treated patients. The most common adverse events reported in the treatment groups were headache, CD aggravation, urinary tract infection, abdominal pain, fever, and nausea. CDP-870 may offer several advantages over inßiximab, such as a more convenient formulation, decreased immunogenicity, and an anticipated lower price. Phase III results from the PRECiSE-1 and PRECiSE-2 trials, which met their primary end points, suggest that CDP-870’s efÞcacy in treating CD is similar to that of inßiximab and adalimumab. Although CDP-870 failed to meet its primary efÞcacy end points in two Phase II trials, many agents in development for CD have had difÞculty meeting primary efÞcacy endpoints because of high placebo response rates, and analysis of subgroups with high CRP levels, presumably patients with the most active disease, has demonstrated signiÞcant efÞcacy over placebo. Nonetheless, CDP-870’s effect on Þstula closure will need to be evaluated if it is going to compete with inßiximab for the treatment of Þstulizing CD. Furthermore, the agent’s uptake may be hindered by its lack of long-term safety data, compared with inßiximab (used in treatment of CD, RA, and other indications for the past seven years) and adalimumab (used in the treatment of RA for two years). Interleukin Inhibitors Overview. The identiÞcation of TNF-α as a key inßammatory mediator for CD and the subsequent launch of the TNF-α inhibitor inßiximab represented a major advance in the treatment of CD. However, numerous cytokines other than TNF-α, including interleukins, are important immune and inßammatory mediators and are also thought to be involved in the pathogenesis of CD. Companies have begun to explore interleukins as therapeutic targets; some of them appear to offer

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promising novel approaches to treatment of CD. Inhibitors of interleukin (IL)12—including Abbott’s injectable MAb ABT-874 (formerly J-695) and Synta’s oral, small-molecule inhibitor STA-5326—are currently in Phase II development for CD with at least preliminary evidence of efÞcacy in the induction of clinical response or remission. In May 2005, Centocor started enrollment for a Phase II CD clinical trial of its MAb that inhibits both IL-12 and IL-23 activity, CNTO1275. Chugai/Roche’s Actemra (tocilizumab; formerly atlizumab, MRA), an IL-6 inhibitor that launched in June 2005 in Japan for Castleman’s disease, is in Phase II development in Japan for CD, but no clinical trial data have been released on this drug’s efÞcacy in treating CD. The two IL-12 inhibitors are discussed in greater detail in following sections. Mechanism of Action. Interleukins are cytokines that have many functions in immune and inßammatory responses. They are secreted predominantly by leukocytes; their primary function is to exert immunoregulatory effects on other leukocytes, such as stimulating the differentiation and proliferation of other immune cells. Research has shown that many interleukins, such as IL-1β, IL-2, IL-6, IL-8, IL-12, and IL-18, are upregulated in the intestinal mucosa of CD patients. Drug compounds in this class inhibit the activity of a particular interleukin, thereby blocking its ability to exert its physiological role in the immune response. ABT-874. Abbott Laboratories is developing ABT-874 (formerly J-695), a fully human MAb against IL-12, as a potential treatment for CD. Phase II trials are under way in the United States and the United Kingdom. No development has been reported for this drug in Japan. ABT-874 is also in Phase II development for treatment of multiple sclerosis (MS) and RA in the United States and Europe. ABT-874 was identiÞed through a collaboration between Knoll Pharmaceuticals (acquired by Abbott in 2001) and CAT. The drug was licensed by CAT to Knoll in the same 1995 agreement that gave Abbott rights to adalimumab (discussed previously). CAT receives milestone payments from Abbott and will receive sales-based royalties if ABT-874 is brought to market. Researchers believe that a cell-mediated immune response predominates in CD. As discussed in “Etiology and Pathophysiology,” the inßammatory response associated with CD is believed to be driven by the activation of T-helper 1 (TH1) cells. IL-12 plays a key role in the differentiation of TH cells into TH1 cells, and IL-12 production appears to be increased in the intestinal tissue of CD patients (Monteleone G, 1997; Parronchi P, 1997). Thus, ABT-874, which is designed to neutralize the activity of IL-12, acts to inhibit TH1 cell differentiation and the resulting disease pathogenesis. Like some of the TNF-α inhibitors, IL-12 inhibitors can also induce apoptosis of T cells in the intestinal mucosa, and this activity may prove to be critical in achieving an effective response in the treatment of CD (Cominelli F, 2004). Although no studies have looked speciÞcally at ABT874’s ability to induce T-cell apoptosis, the fact that it is a bivalent MAb, like inßiximab and adalimumab, suggests that it may have this capability.

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Phase II clinical trial results have demonstrated signiÞcantly greater response rates in ABT-874-treated CD patients, compared with placebo (Mannon PJ, 2004). This double-blind study involved 79 patients with active CD (CDAI score of 220–450) randomized to placebo (n = 16) or to one of two cohorts treated with seven weekly subcutaneous injections of either 1 mg/kg or 3 mg/kg ABT-874 (n = 63). Patients in Cohort 1 received their Þrst and second injections four weeks apart; patients in Cohort 2 experienced no interruption between the administration of the Þrst and second injections. Concomitant CD medications, including corticosteroids, aminosalicylates, and antibiotics, were permitted. Safety was the primary end point; rates of clinical response (deÞned as a 100point or greater decline in CDAI score) and remission (deÞned as a CDAI score lower than 150 points) were secondary end points. The highest response and remission rates were in the group of patients receiving 3 mg/kg ABT-874 in Cohort 2 (n = 16). The response rate of this group was signiÞcantly higher than the response rate of the placebo group after the 7 weeks of treatment (75% versus 25%, respectively), but not at the end of the 18-week follow-up (69% versus 25%). The percentage of patients in this group who achieved remission did not differ signiÞcantly from the percentage in the placebo group, although there was a trend toward higher remission rates in the drug-treated group: 38% versus 0% at both time points. Response and remission rates for all patients in Cohort 1 and for patients in Cohort 2 who were treated with 1 mg/kg ABT-874 did not differ signiÞcantly from the rates for the placebo-treated group. ABT-874 was well tolerated in this Phase II study (Mannon PJ, 2004). Local injection site reactions, most of them mild in nature, were the most frequently reported adverse events and the only events that occurred more frequently in drug-treated patients than in the placebo group (range of 77% to 88% versus 25%, respectively). Other adverse events observed at similar frequencies in the drug-treated and placebo groups were nausea, vomiting, abdominal pain, urinary tract infections, cough, fever, headache, and fatigue. No serious infections were observed. The development of antibodies against ABT-874 was detected in three of the drug-treated patients, a frequency of approximately 5%. Although the efÞcacy results for the group treated with 3 mg/kg injections of ABT-874 for seven consecutive weeks are encouraging, further studies are needed to evaluate ABT-874’s ability to induce response and remission—likely at the 3 mg/kg dose—in a larger patient population. Abbott will likely try to position adalimumab as the Þrst-line TNF-α inhibitor in the treatment of CD, followed by ABT-874 if adalimumab should fail. The ability of ABT-874 to maintain remission and to treat Þstulizing disease would also need to be assessed if the agent is to compete with inßiximab. STA-5326. Synta Pharmaceuticals is developing STA-5326, a speciÞc, smallmolecule inhibitor of IL-12, as a potential treatment for CD. Phase II trials are under way in the United States. Based on promising results from a recently completed Phase IIa trial (see following details), Synta announced in a September 2005 press release that it has initiated a large, randomized, double-blind, placebocontrolled Phase IIb trial. No development has been reported for this drug in

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Europe or Japan. STA-5326 is also in Phase II development for treatment of psoriasis and in early development for MS and RA in the United States. As mentioned previously, IL-12 plays a key role in the differentiation of TH cells into TH1 cells that predominate in the inßammatory response in CD, and IL-12 production appears to be increased in the intestinal tissue of CD patients (Monteleone G, 1997; Parronchi P, 1997). STA-5326 works by inhibiting the transcription of IL-12 but is unlikely to have any additional effect on the induction of T-cell apoptosis in the intestinal mucosa, which may be important in the treatment of CD. In an open-label, dose-ranging Phase IIa study presented at Digestive Disease Week in May 2005, STA-5326 was shown to be generally well tolerated, with potential clinical activity in the treatment of CD (Burakoff R, 2005). This study involved 73 patients with moderate-to-severe active CD (CDAI 220–450); approximately 12 patients were assigned to each of the Þve dose groups, which ranged from 14 mg twice daily to 70 mg daily. Concomitant administration of other CD therapies was permitted. Clinical response (deÞned as a 70-point or 100-point or greater decline in CDAI score) and remission (deÞned as a CDAI score lower than 150 points) were assessed after four weeks of treatment. The group receiving 35 mg STA-5326 daily had the highest response rates (82% for a 70-point or greater decline in the CDAI score, 64% for a 100-point or greater decline in the CDAI score) and the highest remission rates (36%). The most common adverse events reported were dizziness, nausea, headache, and fatigue; seven patients discontinued treatment because of side effects. STA-5326 is the Þrst orally administered small molecule that speciÞcally inhibits IL-12 production. If this drug proves to be efÞcacious in the treatment of CD, its oral daily dosing will give it a signiÞcant advantage over the injectable cytokine inhibitors currently available or in development. Immune Stimulators Overview. One theory of the etiology of CD is that antigenic stimulation within the GI tract of CD patients triggers an aberrant immune response that is prolonged and ampliÞed. Pharmacological therapies currently used to treat CD generally aim to downregulate the production or activity of inßammatory mediators to minimize this aberrant immune response. However, predisposition to the development of CD has been linked to mutations in genes that are important in the innate immune response—for example, NOD2 and toll-like receptors (TLRs) (Ogura Y, 2001; Hugot JP, 2001). (See “Etiology and Pathophysiology” for additional information on these genes and their association with CD.) Researchers have also observed that patients with rare immunodeÞciency diseases (e.g., granulomatous disease; glycogen storage disease Ib; leukocyte adhesion deÞciency; congenital, cyclic, and autoimmune neutropenia) have GI manifestations that resemble those of CD (Couper R, 1991; Dieckgraefe BK, 2002[a]). These Þndings suggest that immune defects, particularly in innate immunity, may contribute to CD’s etiology and pathophysiology by allowing harmless antigens and the normal bacterial

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ßora to penetrate the GI membrane, triggering an aberrant immune response and development of the overactive, chronic adaptive immune response seen in the GI tract of CD patients. Therefore, immune stimulators might offer a new approach to the treatment of CD (Dieckgraefe BK, 2002[b]; Folwaczny C, 2003; Wilk JN, 2002). One immune stimulator—sargramostim (Berlex Laboratories’ Leukine)—has reached late-stage clinical trials for CD. Mechanism of Action. Agents in this class aim to counter defects in innate immunity by stimulating and strengthening a patient’s ability to mount an immune response to pathogens and disease. One way to bolster a patient’s innate immunity is the administration of the exogenous recombinant growth factors involved in hematopoiesis (the formation, differentiation, and proliferation of red and white blood cells from stem cells). Administration of such recombinant growth factors can increase the number of leukocytes (e.g., neutrophils, monocytes, macrophages), which play a vital role in immunological processes, and may help to maintain intestinal integrity by boosting mucosal defenses. Sargramostim. Berlex Laboratories, a subsidiary of Schering AG, is developing sargramostim (Leukine) as a potential therapy for moderate-to-severe CD. Phase III studies are under way in the United States and Europe. The drug has been marketed in the United States since 1991 and is used to accelerate myeloid recovery in patients undergoing autologous and allogenic bone marrow transplantation for a variety of oncology conditions. Schering AG acquired sargramostim from Immunex in July 2002, when Immunex divested the drug upon merging with Amgen. Sargramostim is a yeast-derived, recombinant form of granulocyte-macrophage colony-stimulating factor (GM-CSF), a growth factor that plays an important role in the formation of nonlymphoid blood cells (e.g., erythrocytes, monocytes, neutrophils, eosinophils, basophils, and megakaryocytes). Thus, sargramostim induces the development of cells that play a critical role in immune responses. This activity may help to maintain the GI immune barrier and minimize aberrant immune responses against harmless antigens or normal bacterial ßora, which are thought to be an underlying cause of CD. Berlex is conducting the New Opportunities to Verify Evolving Logic in Crohn’s Disease (NOVEL) program to investigate sargramostim’s potential to treat CD. The Phase II NOVEL 1 trial (discussed later) has been completed. Enrollment for the Phase II NOVEL 2 and Phase III NOVEL 4 trials have been completed; recruitment for the Phase III NOVEL 3 and 5 trials is under way. The randomized, double-blind, placebo-controlled, multicenter NOVEL 2 trial will examine the steroid-sparing properties of daily sargramostim SC injections in steroid-dependent CD patients in the United States and Canada for up to 22 weeks. The NOVEL 3 and 4 trials will be randomized, double-blind, placebocontrolled Phase III trials. NOVEL 3 will evaluate the efÞcacy of sargramostim in response and remission induction in CD patients in the United States who demonstrated a response to an initial cycle of the drug; NOVEL 4 will evaluate

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the ability of the drug to induce a response and remission in CD patients in ten countries outside the United States. NOVEL 5, an open-label trial for all patients previously treated with sargramostim, is designed to assess the drug’s long-term safety. Berlex announced the results of NOVEL 1 in an October 2003 press release; the results were presented that same month at the 68th Annual ScientiÞc Meeting of the American College of Gastroenterology and later published in a peerreviewed journal (Korzenik JR, 2005). The double-blind Phase II trial randomized 124 patients with moderate-to-severe active CD, at a ratio of 2:1, to receive 6 µg/kg of sargramostim or placebo in daily SC injections for 56 days. Concomitant use of antibiotics and aminosalicylates was allowed, but immunosuppressants and glucocorticoids were not permitted. Statistical signiÞcance was not achieved for the primary end point: a decrease of at least 70 points in CDAI at day 57 (54% for the sargramostim-treated group versus 44% for placebo). However, signiÞcantly more sargramostim-treated patients achieved the more stringent secondary endpoints: clinical remission (CDAI of 150 or less) and a decrease of 100 points or more in CDAI at day 57, compared with placebo. Remission was achieved by 40% of patients treated with sargramostim and 19% of placebo recipients; a 100-point decrease in CDAI was achieved by 48% of treated patients and 26% of placebo patients. Mucosal healing was evaluated using the Crohn’s Disease Endoscopic Index of Severity (CDEIS) score in a subset of patients from each group (19 from the sargramostim-treated group and 10 from placebo); median post-treatment CDEIS scores were found to be lower in the sargramostim-treated group than in the placebo group: 1.5 versus 5.6. Complete Þstula closure was observed in four of the eight sargramostim-treated patients who had draining Þstulas at baseline, compared with two out of Þve in the placebo group. Patients maintained clinical response, remission, and improvement in quality of life for 30 days following discontinuation of sargramostim treatment. According to a May 2004 Schering AG press release and presentation at the 2004 Digestive Disease Week conference, approximately 15% of patients maintained a clinical response to sargramostim six months after treatment was discontinued. There was no signiÞcant difference in the overall incidence of adverse events between the sargramostim-treated and placebo groups in the NOVEL 1 trial, and most of the events were mild to moderate in nature (Korzenik JR, 2005). Injection site reactions and bone pain were more common in sargramostim-treated patients than in placebo-treated patients and occurred in 90% and 37%, respectively, of treated patients. Of the 78 sargramostim-treated patients who were tested for antibodies to the agent, one patient tested positive. This patient did not experience any adverse events related to the development of neutralizing antibodies, but sargramostim-induced neutrophilia disappeared in the patient, possibly indicating a loss of drug response. According to the product label for Leukine, fever, nausea, vomiting, diarrhea, and liver abnormalities are common side effects associated with the use of sargramostim in oncology-related clinical studies.

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Preliminary results from the open-label NOVEL 5 trial, designed to assess long-term safety, were presented at Digestive Disease Week in May 2005 (Valentine J, 2005). Patients with active CD who had participated in a previous sargramostim trial were given daily injections of 6 µg/kg sargramostim for up to 6 cycles of treatment over 8 weeks; 42 patients completed at least one cycle of treatment. Concomitant antibiotics, aminosalicylates, and steroids were permitted. After one cycle of sargramostim treatment, 53% of patients achieved a 100-point decrease in the CDAI score and 40% achieved remission (CDAI of 150 or less). The most commonly reported adverse events were injection site reactions, bone pain, nausea, headache, vomiting, joint pain, back pain, and fatigue. The incidence of injection site reactions and bone pain decreased over subsequent treatment cycles. No serious drug-related adverse events were reported. Sargramostim’s beneÞcial effect in CD was also demonstrated in an open-label, dose-escalation, pilot Phase I trial involving 15 patients with moderate-to-severe CD (Dieckgraefe BK, 2002[b]). After eight weeks of sargramostim treatment (4–8 µg/kg per day delivered through self-administered SC injection), 12 of 15 CD patients demonstrated a 100-point or greater decline in CDAI score, and eight of 15 patients achieved clinical remission (deÞned as a CDAI score of 150 points or lower). Health-related quality of life, as measured by improvements in the IBD Questionnaire (IBDQ) scores, signiÞcantly improved for patients during sargramostim treatment. No serious adverse events were noted in this pilot study, but 80% of patients reported localized injection site reactions (e.g., itching, erythema), and 67% of patients experienced bone pain. Sargramostim, a growth factor that promotes the production of immune cells to boost patient immunity, offers a unique therapeutic approach to treatment of CD. Phase I and II studies to date are encouraging, but results from larger Phase III studies are needed to verify the efÞcacy of this novel approach. Even if sargramostim demonstrates efÞcacy against CD in Phase III trials, its sales potential in the CD market will be relatively modest because use of the agent is expected to be limited to the acute setting, given its frequent dosing proÞle (daily SC injections). Lack of long-term efÞcacy results and safety concerns about prolonged administration of sargramostim may also limit the agent’s use to the acute setting. Its high price is likely to limit its use in the United States to treatment of severe disease that is refractory to conventional CD therapies and at least two other biological agents. Cell Adhesion Molecule Inhibitors Overview. Cell adhesion molecules (CAMs) are cell-surface receptors that mediate cell-cell interactions that are critical in many different physiological processes and disease states, including wound healing, immune responses, inßammation, infectious diseases, and cancer metastases. Biopharmaceutical companies are pursuing different approaches, including MAbs and antisense oligonucleotides, to target and block CAMs.

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SigniÞcant setbacks have occurred in the development of drugs in this class. Serious adverse events, some of them fatal, resulted in the market withdrawal of natalizumab (Elan/Biogen Idec’s Tysabri)—a MAb against the α4-integrin protein on the surface of leukocytes that had been launched in the United States for treatment of relapsing-remitting multiple sclerosis (RR-MS)—and an FDA request to suspend clinical trials for all similar CAM inhibitors. These events have brought into question the continued development of α4-integrin antagonists for CD. Millennium’s MLN-02, a MAb that targets α4β7-integrin, is in Phase II development for CD in Canada, but its development in the seven major pharmaceutical markets (United States, France, Germany, Italy, Spain, United Kingdom, and Japan) appears to have been discontinued. Finally, Isis Pharmaceuticals discontinued development of its intracellular adhesion molecule-1 (ICAM-1) antisense inhibitor, alicaforsen (ISIS-2302), for CD in December 2004 because of poor efÞcacy data from two Phase III trials; Isis does plan to continue developing the drug for treatment of ulcerative colitis (UC). The only CAM inhibitor discussed here in greater detail is natalizumab. Mechanism of Action. CAMs include selectins (P-, E-, and L-selectin), integrins (e.g., β2 integrin), and the immunoglobulin superfamily of adhesion molecules (e.g., ICAM-1). Leukocyte migration from the bloodstream into inßamed or infected tissue is mediated by CAMs. These molecules are expressed on leukocytes and endothelial cells (the latter line blood vessels). CAM overexpression correlates with several inßammatory disorders, including CD. Blocking CAMs inhibits leukocyte migration into the intestinal mucosa and submucosa, thereby preventing the accumulation of immune cells that release proinßammatory cytokines and tissue-damaging free radicals known to be important in CD pathogenesis. Natalizumab. Elan and Biogen Idec (formerly Biogen) have been developing natalizumab (Tysabri) as a potential treatment for CD. The companies Þled a marketing authorization application (MAA) for CD with the European Medicines Agency for the Evaluation of Medicinal Products (EMEA) in September 2004. In October 2004, the FDA announced that it would require the results of ongoing Phase III induction studies before it would accept a U.S. application for CD. No development of natalizumab for CD has been reported in Japan. Natalizumab received approval for treatment of RR-MS in the United States in November 2004 after an accelerated priority review. Elan and Biogen Idec also Þled an MAA with the EMEA for MS in June 2004, and approval was expected in 2005. However, on February 28, 2005, Biogen Idec and Elan announced a voluntary withdrawal of natalizumab from the U.S. market and the suspension of all clinical trials involving the drug. These actions were based on two adverse events reported from ongoing natalizumab clinical trials in MS patients: one conÞrmed fatal case and one suspected case of progressive multifocal leukoencephalopathy (PML), a rare and frequently fatal demyelinating disease caused by an opportunistic infection of the central nervous system by the JC virus (JCV).

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Surprisingly, the FDA also requested the suspension of clinical trials for similar agents, even though no cases of PML have been reported in patients taking any drug other than natalizumab. Phase III clinical trials for natalizumab in CD have now been completed in the United States, but no application has yet been submitted, pending a thorough safety review of the agent for further incidence of PML. Natalizumab had also been in Phase II development for RA, but few useful data could be obtained from these trials before they had to be halted in February 2005; new studies would have to be started if the development for RA were to continue. Natalizumab is a humanized MAb that targets the α4 protein, a component of the integrins α4β1 (very late activation antigen [VLA]-4) and α4β7 (mucosal adressin, MAdCAM-1). MAdCAM-1 is an adhesion molecule on mucosal epithelium; VLA-4 is an adhesion molecule expressed on activated T cells and on macrophages that are important for the migration of these cells into the GI tract during inßammation. Therefore, blocking the α4 protein may interfere with the inßux of leukocytes into the intestinal tissue in CD. Three Phase III clinical trials examining natalizumab’s efÞcacy in treating CD were completed prior to the market withdrawal and suspension of clinical trials in February 2005. The Þrst of these trials, known as Evaluation of Natalizumab in Active Crohn’s Disease Therapy-1 (ENACT-1), assessed the drug’s ability to induce a clinical response (deÞned as a 70-point or greater reduction in CDAI score) and remission (CDAI score of 150 points or lower) in moderateto-severe CD patients with active disease. This double-blind, placebo-controlled study randomized 905 patients in a 4:1 ratio to 300 mg natalizumab or placebo. Patients received three doses IV at four-week intervals and were evaluated for a period of 12 weeks. Continuation of immunosuppressants was allowed. Results of this trial, Þrst announced in a July 2003 press release by Biogen Idec and Elan, showed a high placebo-response rate, and treatment with natalizumab did not produce a statistically signiÞcant clinical response, compared with placebo, at week 10, the trial’s primary end point. However, at week 12, there was a signiÞcant difference in response and remission rates between the two groups as well as signiÞcant differences at week 10 on secondary measures of efÞcacy, such as IBDQ scores (a measure of quality of life), time to remission, and mean changes in CDAI score. A subpopulation analysis of patients with active inßammation, as indicated by elevated CRP levels—72% of all patients enrolled in the ENACT-1 trial—revealed signiÞcant differences in response and remission rates at week 10 between patients treated with natalizumab and patients treated with placebo. Endoscopic healing was also examined in a subset of ENACT-1 participants demonstrating a 50% improvement in CDEIS scores by week 10 in the natalizumab-treated group, compared with a 7% decrease in CDEIS scores in the placebo group (Rutgeerts P, 2004[b]). Additionally, 22% of the 53 natalizumabtreated patients evaluated by endoscopy who had ulcers at week 0 were ulcer free by week 10, versus 8% of the placebo-treated patients. The rate of adverse events in the ENACT-1 trial was similar in natalizumab and placebo recipients. Headache, nausea, and abdominal pain were the most

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commonly reported side effects in both treatment groups. Although few adverse events were noted, natalizumab-treated patients demonstrated a sustained increase in the number of circulating lymphocytes, a potentially serious condition known as lymphocytosis: at week 12, the mean total circulating lymphocyte count for the natalizumab-treated group was 2,857 cells/mm3 versus 1,690 cells/mm3 for the placebo-treated group (Rutgeerts P, 2004[b]). A second Phase III induction trial, entitled ENCORE, was conducted in CD patients with elevated CRP levels (greater than 2.87 mg/L), a common strategy for agents that have failed efÞcacy end points in CD trials. In this doubleblind study, 510 patients with moderate-to-severe active CD were randomized to receive 300 mg natalizumab or placebo via intravenous (IV) infusions at weeks 0, 4, and 8; efÞcacy and safety assessments were performed at weeks 4, 8, and 12. According to a June 2005 press release by Elan and Biogen Idec, the trial met its primary end point: clinical response (deÞned as a 70-point or greater reduction in CDAI score) at weeks 8 and 12. ENCORE also met all of its secondary end points, including induction of remission (CDAI score of 150 points or lower) at weeks 8 and 12. There were no signiÞcant differences between natalizumab and placebo in the overall rates of adverse events; the most common side effects were headache, nausea, abdominal pain, and nasopharyngitis. ENCORE participants are being monitored for long-term safety. Natalizumab’s third Phase III trial, known as Evaluation of Natalizumab As Continuous Therapy-2 (ENACT-2), assessed the drug’s ability to maintain clinical response in ENACT-1 participants who had achieved response and/or remission after three natalizumab infusions. The double-blind, placebo-controlled study re-randomized 339 patients to receive either placebo or 300 mg natalizumab as a monthly infusion for up to 12 months; the primary endpoint was the proportion of patients who were able to maintain a response for six consecutive months. Loss of response was deÞned as a 70-point or greater increase in CDAI, a CDAI score of 220 or greater, or the use of rescue intervention at any point. Concomitant use of immunosuppressants was permitted. After six months of additional treatment, 61% of natalizumab recipients and 29% of placebo recipients continued to demonstrate a clinical response; 44% of patients treated with natalizumab maintained clinical remission compared with 26% of patients treated with placebo (Sandborn WJ, 2004[a]). Both the response and remission Þndings were statistically signiÞcant. Natalizumab was able to maintain a clinical response and remission in patients through one year: 54% of natalizumab-treated patients maintained a clinical response and 39% maintained remission (Sandborn WJ, 2005[b]). Corresponding 12-month response and remission rates for the placebo group were 20% and 15%, respectively. Additionally, 49% of patients treated with natalizumab were able to withdraw from steroids at the end of 12 months, compared with 20% of placebo-treated patients (Colombel J, 2005). During the ENACT-2 trial, there was no noticeable difference in the rate of adverse events—either serious or nonserious—between natalizumab- and placebo-treated patients. Headache, nasopharyngitis, nausea, and abdominal pain were the most commonly reported side effects in the 12-month study.

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Natalizumab may have utility in CD patients who are intolerant of or have failed inßiximab therapy, as demonstrated in the ENACT trials that included patients who had been previously treated with inßiximab. In ENACT-1, natalizumab induced a clinical response in 54% of the 360 patients who had received prior inßiximab therapy, compared with 35% of placebo-treated patients. A trend toward increased clinical remission was observed: 33% for natalizumab and 22% for placebo (Sandborn WJ, 2004[b]). In the ENACT-2 trial, in CD patients previously treated with inßiximab and in patients who had previously failed inßiximab therapy, natalizumab also demonstrated signiÞcantly greater maintenance of clinical response and remission than placebo for an additional 12 months (Panaccione R, 2005). Results from a double-blind, placebo-controlled European Phase II trial have also demonstrated natalizumab’s ability to elicit a clinical response and clinical remission and improve quality of life in 244 patients with moderate-to-severe CD (CDAI score 220–450) (Ghosh S, 2003). Patients were randomized to one of four treatment groups: two infusions of placebo; one 3 mg/kg infusion of natalizumab and one infusion of placebo; two 3 mg/kg infusions of natalizumab; and two 6 mg/kg infusions of natalizumab. Patients received their two infusions at weeks 0 and 4 and were followed for 12 weeks from the trial’s inception. Compared with the group receiving two placebo infusions, patients treated with two 3 mg/kg infusions of natalizumab demonstrated the most consistently signiÞcant clinical responses (deÞned as a 70-point or greater decline in CDAI score) and clinical remission (deÞned as a CDAI score of 150 points or lower). At week 6, the prospectively deÞned primary end point of the study, the group treated with two 3 mg/kg infusions of natalizumab demonstrated a response rate of 71% and a remission rate of 44%, compared with rates of 38% and 27%, respectively, for the group treated with two placebo infusions. SigniÞcant Þndings for the group treated with two 3 mg/kg infusions of natalizumab persisted through week 12, at which point the response rate and remission rate were 61% and 42%, respectively; the placebo group demonstrated a response rate of 43% and a remission rate of 27% at week 12. Although the group treated with two 6 mg/kg infusions of natalizumab demonstrated a higher response rate at weeks 6 and 12 than the placebo group, the remission rate for this group was not statistically signiÞcant at these two times. Compared with the group receiving two placebo infusions, all three natalizumab-treated groups in the European Phase II study experienced signiÞcant improvements over baseline scores on the IBDQ at week 6. This improvement persisted at week 12 for the two groups receiving second infusions of natalizumab. Overall, natalizumab was well tolerated by patients in the European Phase II trial, and the number of adverse events did not differ signiÞcantly between drug-treated and placebo groups (Ghosh S, 2003). The most common adverse events experienced by patients in this study were headache (32% of both natalizumab-treated patients and placebo recipients); abdominal pain (15% of natalizumab-treated patients, 17% of placebo recipients); inßuenza syndrome (14% of natalizumab-treated patients, 8% of placebo recipients); colitis (13%

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of natalizumab-treated patients, 14% of placebo recipients); and nausea (9% of natalizumab-treated patients, 16% of placebo recipients). Serious adverse events were reported by 10% of natalizumab-treated patients and 11% of placebo-treated recipients. None of these serious adverse events were considered to be related to treatment; most were related to CD symptoms and complications. However, lymphocytosis was a common effect of natalizumab treatment, with mean lymphocyte counts increasing 1.3 to 1.9 times over baseline values. Currently available data suggest that the efÞcacy of natalizumab in maintaining remission in CD patients is similar to that of inßiximab. Encouraging results from ENACT-2 indicate that natalizumab can maintain a clinical response and remission for up to one year. However, natalizumab’s ability to induce CD remission is less clear. The ENACT-1 induction study missed its primary end point, and signiÞcant remission induction was demonstrated in the ENCORE trial only in a subset of CD patients who had high CRP levels. Many agents in development for CD have had difÞculty meeting primary efÞcacy end points because of high placebo response rates; analysis of subgroups with high CRP levels, presumably patients with the most active disease, has salvaged the efÞcacy results. Therefore, natalizumab appears able to induce remission of active CD, but its efÞcacy in so doing is likely less than that of inßiximab. In addition, inßiximab has many other advantages over natalizumab, including a demonstrated ability to treat Þstulizing disease, less frequent dosing (IV every 8 weeks versus every 4 weeks), and long-term safety data. To gain approval for treatment of CD, natalizumab must also overcome the hurdle of its association with signiÞcant life-threatening side-effects (PML). Since natalizumab’s withdrawal, the suspected PML case has been conÞrmed, and one additional conÞrmed case and two suspected cases of PML have been discovered in clinical trial participants. Of the three conÞrmed natalizumabassociated PML cases, two were in MS patients receiving 300 mg of natalizumab every four weeks in combination with Biogen Idec’s interferon beta-1a (Avonex) (Langer-Gould A, 2005; Kleinschmidt-DeMasters BK, 2005). Natalizumab’s product label for use in RR-MS patients did warn of a minor drug interaction with Avonex that resulted in decreased clearance rates but stated that this alteration in clearance rate did not necessitate reduction of natalizumab’s dose to maintain safety. The third conÞrmed case of PML was fatal, occurring in a CD patient who was taking natalizumab as monotherapy (300 mg every 4 weeks), although the patient had previously taken other immunosuppressive agents, including azathioprine and inßiximab (Van Assche G, 2005). Therefore, the cases of PML appear to be associated with natalizumab treatment, but concomitant use of Avonex or other immunosuppressants likely increases the risk of developing the disease, because of either drug interactions or cumulative immunosuppressive effects. Regulatory authorities are likely to require further safety studies or the development of screening and monitoring tests before natalizumab is relaunched for MS or gains any further approvals. It is possible to test for the presence of the JC virus to rule out natalizumab treatment in patients with latent JCV infection.

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However, the rate of seropositivity for JCV in healthy adults is high, with estimates ranging from 50% to 86% (Knowles WA, 2003; Weber T, 1997), which would severely limit the drug’s utility and market potential. Alternatively, patients taking natalizumab could be tested for increases in plasma JCV levels or monitored for PML lesions by MRI, and therapy stopped if either is detected. In one natalizumab-associated PML case, JCV was detected in the plasma two months prior to clinical onset of PML (Van Assche G, 2005). Likewise, the Þrst PML lesion was detected by MRI in an MS patient two months prior to the development of obvious neurological deÞcits (Langer-Gould A, 2005). This lesion was difÞcult to distinguish from an MS-induced lesion, but such lesions in a CD patient would be highly unusual and indicative of PML onset. While it is possible to monitor for early signs of PML before the onset of clinical disease, natalizumab’s immunosuppressive effects appear to persist for approximately three months after cessation of treatment, and it is not known whether stopping therapy after detection of JCV or PML lesions would prevent full-blown PML (Langer-Gould A, 2005). Biogen Idec and Elan Þnished a thorough review of natalizumab’s safety data in MS trials in August 2005 and in CD and RA trials in October 2005 and found no additional cases of PML. According to a press release in April 2005, the companies are taking preliminary steps to restart MS trials. In September 2005, the companies submitted an application to resume marketing of natalizumab for MS in the United States; an FDA decision is expected within six months. MAP Kinase Inhibitors Overview. The MAP kinase signal transduction pathway is central to the normal physiological functioning of cells, including cell growth, division, and survival. The MAP kinase pathway is also important in cytokine production, including TNF-α production. Overactivation of this pathway is involved in many disease states, such as immune and inßammatory disorders, central nervous system disorders, cardiovascular disease, and cancer. Therefore, drugs that inhibit this signaling pathway have potential applications across several therapeutic areas. MAP kinase inhibitors have the potential to address several unmet needs in inßammatory conditions like CD, including control of aberrant inßammatory cytokine production via an orally available, small-molecule formulation. Small-molecule MAP kinase inhibitors would be more convenient and potentially less expensive than parenteral agents such as the biological TNF-α inhibitors. However, an incomplete understanding of signal transduction biology has hampered the development of safe and effective agents that modulate this pathway because drug discovery has relied on imprecise, high-throughput screens that result in molecules with cross-reactivity to other kinases. Consequently, agents within this drug class have had a high discontinuation rate; the development of many agents within this class has been terminated because of cross-reactivity with unknown, related proteins that results in unanticipated side effects or a lack of efÞcacy. This section focuses on the two MAP kinase inhibitors currently in Phase II development for CD: Procter & Gamble’s RDP-58

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and Cytokine PharmaSciences’ semapimod. Boehringer Ingelheim’s MAP kinase inhibitor, doramapimod, which was in Phase II development in the United States for CD, has had no development reported since March 2003, so it is not discussed in further detail here. Mechanism of Action. The MAP kinase signal transduction pathway—which comprises the p38, JNK, and ERK pathways—is involved in the synthesis of proinßammatory cytokines. This pathway can also be activated by the binding of these cytokines to their respective receptors and thus helps to regulate cellular responses to proinßammatory cytokines. Therefore, drugs that inhibit the MAP kinase pathway have the potential to inhibit the feedback loop common to inßammatory conditions that involve proinßammatory cytokines. Specimens of inßamed mucosa taken from CD patients contain increased quantities of several proinßammatory cytokines (e.g., TNF-α, IL-1β, IL-2, IL-6, IL-8, IL-12, IL-18, and IFN-γ ), and long-term overexpression of these proinßammatory cytokines can result in tissue damage, as occurs in CD. Furthermore, colonic biopsies of patients with severe CD have demonstrated enhanced p38 and JNK activation (Hommes D, 2002). Therefore, MAP kinase inhibitors may represent a promising novel approach for treating CD. Targeting the MAP kinase pathway requires caution, however. Agents in this class must be carefully designed because of the potential for toxicity. An overly robust inhibitor of the MAP kinase pathway may cause side effects (such as blocking normal levels of signal transduction required for cellular homeostasis), whereas a nonspeciÞc MAP kinase inhibitor may not demonstrate any efÞcacy and may provoke unwanted responses. RDP-58. Procter & Gamble (P&G) is developing RDP-58 (delmitide) as a potential treatment for inßammatory bowel disease (IBD; a general term that encompasses CD and the related disease UC). The drug was discovered by the Institut de Transplantation et de Recherche en Transplantation (ITERT) and SangStat Medical in collaboration with Synt:em, using the latter’s computer-aided rational design technology, Acti:map. SangStat had completed Phase II European trials with RDP-58 when the company was acquired by Genzyme in September 2003. In April 2004, Genzyme announced an agreement whereby the company licensed worldwide development, manufacturing, and commercialization rights for RDP-58 to P&G in exchange for up-front and milestone payments, as well as royalties based on sales. (Genzyme retained the rights to RDP-58 for pulmonary conditions and other indications not covered in this agreement and the rights to copromote RDP-58 with P&G for oncology-related disorders.) Clinical studies have been conducted to evaluate RDP-58 as a potential treatment for CD, but according to an April 2004 press release issued by P&G and Genzyme, the former will initially focus on UC as the primary indication for RDP-58. RDP-58 is an orally administered decapeptide composed of nine D-amino acids and glycine. The agent’s D-amino acid composition makes it resistant to degradation by proteases (Sorbera LA, 2004). Although administered orally, RDP-58

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does not appear to be systemically bioavailable and is believed to exert its therapeutic effect locally in the intestines. According to a May 2003 SangStat press release, RDP-58 inhibits the intracellular protein complex TRAF6/MyD88/IRAK (TRAFYK), thereby preventing the activation of p38 MAP kinase and JNK 1 and 2 kinases in the MAP kinase signaling pathway. The TRAFYK complex is also believed to play an important role in the NFκB signaling pathway, important in inßammatory responses. RDP-58’s interference with multiple signaling pathways through its interaction with TRAFYK results in the inhibition of the synthesis of several proinßammatory cytokines, including TNF-α, IFN-γ , IL-2, and IL-12. SangStat announced disappointing efÞcacy results from two European Phase II trials in an April 2003 company press release and a subsequent peer-reviewed journal article (Sorbera LA, 2004). A total of 104 patients with mild-to-moderate CD participated in the blinded, placebo-controlled studies. Patients were randomized to receive an oral solution containing placebo or 100, 200, or 300 mg/day of RDP-58 for 28 days; patients were subsequently followed for an additional month. Using a 70-point or greater decline in the CDAI as a measure of response and a CDAI score of 150 points or lower as a measure of remission, treatment with 200 mg/day RDP-58 was associated with the highest response rate (66%) and highest remission rate (48%), compared with response and remission rates of 43% and 20%, respectively, for the placebo group. However, these Þndings were not statistically signiÞcant. RDP-58 was generally well tolerated; all serious adverse events were related to either disease progression or leukopenia (which occurred in one patient). Nausea and headache were the most commonly reported side effects; the incidence of headache was statistically signiÞcant and was more prominent in the placebo and 100 mg/day groups. At the time the Phase II trial results were released, SangStat believed that increased or longer dosing of RDP-58 might be necessary to produce statistically signiÞcant results in CD patients because of the transmural nature of CD-related inßammation. However, P&G is apparently pursuing UC as the lead indication for RDP-58; the drug’s future development for CD is uncertain. Semapimod. Semapimod (formerly CNI-1493), a synthetic guanylhydrazone MAP kinase inhibitor, is under development by Cytokine PharmaSciences (formed in 1999 by the merger of Cytokine Networks and PharmaSciences). Phase II trials are being conducted for CD in the United States and Europe. The drug is also in Phase II development in the United States for pancreatitis and psoriasis, and studies are under way to examine its potential to treat cancer, congestive heart failure, and pain. No development has been reported in Japan. The company has announced on its Web site that it is seeking licensing partners for semapimod in various indications and regions. Semapimod has been shown to inhibit p38 MAP kinase and JNK phosphorylation (Hommes D, 2002). It has also been shown to inhibit the production of nitric oxide, the proinßammatory cytokines TNF-α, IL-1, and IL-6, and the macrophage inßammatory proteins (MIP)-1α and MIP-1β in both murine models and in vitro studies (Bianchi M, 1995; Bianchi M, 1996).

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The results of an open, randomized, double-blind study that showed a dose response for semapimod in 33 CD patients were presented at the 2004 Digestive Disease Week conference (Buchman AL, 2004). The study randomized patients with moderate-to-severe CD to receive daily IV infusions of placebo or one of two doses of semapimod (30 mg or 60 mg) for Þve days. On day 29, 20%, 31%, and 60% of patients treated with placebo, 30 mg semapimod, and 60 mg semapimod, respectively, demonstrated a response (deÞned as a 70-point or greater reduction in CDAI score). On day 57, response rates were 10%, 46%, and 50% respectively for the three treatment groups. No signiÞcant toxicities associated with the drug were reported in this open study, but there was a signiÞcant increase in infusion site irritation and/or phlebitis in the groups treated with semapimod, compared with placebo-treated patients. The rate of infusion site irritation and/or phlebitis was 69% and 80% in patients treated with 30 mg and 60 mg semapimod, respectively, compared with only 20% in patients treated with placebo. The study was terminated early because of poor accrual on account of the infusion discomfort and dosing schedule. Alternative formulations and shorter durations of treatment are now being evaluated (Gotham S, 2004). In a Phase I pilot study involving 12 patients with severe CD, semapimod was shown to be safe and effective in reducing CDAI scores (Hommes D, 2002). Study participants were randomized to receive 8 mg/m2 or 25 mg/m2 of semapimod once daily via IV infusion; study medication was administered for 12 days and patients were followed for 4 months. Safety was the trial’s primary end point, and patients were allowed to continue stable doses of other CD therapies (e.g., corticosteroids, aminosalicylates, antibiotics, immunosuppressants) during the course of the study. Eight of the 12 patients completed the full 12 days of the study. Four patients discontinued the study because of elevated liver enzymes (alanine aminotransferase levels in two patients), catheter-related infection (one patient), or worsening CD (one patient). The most common side effects associated with semapimod treatment in this study were digestive/hepatic, cardiovascular, or hematologic in nature. Hommes and colleagues also evaluated semapimod in a small, uncontrolled study of 12 CD patients, with measures of efÞcacy as secondary end points. The highest response rate (deÞned as a 25% or greater reduction in CDAI score and 70-point or greater reduction in CDAI occurring at least once after the start of the trial) was 67% and occurred 15 and 29 days after the start of the study. The highest remission rate (deÞned as a CDAI score of 150 points or lower at least once after the start of the trial) was 50% and occurred at day 112 in the study. At week 4, all but one of the study participants experienced rapid endoscopic healing, and 4 of the 5 patients with Þstulizing CD experienced Þstula closure during the course of the study. If future clinical studies conÞrm the Þndings of this pilot study, then semapimod could become an important therapy in CD treatment. However, there is currently no statistically signiÞcant evidence for the efÞcacy of the drug in inducing a clinical response or remission in CD patients compared with placebo.

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Velayos FS, et al. Effect of 5-aminosalicylate use on colorectal cancer and dysplasia risk: a systematic review and meta-analysis of observational studies. American Journal of Gastroenterology. 2005;100(6):1345–1353. Veloso FT, et al. Clinical outcome of Crohn’s disease: analysis according to the Vienna classiÞcation and clinical activity. Inßammatory Bowel Diseases. 2001;7:306–313. Verispan, Scott-Levin. Physician Drug & Diagnosis Audit, December 2003-December 2004. Walker LJ, et al. Anti-saccharomyces cerevisiae antibodies (ASCA) in Crohn’s disease are associated with disease severity but not NOD2/CARD15 mutations. Clinical and Experimental Immunology. 2004;135(3):490–496. Weber T, et al. Analysis of the systemic and intrathecal humoral immune response in progressive multifocal leukoencephalopathy. Journal of Infectious Diseases. 1997;176: 250–254. Wehkamp J, et al. Innate immunity and colonic inßammation: enhanced expression of epithelial alpha-defensins. Digestive Disease Sciences. 2002;47(6):1349–1355. Wehkamp J, et al. NOD2 (CARD15) mutations in Crohn’s disease are associated with diminished mucosal alpha-defensin expression. Gut. 2004;53(11):1658–1664. Wehkamp J, et al. Defensin deÞciency, intestinal microbes, and the clinical phenotypes of Crohn’s disease. Journal of Leukocyte Biology. 2005;77(4):460–465. Wenckert A, et al. The long-term prophylactic effect of salazosulphapyridine (Salazopyrin) in primarily resected patients with Crohn’s disease. A controlled double-blind trial. Scandinavian Journal of Gastroenterology. 1978;13(2):161–167. West RL, et al. Clinical and endosonographic effect of ciproßoxacin on the treatment of perianal Þstulae in Crohn’s disease with inßiximab: a double-blind placebo-controlled study. Alimentary Pharmacology and Therapeutics. 2004;20:1329–1336. Wilk JN, Viney JL. GM-CSF treatment for Crohn’s disease: a stimulating new therapy? Current Opinion in Investigational Drugs. 2002;3(9):1291–1296. Williams CN. Role of rectal formulations: suppositories. Scandinavian Journal of Gastroenterology. 1990;172:60–62. Williams JB, et al. Long-term inßiximab maintenance infusion regimens and rates of hospitalization, surgery, and disability in Crohn’s disease patients. Digestive Disease Week. May 14–19, 2005; Chicago, IL. Willoughby JMT, et al. Controlled trial of azathioprine in Crohn’s disease. Lancet. 1971;2(7731):944–947. Winter TA, et al. Intravenous CDP870, a PEGylated Fab’ fragment of a humanized antitumor necrosis factor antibody, in patients with moderate-to-severe Crohn’s disease: an exploratory study. Alimentary Pharmacology & Therapeutics. 2004;20:1337–1346. Witte J, et al. Disease outcome in inßammatory bowel disease: mortality, morbidity and therapeutic management of a 796-person inception cohort in the European Collaborative Study on Inßammatory Bowel Disease (EC-IBD). Scandinavian Journal of Gastroenterology. 2000;35:1272–1277. Workman EM, et al. Diet in the management of Crohn’s disease. Human Nutrition. Applied Nutrition. 1984;38(6):469–473. World MJ, et al. Mesalazine-associated interstitial nephritis. Nephrology Dialysis Transplantation. 1996;11(4):614–621.

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Yacyshyn BR, et al. A randomized, placebo-controlled trial of an antisense ICAM-1 inhibitor (ISIS-2302) in steroid-dependent Crohn’s disease showed clinical improvement at high serum levels. Gastroenterology. 2001;120:5(suppl 1). Abstract 1447. Yang SK, et al. Epidemiology of inßammatory bowel disease in Asia. Inßammatory Bowel Diseases. 2001;7:260–270. Yao T, et al. Crohn’s disease in Japan: diagnostic criteria and epidemiology. Diseases of the Colon and Rectum. 2000;43:S85–S93. Yoshida EM. The Crohn’s Disease Activity Index, its derivatives, and the Inßammatory Bowel Disease Questionnaire: a review of instruments to assess Crohn’s disease. Canadian Journal of Gastroenterology. 1999;13:65–73. Yoshida Y, Murata Y. Inßammatory bowel disease in Japan; studies of epidemiology and etiopathogenesis. Medical Clinics of North America. 1990;74:67–90.

Psoriasis

ETIOLOGY AND PATHOPHYSIOLOGY Introduction Psoriasis, derived from the Greek word psora, meaning “itch,” was Þrst classiÞed as a unique skin disorder in the late 18th century. It is a chronic disease that encompasses a wide range of severities. This dermatologic disease causes tremendous morbidity and may be associated with increased mortality. Skin involvement ranges from mild to very severe, and skin disease is sometimes accompanied by the debilitating joint disease psoriatic arthritis (PsA). Psoriasis is now understood to be an inßammatory disease, and current research is largely directed to understanding the roles of T cells and inßammatory cytokines in the pathology of psoriasis. Pathophysiology Psoriasis may affect the skin and nails; psoriatic arthritis may also develop in a subset of these patients and can become debilitating. Clinical presentation varies greatly among affected patients and may even vary markedly in an individual patient during the course of the disease. Psoriasis presentation may be as innocuous as a single pit in a Þngernail or as serious as generalized, disÞguring skin lesions and disabling PsA. Psoriasis is thought to arise from a combination of factors, including genetics, environmental triggers (e.g., climate, trauma), associated diseases (particularly infection), immunological status, and concurrent medications. The hallmark psoriatic skin lesions are sharply demarcated, thick, erythematous (red) patches that are covered by silvery scales. These lesions are often Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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PSORIASIS

Scalp Ears

Chest Elbows

Trunk

Umbilicus Gluteal cleft

Nails

Genitalia Knees

Toenails FIGURE 1. Typical locations of psoriasis lesions.

pruritic (itchy), may be painful, and typically bleed slightly upon removal of the scale (a phenomenon known as the Auspitz sign). Psoriasis lesions may occur anywhere on the body; Figure 1 depicts typical sites. The vast majority of patients have chronic plaque psoriasis. The disease course is quite variable, regardless of severity. Some patients experience periods of remission followed by exacerbation, both of which can vary widely in duration; in others, lesions are always present. Table 1 describes the classiÞcation of the clinical forms of psoriasis, which are based principally on the patterns and shapes of skin lesions. Pathophysiological characteristics of psoriatic skin include increased keratinocyte activity and lymphocyte inÞltration. The result is hyperproliferation and accelerated maturation of the epidermis; inßammation involving T cells, T-cell products (e.g., tumor necrosis factor alpha [TNF-α]), other cells (e.g., neutrophils), and cell products of inßammation; and vascular alterations. In clinical trials, a wide variety of measures are used to evaluate the severity of psoriasis and response to therapy. They include individual scores for signs such as erythema, plaque thickness, or scaling, as well as pooled indices of signs of psoriasis. The most commonly used pooled index in clinical trials is the Psoriasis Area and Severity Index (PASI). The Overall Lesional Assessment (OLA) is a measure of a physician’s judgment of a patient’s overall lesional severity, with emphasis on plaque elevation, and is used in some clinical trials. The Physicians Global Assessment (PGA), a more general evaluation tool, is also sometimes employed. However, in everyday medical practice such measures are seldom used and no single deÞnition of disease severity is uniformly accepted. Table 2

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TABLE 1. Clinical Forms of Psoriasis Percentage of Psoriasis Patients Affected

Type

Cutaneous manifestations Plaque-type 75–85 psoriasis (Psoriasis vulgaris, chronic stationary psoriasis)

Guttate psoriasis (eruptive psoriasis)

15–20

Pustular psoriasis

2

Erythrodermic 1–2 psoriasis (exfoliative dermatitis, psoriatic erythroderma)

Description of Lesions • First appears as small papules, then grows to form red-violet round or oval plaques overlaid with silvery scales. • May be hyperpigmented with brown or black coloration in darkly pigmented patients. • Comprises droplet-shaped (dotlike) lesions ranging in diameter from 0.1 to 1.0 cm. • Lacks the scaliness and induration that is common to plaque psoriasis. • Found predominantly on the trunk and proximal areas of extremities and likely to involve the face. • Sterile pustules are frequently localized to the palms and soles (Pustulosis palmaris et plantaris) but may be generalized (von Zumbusch variant). • Localized form may develop from vesicles to vesicopustules to frank pustules to dried brown maculopapules. • Generalized form has irregular red patches over which thousands of 1–2 mm superficial pustules develop and coalesce to form large collections of pus. • Inflammatory lesions may cause extreme reddening (erythema) of all or most of the body surface. • Large areas of skin may slough off during the exfoliative phase.

Comments • Most commonly affected areas: knees, elbows, scalp, behind the ears, sacrum (gluteal cleft), umbilicus, intergluteal cleft, genitalia.

• May be the initial manifestation or a flare of chronic plaque psoriasis. • May be precipitated by Streptococcus groups A, C, and G, by viral infection, or by immune system compromise. • The most common form in children; characteristic of early- onset psoriasis. • May present de novo or as a flare of plaque psoriasis. • Average age of onset is 50 years. • Localized form occurs more frequently in females, is difficult to treat, and frequently recurs. • Generalized pustular psoriasis is very severe, is difficult to treat, may require inpatient care, and may cause death. • Usually occurs in people with chronic plaque psoriasis but may occur de novo. • Average age of onset is 50 years. • Occurs more frequently in males.

580

PSORIASIS

TABLE 1. (continued)

Type

Percentage of Psoriasis Patients Affected

Description of Lesions • Pustules may develop, leading to generalized pustular psoriasis.

Scalp psoriasis Approximately 50

Inverse psoriasis

—

• Plaque-type psoriasis lesions on the scalp.

• Usually smooth and erythematous, not scaly. • Occurs on flexural sides of the extremities, in contrast to the distribution of plaque psoriasis.

Extracutaneous manifestations • Nail psoriasis Up to 50 (80 in patients with • psoriatic arthritis)

Psoriatic arthritisApproximately 20

Large, deep, random pits of the nail plate are the most common manifestation. Nail thickening, ridging, crumbling, and splinter hemorrhaging are common. • Loss of the nail plate may occur in severe lesions. • Fingernails are more commonly affected than toenails. • Similar pathology to rheumatoid arthritis (may progress to severe joint destruction and deformity).

Comments • Precipitating factors include excessive use of potent topical or systemic corticosteroids, systemic illness, psychological stress, and alcoholism. • Very difficult to treat because topical formulations generally do not penetrate well. • Typically appears in the armpit or groin, under pendulous breasts, or in the skin folds of obese patients.

• Nail psoriasis may closely resemble onychomycosis (fungal infection of the nail), which must be ruled out first.

• Arthropathy that typically affects asymmetrical joints (unlike rheumatoid arthritis).

provides some commonly used deÞnitions of psoriasis severity and estimated percentages of psoriasis patients with each severity. Keratinocyte Hyperproliferation. The epidermal cell cycle is dramatically shortened from the 26–28 days normally required to form the epidermis to 4–6 days in people with psoriasis. Cell division in the basal layer occurs every 1.5 days, and migration of keratinocytes to the stratum corneum (the outer layers of the epidermis) occurs within approximately 4 days. This much-shortened cycle of cell division is further exacerbated by the proportion of germinal center cells entering the growth fraction—essentially 100% in patients with psoriasis, compared with 60–70% in normal skin. This high proportion results in a net increase

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TABLE 2. Psoriasis Severity Definitions in Clinical Practice Percentage of Psoriasis Patients Affectedb

Body Surface Area Covered (%)

Mild

65–80

<2

Moderate

15–25b

2–10

Severe

5–10b

>10

Severitya

Description Generally, isolated patches of psoriasis appear on the knees, elbows, hands, feet, and scalp. Topical therapies, including over-the-counter and prescription creams, ointments, and shampoos, are usually effective. Psoriatic skin lesions may occur on arms, legs, torso, scalp, and other areas. Topical agents, phototherapy, and/or oral drug therapy may be used, depending on the location and extent of the lesions, and on individual factors. Extensive areas of skin may be covered with psoriatic plaques or pustules. Alternatively, widespread erythrodermic lesions may cause severe peeling of the skin. Concomitant psoriatic arthritis often occurs. Therapeutic options intensify to include powerful oral agents with significant risk of side effects.

a In clinical trials, measures including the Psoriasis Area and Severity Index (PASI), the Physicians Global

Assessment (PGA), and the Overall Lesional Assessment (OLA) are used to assess disease severity and quantify response to treatment. b The National Psoriasis Foundation estimates a 20–35% share for moderate and severe psoriasis. Note: Percentages add up to 95–110%, underscoring the overlap between severities. Source: Modified from National Psoriasis Foundation estimates.

in the number of cells produced per day per square millimeter of skin surface. The rapid arrival of newly formed keratinocytes at the skin surface precludes proper differentiation and maturation of these cells, resulting in deÞcient keratinization of the stratum corneum and a three- to Þvefold increase in thickness of the psoriatic epidermis, which is full of scaly cells. Blood vessel dilation in the papillary layer of the dermis occurs, along with inÞltration of inßammatory cells (e.g., T cells, neutrophils). It has been suggested that an intrinsic regulatory defect that is critical to psoriatic lesion development is present in the epidermis itself, including nonlesioned skin. Indeed, one study found that uninvolved (i.e., nonlesioned) skin of psoriatics demonstrates altered epidermal kinetics when grafted onto athymic mice (Fraki JE, 1982). Koebner’s phenomenon—a hallmark of the disease—is the development of psoriatic plaques following mechanical trauma to nonlesioned skin and occurs in approximately 20% of patients with psoriasis. This phenomenon provides additional support for this theory. Keratinocytes participate in immunomodulation through the production of proinßammatory and inhibitory cytokines (e.g., interleukin-1 [IL-1], -6, and -8

582

PSORIASIS

and TNF-α). Cytokine production by hyperproliferating keratinocytes appears out of balance in psoriasis; TNF-α and IL-8 levels are increased. Keratinocytes also produce angiogenic factors that may account for capillary proliferation observed in psoriatic lesions (addressed later in this section). T Cells. Psoriasis is now thought to be principally a T-cell-mediated disease, and many theories regarding the role(s) of T cells in the pathophysiology of psoriasis have been proposed. One theory holds that cytokines released by keratinocytes (which may have been activated by Koebner’s phenomenon) activate T cells, which then produce more cytokines, amplifying the inßammatory, T-cell, and keratinocyte responses. Another theory holds that epidermal Langerhans cells interact with helper T cells, which activate keratinocytes and the ampliÞcation loop just described. A further theory of T-cell involvement proposes an autoimmune mechanism whereby killer (CD8+ ) T cells attack and activate keratinocytes. The Þrst evidence of a major role for T-cell dysregulation in psoriasis was the serendipitous discovery that inhibiting T-cell activation with cyclosporine (an immunosuppressant) was effective for treating the disease. Further support was provided by a study in which the destruction of activated T cells with a speciÞc toxin (consisting of the receptor-binding domain of IL-2 coupled with diphtheria toxin) caused remission of psoriatic symptoms and a reduction in epidermal CD8+ T cells (Gottlieb SL, 1995). CD4+ and CD8+ T cells have both been implicated in psoriasis disease progression. It is thought that inÞltrating CD4+ T cells localize in the epidermis during plaque development and send a signal that activates dormant CD8+ T cells already present in the epidermis. The observation of an increase in circulating CD8+ T cells in psoriatic patients provides further evidence. T cells positive for cutaneous lymphocyte antigen (CLA), indicating they are primed for cutaneous accumulation, are found in high concentrations in psoriatic lesions. Psoriasis appears to be mainly a TH 1-mediated disorder because both circulating and lesional CD8+ and CD4+ T cells produce type I cytokines including IL-2 and interferon-gamma (IFN-γ ), with an associated increase in TNF-α, IL-6, and IL-8; only a small proportion of cells produce the TH 2 cytokines IL-4 and IL-10. CD4+ T cells appear to be involved in plaque initiation (by preparing the microenvironment), while CD8+ T cells produce the TH 1 cytokines (e.g., IFN-γ ) that are involved in plaque persistence. The lack of structural uniformity in psoriatic lesions has led some researchers to speculate that different subsets of T-cell products may play different roles in lesion pathogenesis at different stages of lesion progression. T cells release lymphokines in the relatively quiescent center of plaques that can initiate a positive feedback loop with keratinocytes (described previously), which respond to these T-cell molecules and produce T-cell-activating cytokines. This T-cell loop is important for maintaining chronic lesions. However, chemokines (such as IL-8) and complement factors (such as C5a/C5a des Arg) are released by keratinocytes at the highly inßamed edges of lesions and are powerful attractants for neutrophils. Neutrophils express the human leukocyte antigen HLA-DR and subsequently induce the growth and differentiation of the surrounding keratinocytes

ETIOLOGY AND PATHOPHYSIOLOGY

583

Keratinocytes

HLA Cytokines Other Activated mediators

Activation of T cells and Keratinocytes

Production of chemoattractants (e.g., IL-8, C5a) Neutrophil

T cells

Leukocyte recruitment

Hyperproliferation

Blood vessel

T-cell adhesion

HLA = Human leukocyte antigen. IL = Interleukin.

FIGURE 2. Inflammation in psoriasis.

and activation of T cells. The activated T cells then induce the transendothelial migration of more neutrophils and set up a positive feedback loop that leads to excessive cytokine release and inßammation. Thus, the addition of the T-cell/neutrophil loop creates a unique environment at the border of plaques that can perpetuate acute inßammation (Figure 2). Cellular inÞltrates in skin lesions and PsA joints contain aberrantly active T-cell populations, supporting the hypothesis that psoriasis and its related pathology, PsA, are immunologic in origin. Indeed, impaired T-cell suppressor function; overexpression of the IL-2 receptor, HLA-DR, and various cell-adhesion molecules (CAMs); and proinßammatory cytokine (particularly IL-6) secretion are associated with psoriasis. Researchers theorize that these cytokines (secreted by activated T cells and other mononuclear inßammatory cells) induce hyperproliferation of epidermal and synovial Þbroblasts that, in turn, secrete IL-1β, IL-2, IL-6, and platelet-derived growth factor (PDGF), contributing to disease pathology. As in psoriatic skin lesions, various cytokines found in affected joints of people with PsA exhibit conßicting activities (e.g., IL-2 is pro-inßammatory, and IL-13 is anti-inßammatory). TNF-α. TNF-α is present in elevated levels in psoriatic skin plaques (along with the TH 1 cytokines IL-2, IFN-γ , IL-6, and IL-8). This overexpression of TNF-α is implicated as a key factor in keratinocyte activation and proliferation, as well as stimulating the overexpression of endothelial adhesion molecules. However, the pathogenic role TNF-α plays is not entirely clear. Research during the past decade has implicated T cells in the etiology and pathogenesis of psoriasis. In psoriatic skin plaques, mononuclear cells (principally T cells) inÞltrate the dermis and produce cytokines that cause keratinocyte hyperproliferation and plaque formation. Infection may trigger both skin lesions, and some researchers believe

584

PSORIASIS

that TNF-α produced by monocytes and macrophages during infection plays a major role in the T-cell response, causing activation and proliferation of T cells and serving as a link between etiology and pathogenesis. Although clinical studies have demonstrated the effectiveness of TNF inhibitors against psoriatic skin, their mechanisms of action are not clear. Experimental models have demonstrated a connection between TNF-α and infection in psoriatic skin. In one of the most convincing experiments, streptococcal and staphylococcal superantigens induced an inßammatory response in uninvolved skin from patients with psoriasis (but not other skin disorders), and TNF-α was dramatically upregulated in the epidermis, indicating that it plays a role in driving the inßammatory response (Travers JB, 1999). The precise role is not clear, however, and in one study, TNF-α had no effect on proliferation or differentiation of healthy and psoriatic keratinocytes (Fransson J, 2000). According to some researchers, this Þnding provides further evidence that TNF-α may act in psoriasis principally through its effects on T cells (among other inßammatory effects) and that T cells, rather than abnormal keratinocytes, cause the skin changes of psoriasis. Other inßammatory effects of TNF-α that may contribute to skin lesions and damage cartilage and bone in articular lesions include the following: increased adherence of T cells to postcapillary venules; activation and proliferation of polymorphonuclear leukocytes (PMNs, or neutrophils); expression of human leukocyte antigens (HLAs); and B-cell differentiation and antibody formation in joints. Aberrant Angiogenesis. Dermal capillary proliferation is a pathological characteristic of psoriatic lesions and explains why bleeding occurs easily when lesions are removed. Keratinocytes produce angiogenic factors—endothelial-cellstimulating angiogenesis factor (ESAF) and vascular endothelial growth factor (VEGF)—both of which have been reported to be elevated in patients with psoriasis, compared with controls; increases in these levels correlate with disease severity (Bhushan M, 1999). For example, according to results presented by Regeneron Pharmaceuticals at the 2002 Society for Investigative Dermatology in Los Angeles (Xia AY, 2002), VEGF-transgenic mice that chronically express VEGF in the skin developed a skin condition resembling psoriasis. Single-nucleotide polymorphisms in the VEGF gene were recently identiÞed in patients with early onset chronic plaque psoriasis, which also correlated with elevated plasma levels of VEGF (Young HS, 2004). One of the therapeutic mechanisms of retinoids in psoriasis may be the inhibition of angiogenesis; studies have demonstrated that retinoids inhibit the action of VEGF at the transcriptional level (Diaz BV, 2000). Researchers have worked extensively on elucidating the role of angiogenesis in cancer. Further study of a potential role of anti-angiogenesis is warranted because some of the many anti-angiogenic drugs in development as cancer therapeutics may prove useful in psoriasis, particularly if topical delivery is feasible. Table 3 lists mechanisms thought to be involved in the pathogenesis of psoriasis that serve as targets for drug development. These mechanisms are addressed in more detail in “Emerging Therapies.”

ETIOLOGY AND PATHOPHYSIOLOGY

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TABLE 3. Select Potential Therapeutic Targets in Psoriasis and Psoriatic Arthritis Targets of Developmental Agents TNF-α IL-1β IL-6 IL-8 IL-10 IL-11 Pyrimidine synthesis T-cell vaccines Complement components

Suspected Mechanisms of Action Inhibits T-cell activation and proliferationa Inhibits T-cell activation and proliferation Inhibits T-cell activation and proliferation Reduces recruitment of T cells, chemoattraction of T cells and neutrophils, angiogenesis, and keratinocyte proliferation Suppresses synthesis of pro-inflammatory cytokines Suppresses synthesis of pro-inflammatory cytokines Inhibits DNA and RNA synthesis Suppresses T-cell response Inhibits complement activation and reduces the overproduction of C5a observed in psoriasis

a See ‘‘TNF-α ’’ section for more details. IFN = Interferon. IL = Interleukin. TNF = Tumor necrosis factor.

Etiology The precise causes of psoriasis are not known. Genetic defects that cause aberrant inßammatory and immune-mediated responses are thought to be the basis of the abnormally rapid keratinocyte proliferation and inßammation that lead to the development of psoriatic lesions. Risk factors associated with psoriasis include compromised immunity, the presence of bacterial antigens, age, and environmental factors (e.g., climate, trauma, infection, diseases, psychological stress, concurrent medications). Genetic Factors. Substantial data derived from studies of familial inheritance and genetic association patterns with psoriasis suggest that genetics play a role in the development of this disease. Studies of family history have found that in approximately 35% of people afßicted with psoriasis, one or more family members also have the disorder. A study of the lifetime risk for developing psoriasis estimated the risks at 4% for a person with no family history, 28% when one parent is affected, and 68% when both parents are affected (Swanbeck G, 1997). Studies of twins similarly demonstrate increased risk; a concordance rate of 65% for monozygotic twins and 30% for dizygotic twins was observed in one study; other studies have derived different Þgures, but none have shown a rate of 100% concordance (Krueger GG, 1994; Henseler T, 1997). The lack of 100% concordance between monozygotic twins and other genetic Þndings demonstrate that additional factors play a role in the risk of developing psoriasis. Genetic factors are thought to increase an individual’s susceptibility to environmental risk factors such as psychological stressors, climate, and infections. Most researchers believe psoriasis to be a polygenic disorder, although the precise gene locus or loci associated with increased susceptibility or initiation of

586

PSORIASIS

psoriasis are not known. A potential role for the human leukocyte antigen (HLA) region on chromosome 6 was reported almost 30 years ago. More-recent studies have suggested roles for regions on chromosomes 1, 2, 3, 4, 8, 10, 16, 17, and 20. Some of these putative susceptibility loci have been named psoriasis susceptibility loci 1 through 8, or PSORS1-8. Most research, however, has focused on the major histocompatibility (MHC) class I region (coded for by genes on chromosome 6) and, in particular, the HLA-Cw6 allele (Nickoloff BJ, 1999; Guedjonsson JE, 2002). Other genes associated with psoriasis pathophysiology include the gene encoding the CD1d molecule, which is located near other implicated genes on chromosome 1, and genes that encode nonclassical MHC antigens, including MICA, PERB 11, and the S gene, which are located on chromosome 6. The importance of the MHC class I region was underscored by the International Psoriasis Genetics Study (The International Psoriasis Genetics Consortium, 2003). To conÞrm previously reported linkages to psoriasis, 942 affected sibling pairs from 710 family pedigrees were analyzed for polymorphic satellites spanning 14 psoriasis candidate regions. The alleles that appeared to be shared by affected siblings at the most statistically meaningful frequencies were located within the MHC class I region. Two other loci on chromosomes 10q (D10S2327 ) and 16q (D16S3032 ) were also identiÞed in the same study as potentially promising candidates for more detailed examination. Genotypic analysis of 644 subjects (from 195 Swedish families with a history of psoriasis) revealed an association of SCL12A8 (a gene that encodes a member of the solute carrier family 12 proteins) with psoriasis. This analysis used a single-nucleotide polymorphism (SNP) map spanning 900–1,200 kb of chromosome 3q21, which maps to the psoriasis susceptibility locus PSORS5. SCL12A8 belongs to a class of genes that was previously unrecognized as playing a role in psoriasis pathogenesis (Hewett D, 2002). Age. The onset of psoriasis symptoms can occur at any age, but two peaks in onset have been observed. The initial peak occurs during the teen years: approximately 40% of patients Þrst experience the disease before the age of 20, and initial onset before age 10 occurs in approximately 10% of cases. A second peak has been reported in patients who are in their mid 50s. Onset at an earlier age (before age 40) predicts a more severe disease course. An association between HLA haplotype and age of onset for psoriasis has been reported (Table 4). Climate. Cold, dry weather is known to provoke ßare-ups of psoriasis, whereas sunny, hot, and humid weather helps relieve symptoms in most patients. In the case of photosensitive psoriasis, however, the reverse is true: the sunlight of summer may exacerbate the condition, and symptoms may improve during winter. Trauma. Cuts, burns, injections, and even very mild abrasions and other trauma to the skin may cause psoriatic skin plaques to erupt. Koebner’s phenomenon—a hallmark of the disease—is the development of psoriatic plaques following mechanical trauma to nonlesioned skin and occurs in approximately 20% of

ETIOLOGY AND PATHOPHYSIOLOGY

587

TABLE 4. Select Genetic Factors in Psoriasis Genes

Comment

HLA-Cw6, HLA-B13, HLA-B17 HLA-B27, HLA-Cw2, HLA-B44, HLA-Cw5 Chromosome 3q21 (SCL12A8) Chromosome 10q22-q23 (D10S2327)

Associated with earlyonset psoriasis Associated with late-onset psoriasis Associated with psoriasis

Chromosome 16q (D16S3032)

Associated with psoriasis

Associated with psoriasis

Reference Szczerkowska-Dobosz A, 1996 Szczerkowska-Dobosz A, 1996 Hewett D, 2002 Nair RP, 1997; The International Psoriasis Genetics Consortium, 2003 Nair RP, 1997; The International Genetics Consortium, 2003

HLA = Human leukocyte antigen.

patients with psoriasis. Koebner’s phenomenon may explain the increased frequency of lesions on the elbows and knees. Psychological Stressors. A strong association between psoriasis and emotional disorders (including anxiety and depression), repressed anger, and psychological stress has been reported. Nearly 40% of patients in one study could recall a speciÞc stressor that occurred within one month of a psoriatic ßareup. Researchers are beginning to investigate potentially aberrant biochemical responses to stress in these patients. Infections. Both viruses and bacteria have been implicated as potential etiologic agents in the development of psoriasis. SpeciÞcally, streptococcal infections of the upper respiratory tract (e.g., “strep” throat, tonsillitis, sinusitis) are known to trigger the disease, particularly guttate (eruptive) psoriasis in individuals who express the MHC molecule HLA B-13. HIV infection has been linked to psoriasis; compromised immunity (i.e., T-cell suppression) is the suspected mechanism. Bacterial Superantigens. Bacterial superantigens (SAs) are bacterial products that nonspeciÞcally activate subsets of T cells by binding directly to MHC class II molecules and Vβ -chains of T-cell receptors outside of the antigen recognition domains of these molecules. This interaction of superantigens with T cells results in the nonspeciÞc activation of as much as 20% of all T cells—a percentage exponentially higher than that activated by an antigen that speciÞcally interacts with MHC or T-cell receptor molecules. SAs have been implicated in inßammatory skin disorders, such as atopic dermatitis. Researchers speculate that these SAs may contribute to the development of psoriasis through T-cell mechanisms. SigniÞcant levels of the gram-positive Staphylococcus aureus bacteria, which produces a potent SA, have been detected on the skin of more than 50% of patients with plaque psoriasis compared with 5–10% of the nonpsoriatic population. However, Þndings regarding the importance of bacterial SAs in exacerbating psoriasis do not conÞrm the role of SAs in the development of psoriasis. Despite increased humoral and cellular immunity against the gram-positive bacteria found in psoriatic plaques, a cause-and-effect relationship is not clear

588

PSORIASIS

(i.e., plaques may be colonized or secondarily infected, rather than caused by these organisms). Furthermore, the T-cell response to streptococcal antigens is similar in both PsA and rheumatoid arthritis (RA) patients, indicating that it may be a generalized response regardless of the etiology. Results from an in vivo study of the humoral (immunoglobulin G [IgG] antibody) response to Streptococcus pyogenes antigens (speciÞc recombinant proteins) suggest that streptococcal infection may be an etiologic agent in the development of PsA (Muto M, 1996[a]). Thus, T cells recognizing streptococcal antigens may cross-react with auto-antigens found in the skin, such as on keratinocytes. This phenomenon is perhaps most classically recognized in the case of the streptococcal M protein and proteins present on the heart. Here, cross-reactive T cells targeting the bacteria also target the heart, causing permanent heart damage. Concurrent Medications. Drugs that can exacerbate psoriasis include β-adrenergic blockers, angiotensin-converting enzyme (ACE) inhibitors, progesterone, lithium, systemic (or high-potency topical) corticosteroid withdrawal, and chloroquine. Nonsteroidal anti-inßammatory agents (NSAIDs) can also trigger psoriasis or exacerbate symptoms in some patients, although some NSAIDs (e.g., meclofenamate) improve the condition in certain patients. Drugs that cause skin rashes can also trigger psoriasis via Koebner’s phenomenon because of trauma induced by scratching the rash. CURRENT THERAPIES Psoriasis is a chronic disease for which there is no cure. Drug therapies and phototherapies may clear lesions and relieve uncomfortable symptoms, but remission is usually short-lived. Most patients experience one or more exacerbations or relapses per year. Exacerbations occur when drug therapy is discontinued (because of side effects and/or patient noncompliance) or tolerance to therapy develops, necessitating a change in therapies. Table 5 lists treatments in their typical order of application in the management of psoriasis, based on disease severity and response to therapy. Table 6 lists selected current therapies for psoriasis. Table 7 lists topical corticosteroids used in the treatment of psoriasis. Although many of the agents prescribed for psoriasis are used as monotherapies, the potential for adverse effects and tachyphylaxis, as well as the need to maximize efÞcacy, have often prompted physicians to resort to combination therapy. Recent clinical trials involving long-established drug classes have focused on identifying combinations of drugs that minimize adverse effects while maximizing clearance of plaques. Guidelines for using combination therapies are summarized in Table 8, and selected combinations are described in detail later in this section. Many of the topical treatments described in this section are used in combination with emollient and keratolytic agents that supply moisture and help remove scales. Because these agents are predominantly used as adjunct therapy, they are not described in detail here. Other agents that are prescribed only for limited subtypes of psoriasis, such as antibiotics prescribed for pustular psoriasis, are not discussed here.

CURRENT THERAPIES

589

TABLE 5. Treatments for Psoriasis Treatment

Common Members in Class

Frequency of Use

Topical therapies

Vitamin D3 analogues (calcipotriene, High: nearly all patients are treated others); topical corticosteroids; with topical agents retinoids (tazarotene); anthralin Phototherapies Ultraviolet B (UVB) light; UVB Moderate: prescribed only when monochromatic light (311 nm); UVB disease is moderate to severe or light + coal tar (Goeckerman when topical therapy fails regimen); UVB light + anthralin + coal tar (Ingram regimen) Psoralen + UVA light; psoralen Systemic Moderate: prescribed only when therapies + UVA light + UVB light; oral disease is moderate to severe or retinoids-acitretin (may be combined when topical therapy fails with UVB light or psoralen + UVA light); methotrexate; cyclosporine; alefacept; efalizumab

Corticosteroids Overview. Topical corticosteroids are Þrst-line treatments for psoriasis, particularly mild and mild-to-moderate psoriasis. They may also be used to supplement systemic therapies for patients with moderate-to-severe psoriasis. Patients readily accept topical corticosteroids because these agents—formulated into ointments, creams, lotions, and gels—cause no irritation, have no odor, and are frequently effective (especially in mild cases). Also, side effects at low doses are often minimal. Corticosteroids are available in a range of potencies and concentrations that permit physicians to optimize efÞcacy and minimize side effects. A comprehensive list of topical corticosteroids according to potency is provided in Table 7. The potency of a corticosteroid is assessed by its ability to induce vasoconstriction, with class I (superpotent) corticosteroids being the most potent and class VII (low potency) being the least potent. The corticosteroids prescribed for psoriasis are usually class I to class IV (medium potency) corticosteroids. The topical corticosteroids prescribed most frequently for psoriasis in the major markets are clobetasol propionate (Oclassen Pharmaceuticals’ Cormax, GlaxoSmithKline’s Temovate, Connetics’ Olux), optimized betamethasone propionate (Schering-Plough’s Diprolene), dißorasone diacetate (Dermik’s Psorcon), halobetasol (Bristol Myers Squibb’s Ultravate), and mometasone furoate (Schering-Plough’s Elocon). Because many of these agents have very similar properties, only betamethasone dipropionate and mometasone furoate, two of the most frequently prescribed topical corticosteroids, are discussed in detail here as representative examples of corticosteroids commonly used to treat psoriasis. Most of the topical corticosteroids used to treat psoriasis are available as generic drugs, particularly in the United States. The exceptions are mometasone furoate, halobetasol propionate, and the augmented form of betamethasone dipropionate, each of which will lose market exclusivity in the next ten years. Side effects associated with topical corticosteroid use include allergic contact dermatitis, skin atrophy (thinning), striae (stretch marks), telangiectasias (spider

590

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TABLE 6. Current Therapies Used for Psoriasis Agent

Company/Brand

Topical corticosteroids Betamethasone Schering-Plough’s dipropionate Diprolene, generics

Mometasone Schering-Plough’s furoate Elocon Vitamin D3 analogues Calcipotriene Leo Pharmaceuticals/ Bristol-Myers Squibb’s Dovonex

Retinoids Acitretin

Tazarotene

Roche’s Soriatane/Neotigason

Allergen’s Tazorac

Antimetabolites Methotrexate Wyeth’s Rheumatrex; generics Anthralin

Bioglan Pharma’s Micanol, generics

Daily Dose

Availability

For all topical corticosteroids, US, F, G, I, S, twice-daily application to start; UK, J empirical observations for optimal interval between dosing; use formulation from lowest-potency class that clears skin. US, G, S, UK, J

0.005% cream or ointment; topical application twice daily; not more than 5 mg per week (equal to 100 g of vehicle base); can use for up to eight weeks; not to be used directly after salicylic acid treatment; apply after UV treatment for combination therapy.

US, F, G, I, S, UK, J

Oral administration of US, F, G, I, S, 0.3–0.5 mg/kg/day initially; UK increased in 3–4 week intervals to 0.75 mg/kg/day; pustular psoriasis requires initial dose of 1 mg/kg/day tapered to 0.5 mg/kg/day for maintenance. 0.05–1.0% applied topically once US, F, G, I, S, daily at bedtime; can be used UK, J for up to 12 weeks. 10–25 mg once weekly in three US, F, G, I, S, doses every 12 hours for 36 UK, J hours 0.05–0.1% ointment used US, F, G, I, S, initially; increased in UK, J increments to 5% until plaques resolve

Immunosuppressants Cyclosporine Novartis’s Oral administration of 2.5–3.0 US, F, G, I, S, Neoral/Sandimmune, mg/kg/day in two divided UK, J SangStat’s SangCya doses, ≤5 mg/kg/day; dosage tapered once clinical effect is seen. T-cell modulators Alefacept Biogen Idec’s Amevive 7.5 mg once-weekly bolus IV or US 15 mg once-weekly IM Efalizumab Genentech/Xoma’s 1 mg/kg once-weekly SC US Raptiva

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TABLE 6. (continued) Agent TNF-α inhibitors Etanercept

Company/Brand Amgen/Immunex/ Wyeth’s Enbrel

Photosensitizers 8-methoxypsoralen ICN Pharmaceuticals’ (8-mop) Oxsoralen

Coal tar

Various

Daily Dose 50 mg/kg once-weekly SC

Availability US

Topical: applied 20 minutes F, G, I, S, UK, J prior to UVA treatment. Oral: 0.6–0.8 mg/kg taken 2 hours prior to UV treatment. Available in many US, F, G, I, S, concentrations; 5% works UK, J best.

IM = Intramuscular; IV = Intravenous; SC = Subcutaneous; UV = Ultraviolet. US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan.

veins), hypopigmentation, rosacea, acne, perioral dermatitis, and increased Þne hair growth. The side effects of a topical corticosteroid are directly proportional to its potency, though some evidence suggests that newer formulations may provide enhanced beneÞt-to-risk ratios. Topical corticosteroids must be used carefully near the eyes because of an increased risk of cataracts and glaucoma. Physicians must also be aware of the potential for systemic absorption, which may lead to adrenal suppression, a serious side effect that is often incorrectly associated only with systemic corticosteroids. The risk of systemic side effects increases as the surface area of skin treated (i.e., a larger quantity of drug) and potency of the corticosteroid used increases. Other factors that affect the risk of side effects from topical corticosteroids include the drug vehicle (which affects the quantity of corticosteroid available to the skin), the concomitant use of occlusion therapy, the integrity of the skin, and the area being treated (e.g., absorption is greater in skin folds). Oral corticosteroids used at doses considered effective for treating psoriasis are associated with adverse side effects. Therefore, they are very seldom prescribed for psoriasis and are not discussed here. Mechanism of Action. The beneÞcial actions of topical corticosteroids in psoriasis include vasoconstriction, immunosuppression, and antiproliferative and anti-inßammatory effects. The potency of topical corticosteroids is determined by their ability to induce blood vessel constriction, with more-potent corticosteroids inducing greater vasoconstriction. Vasoconstriction results in decreased vascular permeability of dermal capillaries. It decreases edema and erythema and reduces leukocyte migration into affected skin. Topical glucocorticoids prevent cellular proliferation by inhibiting DNA synthesis and subsequently blocking mitosis. The anti-inßammatory and immunosuppressive actions of corticosteroids result largely from the inhibition of particular functions of leukocytes. Corticosteroids enter cells, bind to glucocorticoid receptors, and are transported to the nucleus as a corticosteroid-receptor complex. The corticosteroid-receptor

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TABLE 7. Select Topical Corticosteroid Preparations Corticosteroida

Concentration (%)

Clobetasol propionate Betamethasone dipropionate (optimized) Diflorasone diacetate Halobetasol propionate

0.05 0.05

Temovate (c/o) Diprolene (c/o)

0.05 0.05

Psorcon (o) Ultravate (c/o)

II. High potency

Amcinonide Betamethasone dipropionate Fluocinonide Desoxymetasone Mometasone fuorate Diflorasone diacetate Halcinonide

0.1 0.05 0.05 0.25 0.1 0.05 0.1

Cyclocort (o) Diprosone (o) Lidex (g/c/o) Topicort (c/g/o) Elocon (o) Florone (o), Maxiflor (o) Halog (c)

III. Potent

Triamcinolone acetonide Fluticasone propionate Amcinonide Betamethasone dipropionate Diflorasone diacetate Halcinonide Fluocinonide Betamethasone valerate

0.1 0.005 0.1 0.05 0.05 0.1 0.05 0.1

Aristocort A (o) Cutivate (o) Cyclocort (c/l) Diprosone (c) Florone (c), Maxiflor (c) Halog (o) Lidex E (c) Valisone (o)

IV. Medium potency

Flurandrenolide Mometasone fuorate Triamcinolone acetonide Fluocinolone acetonide Hydrocortisone valerate

0.05 0.1 0.1 0.025 0.2

Cordran (o) Elocon (c) Kenalog (c) Synalar (o) Westcort (o)

V. Midstrength

Flurandrenolide Fluticasone propionate Betamethasone dipropionate Triamcinolone acetonide Hydrocortisone butyrate Fluocinolone acetonide Betamethasone valerate Hydrocortisone valerate

0.05 0.05 0.05 0.1 0.1 0.025 0.1 0.2

Cordran (c) Cutivate (c) Diprosone (l) Kenalog (l) Locoid (l) Synalar (c) Valisone (c) Westcort (c)

VI. Mild

Aclometasone dipropionate Triamcinolone acetonide Desonide Fluocinolone acetonide Hydrocortisone

0.05 0.1 0.05 0.01

Aclovate (c/o) Aristocort (c) DesOwen (o), Tridesilon (c) Synalar (c)

2.5, 1

Hytone (c/o) (generics)

Class I. Superpotent

VII. Low potency Hydrocortisone, dexamethasone, flumethasone, prednisolone, and methylprednisolone

Brand Name

a Different preparations of the same corticosteroid (ointment, cream, lotion, gel) can be members of different

potency classes. c = Cream; g = Gel; l = Lotion; o = Ointment.

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TABLE 8. Combination Therapies for Psoriasis

Therapy

Weeks Required for Full Efficacy of Monotherapy Efficacy of to (1) Clear Lesions and Monotherapy (2) Maintain Clearance

Anthralin

3–16

(1) Moderate to excellent (2) Moderate

Anthralin + acitretin Anthralin + calcipotriol Anthralin + cyclosporine Anthralin + methotrexate Anthralin + coal tar

Acitretin

Recommended Recommended Recommended Recommended Recommended to strongly recommended 8–12

(1) Poor (2) Excellent

Acitretin + cyclosporine Acitretin + coal tar Acitretin + methotrexate Acitretin + calcipotriol Acitretin + PUVAa Acitretin + UVB Calcipotriol

Contraindicated Recommended Recommended Strongly recommended Strongly recommended Strongly recommended 8

(1) Moderate to excellent (2) Moderate

Calcipotriol + coal tar Calcipotriol + methotrexate Calcipotriol + topical corticosteroids

Recommended Recommended Recommended to strongly recommended Strongly recommended Strongly recommended Recommended

Calcipotriol + cyclosporine Calcipotriol + PUVAa Calcipotriol + UVB Coal tar

Variable

(1) Moderate (2) Poor

Coal tar + methotrexate Coal tar + topical corticosteroids Coal tar + PUVAa Coal tar + UVB

Cyclosporine Cyclosporine + coal tar Cyclosporine + topical corticosteroids Cyclosporine + methotrexate Cyclosporine + PUVAa Cyclosporine + UVB

Guidelines for Using Combination

Recommended Recommended Contraindicated Recommended to strongly recommended 4–8

(1) Excellent (2) Not recommended Recommended Recommended Contraindicated Contraindicated Contraindicated

594

PSORIASIS

TABLE 8. (continued)

Therapy Topical corticosteroids

Weeks Required for Full Efficacy of Monotherapy Efficacy of to (1) Clear Lesions and Monotherapy (2) Maintain Clearance 2–4

(1) Good (2) Excellent

Topical corticosteroids + acitretin Topical corticosteroids + methotrexate Topical corticosteroids + PUVAa Topical corticosteroids + anthralin Topical corticosteroids + UVB PUVAa

Recommended Recommended Recommended Recommended Not recommended 4–8

(1) Excellent (2) Not recommended

PUVA + anthralin PUVA + methotrexate UVB

Recommended Contraindicated 3–8

(1) Good (2) Good

UVB + methotrexate UVB + anthralin Methotrexate

Guidelines for Using Combination

Recommended Not recommended 8–16

(1) Excellent (2) Excellent

a The most common form of UVA phototherapy includes pretreatment with the photosensitizing agent pso-

ralen and is known by the acronym PUVA. Source: Adapted from Van de Kerkhorf PC. Combinations and comparisons. Clinical Dermatology. 1997;15:831–834; and Lebwohl M, et al. Combination therapy to treat moderate to severe psoriasis. Journal of the American Academy of Dermatology. 2004;50:416–430.

complex modulates transcription of corticosteroid responsive genes. Inhibited genes include those encoding the inßammatory cytokines IL-1, IL-2, IL-3, IL-6, granulocyte-macrophage colony-stimulating factor (GM-CSF), IFN-γ , and TNFα, which are produced by T cells (TH 1 and TH 2), B cells, monocytes, and macrophages. Corticosteroids also inhibit the expression of AP-1 and nuclear factor-kappa B (NF-κB), transcription factors that regulate expression of several inßammatory cytokines. Formulation. Topical corticosteroids are available as ointments, creams, lotions, and foams. Solutions for the treatment of scalp lesions are also available. Various vehicles are used in topical corticosteroid medications. Neutral vehicles, such as petrolatum, are popular. Some physicians avoid propylene glycol, which enhances penetration of the active drug, because it can irritate the skin. Halogenated corticosteroids (ßuorinated or chlorinated) are seldom prescribed. Although a halogen increases potency, it also increases systemic absorption and the potential for

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systemic side effects. Newer, highly lipophilic formulations (e.g., ßuticasone propionate and mometasone furoate) have been developed to promote absorption through the skin and increase potency while promoting rapid metabolism to inactive metabolites in the circulatory system and limiting systemic effects. Betamethasone Dipropionate. Betamethasone dipropionate (Schering Plough’s Diprolene/Diprosone, generics) is among the most popular topical corticosteroids. Betamethasone dipropionate is available in a variety of formulations and vehicles that range in potency from class I to class IV. Augmented betamethasone dipropionate ointment is a class I corticosteroid known for its rapid onset of action. The formulation differs from other formulations of the drug because it contains propylene glycol, which enhances skin penetration. Like other topical corticosteroids, betamethasone dipropionate has antiinßammatory and antiproliferative properties. Betamethasone dipropionate suppresses the production of inßammatory cytokines and reduces leukocyte migration into psoriatic skin. Dosage of class I corticosteroids must be managed carefully to minimize adverse effects. The FDA-approved labeling for augmented betamethasone dipropionate and for the class I corticosteroids clobetasol propionate and halobetasol propionate recommends limiting the duration of treatment with these agents. Pulse therapy, a limited, long-term dosing strategy, involves an initial period of daily application of a class I corticosteroid until the plaque being treated is ßattened (Katz HI, 1991). In a randomized double-blind, placebo-controlled study, an intermittent dosing regimen was tested for augmented betamethasone dipropionate (Katz HI, 1991). The initial phase of the study was an open-label screening phase during which psoriasis patients with ≤10% skin involvement applied augmented betamethasone dipropionate to their lesions twice a day for three to four weeks. After this initial phase, study participants were qualiÞed to proceed to the maintenance phase of the trial if their overall disease status was cleared or slight. Ninety of the original 123 patients enrolled in the trials were evaluated for efÞcacy during the maintenance phase of treatment. Of these, 46 were treated with augmented betamethasone and 44 were treated with placebo. Patients applied betamethasone or placebo three times per week at 12-hour intervals for a maximum treatment period of six months. Evaluation of clinical beneÞt was based on the length of time until treatment failure occurred, an end point assessed by changes in weighted scores of disease signs, overall disease status, and the patients’ evaluation of treatment effectiveness. Sixty-Þve percent of the patients treated with augmented betamethasone remained in remission for the duration of the sixmonth treatment period, while 20% of the latter group remained in remission. None of the patients in either treatment group exhibited any signs of cutaneous atrophy at the end of treatment. No trends suggestive of HPA axis suppression were evident during the course of the six-month treatment period. Therefore, intermittent therapy with high-potency topical corticosteroids appears to provide an extended period of disease remission while minimizing adverse effects.

596

PSORIASIS

FIGURE 3. Structure of mometasone furoate.

Betamethasone dipropionate is frequently combined with other topical medications, including vitamin D3 analogues and topical retinoids, to reduce the effective dose of each agent, improve overall efÞcacy, and extend the period of remission following treatment. Topical corticosteroids can also alleviate skin irritation induced by vitamin D3 analogues or topical retinoids. Like all topical corticosteroids, betamethasone dipropionate can cause hypothalmic-pituitary-adrenal (HPA) axis suppression when applied under occlusive dressings or when used daily for extended periods of time. As with other topical corticosteroids, skin atrophy, thinning, and striae are possible, particularly at high doses or when used for extended periods. Tachyphylaxis is a concern with betamethasone dipropionate, as it is with other topical corticosteroids. Mometasone Furoate. Mometasone furoate (Schering-Plough’s Elocon) (Figure 3) is a class II corticosteroid formulated as an ointment and a class IV corticosteroid formulated as a cream. Mometasone furoate has a relatively low potential for producing local and systemic side effects, including adrenal suppression. Like other topical corticosteroids, mometasone furoate enters cells and binds to the corticosteroid receptor. The corticosteroid-receptor complex modulates transcription of corticosteroid responsive genes, including inßammatory cytokines, AP-1, and NF-κB. Mometasone furoate is believed to penetrate human skin slowly, allowing for less frequent applications of the agent. Because mometasone furoate has been on the market for some time, few randomized clinical trials using contemporary methodology and end points are available to detail the safety and efÞcacy of the drug. However, a comparison with well-established agents indicated that mometasone furoate can induce similar clearing of psoriatic lesions with less frequent application than triamcinolone acetonide or ßuocinolone acetonide. The combined results of four randomized, single-blind, parallel-group studies compared the efÞcacy of mometasone furoate ointment and cream with the ointment and cream formulations of ßuocinolone acetonide and triamcinolone acetonide (Medansky RS, 1988). A total of 823 patients with chronic moderate to severe psoriasis were enrolled in the study and followed for 21 days of treatment. Mometasone ointment applied once daily was found to have a more rapid onset of action and produced a greater improvement in

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disease than triamcinolone ointment applied twice daily or ßuocinolone ointment applied three times daily. Mometasone cream applied once daily demonstrated a similarly rapid onset of action, was more effective in reducing signs of disease than ßuocinolone cream applied three times daily, and showed comparable efÞcacy with triamcinolone cream applied twice daily. Therefore, mometasone furoate produced comparable or superior results with a more favorable dosing schedule. A limited number of side effects were judged to be related to treatment with mometasone furoate. The most commonly observed were burning, itching, and dryness, which occurred at similar or lower frequency compared with triamcinolone or ßuocinolone (Medansky RS, 1988). Mild to moderate instances of skin atrophy were observed in less than 1% of patients treated with mometasone, in 2.5% of the patients treated with triamcinolone, and in no patients treated with ßuocinolone. Clinical laboratory tests revealed few values outside of the normal range, and these were deemed to be of no clinical signiÞcance or were judged by investigators to be unrelated to treatment. Therefore, once-daily application of mometasone furoate was judged to be safe. Like all topical corticosteroids, mometasone furoate should not be used at very high doses. The FDA-approved labeling for mometasone furoate cautions that treatment of greater than 30% of body surface area can result in HPA axis suppression. Also, tachyphylaxis, skin thinning, skin atrophy, and striae are potential side effects of treatment with mometasone furoate. Vitamin D3 Analogues Overview. Topical preparations of vitamin D3 analogues are Þrst-line treatments for mild psoriasis, particularly in children, because they do not present the risk of HPA axis suppression. Few adverse effects are observed. The most common adverse effect observed with topical vitamin D3 analogues is contact irritation (for this reason, topical use is contraindicated in sensitive areas such as the face and groin), but combining a vitamin D3 analogue and a topical corticosteroid can ameliorate the irritation. Oral vitamin D3 supplements cause adverse hypercalcemic effects and are therefore seldom prescribed. Calcipotriene (Leo Pharmaceuticals/Bristol-Myers Squibb’s Dovonex) is the most widely used vitamin D3 analogue, but two other agents, tacalcitol and calcitriol, are available outside the United States. Tacalcitol (Teijin’s Bonalfa), a topical vitamin D3 analogue, is frequently prescribed in Japan and in some European countries, but is not as widely available as calcipotriene. Calcitriol is available in both topical (Savient/Galderma’s Silkis) and oral (Roche’s Rocaltrol) formulations. Orally administered calcitriol has been largely displaced by calcipotriene because of the hypercalcemic effects associated with systemic administration of the former. In an effort to counter these negative effects, calcitriol has been reformulated as an ointment; nonetheless, calcipotriene is likely to remain the most widely used vitamin D3 analogue and is thus the agent discussed in detail in this section.

598

PSORIASIS

FIGURE 4. Structure of calcipotriene.

Mechanism of Action. Vitamin D3 and vitamin D3 analogues suppress growth and stimulate the terminal differentiation of keratinocytes. The high-afÞnity vitamin D receptor (VDR), a mediator of transcription, is expressed in a number of cell types, including all epidermal layers of skin, T lymphocytes, macrophages, and Langerhans’ cells. Several genes involved in cell proliferation and differentiation and in inßammation show altered expression levels when exposed to vitamin D3 analogues. Expression of involucrin, a marker protein of keratinocyte differentiation, increases, as does expression of the anti-inßammatory cytokine IL-10, while expression of the proinßammatory cytokine IL-8 decreases. Calcipotriene. Calcipotriene (Figure 4) is a vitamin D3 analogue that blocks proliferation of skin cells, enhances keratinocyte maturation, and exhibits antiinßammatory effects. It is available formulated as an ointment or a cream and is also available in a premixed combination with the topical corticosteroid betamethasone dipropionate (Leo Pharmaceuticals’ Dovobet). Calcipotriene is currently marketed exclusively as a branded drug, but will lose market exclusivity in the United States in 2007; in France in 2009; in Germany, Italy, and Spain in 2006; and in Japan in 2010. The efÞcacy and safety of calcipotriene were examined in a randomized, double-blind, right/left comparison of calcipotriene ointment (50 µg/g) on one side of the body and a placebo (vehicle) ointment on the other side (Dubertret L, 1992). Sixty-Þve patients were evaluated using a modiÞed Psoriasis Area and Severity Index (PASI) score, a measure used to evaluate the severity of psoriasis and response to therapy. Participants were seen at two-week intervals for a period of four weeks. While the average PASI score decreased on both sides of the body, a signiÞcantly greater improvement was observed with calcipotriene. The most frequent adverse events were lesional irritation, and one patient treated with calcipotriene became marginally hypercalcemic. Vitamin D3 analogues are often combined with topical corticosteroids. Leo Pharmaceuticals has developed a topical preparation combining the corticosteroid betamethasone dipropionate (0.5 mg/g) with calcipotriol (50 µg/g)

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(Dovobet/Daivobet). Clinical trial data support the effectiveness of this combination therapy. A randomized, double-blind trial enrolling 1,103 patients presented at the 60th Annual American Academy of Dermatology in New Orleans conÞrmed that this combination was superior to either agent alone. The average improvement in PASI score at the end of the four-week trial with the combination ointment was superior to that observed with either agent alone: 74.4% improvement for the combination, 55.3% improvement for calcipotriol, and 61.3% improvement for the steroid (p < 0.001) (Gollnick H, 2002). This combination appears to provide efÞcacy similar to that of individually applied corticosteroids and vitamin D3 analogues but with only a single application per day—an improvement over current dosing regimens (Kragballe K, 2002). Combining topical corticosteroids and vitamin D3 analogues can also improve the duration of remission observed with topical corticosteroids. In a doubleblind, placebo-controlled, parallel group study, 44 patients with mild to moderate psoriasis were treated with a two-week combination regimen of calcipotriene ointment in the morning and halobetasol ointment (a class I steroid) in the evening (Lebwohl M, 1998). From that group, 40 patients demonstrated at least a 50% improvement on a 0 to 8 scale used to score disease severity, and those 40 patients were rerandomized to two groups: 20 treated with halobetasol twice daily on weekends and calcipotriene twice daily on weekdays, and 20 that received halobetasol ointment only. Of the patients treated with the combination regimen, 76% remained in remission for six months, compared with 40% treated with halobetasol. Calcipotriene causes local skin irritation and is therefore not suitable for use on the face, groin, or other sensitive areas of the body. At very high doses, as seen in patients using more than 200 grams of ointment (50 µg/g of ointment) per week, patients may become hypercalcemic. Elevated calcium levels return to normal with cessation of treatment. Retinoids Overview. The topical and oral retinoids used to treat psoriasis are synthetic vitamin A derivatives. The oral retinoid etretinate (Roche’s Tegison/Tigason) has largely been replaced by the newer oral retinoid acitretin (Roche’s Soriatane/Neotigason), but it is the only oral retinoid available in Japan. The primary difference between acitretin and etretinate is the period of time the drug is retained by the body after treatment is discontinued. Etretinate is still detectable for three years after therapy is discontinued, while acitretin drops to undetectable levels within a few weeks. Because etretinate is no longer available in most of the markets under study, it will not be discussed in detail here. Mechanism of Action. Retinoids bind to speciÞc receptors called retinoid receptors (RARs and RXRs) in the cell nucleus, affecting the transcription of genes involved in keratinocyte differentiation, cell proliferation, and inßammation. Clinical evidence and in vitro studies demonstrate that retinoids downregulate transcription of proinßammatory cytokines, halt proliferation of cultured cells, and alter expression of indicators of keratinocyte differentiation.

600

PSORIASIS

Acitretin. Acitretin (Roche’s Soriatane/Neotigason) is marketed in the United States and Europe for severe psoriasis and is often prescribed for patients who do not respond to other treatments. Acitretin is effective in severe psoriasis, particularly pustular and erythrodermic psoriasis. This agent is often used in combination with other psoriasis therapies, including topicals and UVB light or PUVA. However, combining acitretin and cyclosporine requires close monitoring because both agents can raise triglyceride and serum cholesterol levels. While acitretin is currently marketed as a branded drug, patent protection and market exclusivity have expired in all markets under study. No generics manufacturer has announced plans to market a generic version of acitretin to date. Acitretin, a second-generation oral retinoid, is the principal metabolite of etretinate, the Þrst retinoid that was developed to treat psoriasis. Acitretin activates the retinoid receptors RAR-α, RAR-β, and RAR-γ , but does not activate the retinoid receptors RXR-α, RXR-β, and RXR-γ (Duvic M, 2004). Activation of RAR-α is associated with adverse effects, including elevated serum lipid and triglyceride levels (Duvic M, 2004). It is not yet known whether metabolites of acitretin activate the RXR receptors, which are also associated with adverse effects. Acitretin downregulates transcription of proinßammatory cytokines, halts proliferation of cultured cells, and alters expression of indicators of keratinocyte differentiation. The efÞcacy of acitretin was compared with that of its predecessor, etretinate, in a randomized, double-blind study in Germany (Gollnick H, 1988). A total of 175 patients with severe psoriasis of different types were treated with 10 mg, 25 mg, or 50 mg of acitretin and compared with patients receiving 50 mg etretinate over a period of eight weeks. Complete remission occurred among patients receiving 50 mg of acitretin or etretinate, and an average improvement in PASI score of 53.8% was seen with acitretin, compared with 61.1% with etretinate. However, enlargement of psoriatic lesions was observed in some patients with both agents despite overall improvements in PASI scores. Like all systemic retinoids, acitretin has potentially serious side effects similar to those observed with megadoses of vitamin A. Common, less serious side effects include dry eyes, nose, and lips; peeling skin of the palms and soles; thinning hair; bruising and nose bleeds; bone and joint pain; and headache. Vision-related symptoms may include decreased night vision, blurred vision, and cataracts. Adverse musculoskeletal effects include bone overgrowth in the knee and ankle joints and the pelvic area. Cholesterol levels may also rise. More rarely, patients experience nausea, vomiting, and buildup in intracranial pressure. Acitretin is teratogenic and should not be used by women who may become pregnant. Concomitant ingestion of alcohol with acitretin therapy results in its conversion to etretinate, a form retained by the body for several years; thus, alcohol use is contraindicated for women of childbearing age taking acitretin. Although acitretin is cleared more rapidly than etretinate, it is nevertheless recommended that women taking acitretin avoid becoming pregnant for three years after therapy is discontinued. Because teratogenicity is a serious concern, acitretin is seldom prescribed to women of childbearing age.

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Tazarotene. Tazarotene (Allergan’s Tazorac), the Þrst topical retinoid found to be effective for moderate psoriasis, is used either as a single agent or in combination with UV light or anthralin. Tazarotene acts rapidly; improvement often occurs within one week of use and is generally seen within 10–12 weeks, and success rates of up to 70% of treated patients have been reported. Used either alone or in combination with topical corticosteroids, tazarotene can induce longer periods of remission compared with topical steroid monotherapy (Lebwohl M, 1998). Local skin irritation, pruritus, and erythema may occur, but topical retinoids are considered safe and are not prone to the serious side effects that corticosteroids may cause. Topical tazarotene is currently marketed only as a branded drug; market exclusivity will expire in all markets discussed here during the next ten years. Tazarotene is a third-generation retinoid that selectively binds to retinoic acid receptor (RAR)-β and RAR-γ for the treatment of moderate to severe psoriasis. Binding to tazarotene’s cognate RARs downregulates transcription of proinßammatory cytokines and suppresses keratinocyte proliferation. Because it does not activate RAR-alpha and retinoid X receptors (RXRs), oral tazarotene is not associated with some side effects—such as hypertriglyceridemia, hypothyroidism, and hepatoxicity—that occur with other oral retinoids (Duvic M, 2004). Two multicenter, double-blind, randomized, vehicle-controlled studies of the safety and efÞcacy of tazarotene cream evaluated a total of 892 patients with mild to moderate plaque psoriasis, deÞned as involving at least 2% of the total body surface area and lesions of at least moderate severity (Weinstein GG, 2003). Clinical success was evaluated using the Overall Lesional Assessment (OLA), a measure that scores a patient’s psoriasis with respect to plaque elevation, scaling, and erythema. An OLA score of none, minimal, or mild was the primary efÞcacy variable. Patients were treated with 0.1% or 0.05% tazarotene cream or with vehicle for a period of 12 weeks and were followed for an additional 12 weeks after completing treatment. More patients treated with tazarotene achieved clinical success than did patients treated with placebo at both concentrations of tazarotene and in both studies, with success rates ranging from 39.4% to 50.7%, versus values of 24.5% and 26.2% for vehicle. The therapeutic effect of tazarotene continued to be statistically signiÞcant throughout the post-treatment period. Topical tazarotene can cause local skin irritation and therefore cannot be used in sensitive areas such as the face or groin. Topical tazarotene should not be used by pregnant women because of the risk of teratogenicity, and it can increase sensitivity to sunlight, requiring that patients avoid excessive sun exposure. Antimetabolites Overview. Antimetabolites interfere with the rapid proliferation of cells and decrease inßammation associated with psoriasis. Antimetabolites exert their effects largely by interrupting cellular metabolism, particularly nucleic acid synthesis. Methotrexate (Wyeth’s Rheumatrex, generics), a systemic antimetabolite, and anthralin, a topical agent, are the antimetabolites most commonly used to treat psoriasis.

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H2N

N

N CH3

N

N N H N

NH2 O

COOH COOH

FIGURE 5. Structure of methotrexate.

Systemic antimetabolites can be extremely successful at improving severe cases of psoriasis, but toxicity limits their use. Methotrexate is the most extensively used agent in this class, but it requires careful monitoring of liver function in patients who receive long-term therapy. Other systemic antimetabolites, such as 6-thioguanine and hydroxyurea, are prescribed in rare instances where other therapies have failed, but high toxicity and signiÞcant adverse effects limit their use. Because they are used very infrequently, 6-thioguanine and hydroxyurea are not discussed here. Mechanisms of Action. The most commonly used antimetabolites halt division of proliferating cells by interfering with nucleic acid metabolism and blocking DNA synthesis. Methotrexate. Methotrexate (Wyeth’s Rheumatrex, generics) (Figure 5) is a synthetic analogue of folate, a vitamin that is essential for production of purine DNA bases. Methotrexate is an antimetabolite and chemotherapy drug that interferes with cell proliferation by blocking purine nucleotide synthesis and exerts anti-inßammatory actions. It is effective in, and usually reserved for, patients with severe disease that is resistant to topical therapy and phototherapy. Methotrexate therapy decreases the epidermal cell proliferation rate, monocyte and neutrophil chemotaxis, Langerhans’ cell activity, and leukotriene B4 production by neutrophils. Methotrexate has been approved for the treatment of psoriasis since 1972, and few randomized clinical trials measuring its efÞcacy using contemporary study end points are available from that era. However, it is widely regarded by clinicians as a very effective treatment for severe psoriasis. In a retrospective case-report study, researchers found that among 113 psoriatic patients treated over a 20-year period in one clinic with 15 mg/week of methotrexate, 82% of patients had “prolonged clearance or near clearance” (Van Dooren-Greebe RJ, 1994). Methotrexate is frequently prescribed for patients who fail topical therapy and UV therapy and is often part of combination or rotational regimens. Methotrexate is usually well tolerated in low doses, but it can cause many side effects and has the long-term potential of damaging the liver. Nausea and vomiting, skin rash, itching, and stomatitis (mouth sores) are the most common side effects. Gastrointestinal (GI) side effects are minimized by the use of H2 histamine antagonists, and stomatitis can be minimized by concomitant folic acid administration.

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FIGURE 6. Structure of anthralin.

Potential hepatotoxicity is of great concern, and the total dose must be carefully monitored. Liver enzymes may become elevated, necessitating (usually monthly) monitoring. The American Dermatology Society recommends that a liver biopsy be performed after administration of the Þrst 2.5 g of methotrexate and then at every 1.5 g thereafter. Very rarely, frank liver Þbrosis and cirrhosis may develop, necessitating liver biopsy. Liver damage is most likely to occur in the subset of patients who have received a cumulative dose of more than 1.5 g and demonstrate other risk factors. These factors include taking NSAIDs or other medications that are contraindicated during methotrexate therapy; alcoholism or substance abuse; existing liver or kidney problems; diabetes mellitus; obesity; or old age. Anthralin. Anthralin (Figure 6), or dithranol (Bioglan Pharma’s Micanol, generics), a topical antimetabolite, is a synthetic derivative of chrysarobin (obtained from the Brazilian tree Andira araroba) and has been used to treat psoriasis since the mid 1800s. Anthralin is very effective for mild to moderate psoriasis. As a photosensitizer, it is sometimes used in combination with UV light monotherapy (without psoralen). Although the mechanism of action of anthralin is not well understood, it exerts an anti-inßammatory action, promotes cellular differentiation, and reduces cellular proliferation (as demonstrated by changes in markers of psoriasis, T-cell activation, and chemotaxis). In recent years, conventional applications of anthralin have been replaced by short-contact anthralin therapy, in which a high concentration of anthralin (1% or greater) is applied for several minutes to one hour. However, comparisons with corticosteroids and vitamin D3 analogues have revealed few efÞcacy advantages, and patients favor other agents that are less irritating and are less likely to stain skin or clothing. In a multicenter, randomized, open study, 306 patients of either gender with stable chronic plaque psoriasis covering at least 100 cm2 of surface area applied calcipotriol (50 µg/mg) twice daily or shortcontact anthralin (0.1–2%) for up to three months. The number of patients who were assessed by physicians as being “cleared” or showing “marked improvement” at the end of treatment was greater in the calcipotriene group than in the anthralin group (60.1% versus 51.1%), but both groups showed improved quality of life as assessed by the Psoriasis Disability Index (PDI) and the Sickness Impact ProÞle (SIP) (Wall AR, 1998). However, calcipotriene provoked less contact irritation, and patients found it more appealing to use. Anthralin can cause nonpsoriatic skin to become irritated and inßamed. While anthralin is not known to cause any serious adverse effects, it does not offer

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efÞcacy advantages over other topical treatments, and other agents that are less prone to staining clothing are preferred by patients. Immunosuppressants Overview. Immunosuppressants may be very effective in psoriasis, but their use is limited to severe cases because of potentially serious side effects. Cyclosporine (Novartis’s Sandimmune/Neoral, SangStat’s SangCya, generics) is the immunosuppressant most widely used to treat severe cases of psoriasis. More toxic drugs, such as mycophenylate mofteil and azathioprine, are used infrequently because of high toxicity and only moderate efÞcacy. Mechanisms of Action. Cyclosporine and similar immunosuppressants bind to cytoplasmic receptors, which are isomerases known as immunophilins. The drug-immunophilin complex exerts its primary immunosuppressive activity by binding to and inhibiting calcineurin, a protein that plays an essential role in calcium-dependent signal transduction in T cells. This signal transduction pathway activates expression of cytokine genes, among others. Decreased cytokine production concomitantly decreases T-cell (TH 1 and TH 2) activation, including IL-2 production, growth, migration, and cytotoxic T-cell degranulation. Reduced cytokine secretion also affects the functions of keratinocytes, antigen-presenting cells, and neutrophils. Calcineurin is involved in T-cell-derived IL-2, IL-4, GMCSF, TNF-α, and IFN-γ production. Therapy with cyclosporine or tacrolimus reduces the synthesis of all of these cytokines. See “Pathophysiology” for more information on the role of cytokines in psoriasis. Cyclosporine. Cyclosporine (Novartis’s Sandimmune [an oil-in-water emulsion] and Neoral [a microemulsion], SangStat’s SangCya [a microemulsion], generics) plays an important role in the treatment of psoriasis. The effectiveness of cyclosporine for psoriasis was discovered serendipitously and was a pivotal Þnding with regard to the development of T-cell hypotheses of the pathogenesis of psoriasis (see “Pathophysiology”). Cyclosporine binds to an intracellular receptor, the immunophilin cyclophilin. The immunophilin-cyclosporine complex inhibits the phosphatase, calcineurin, and cytokines activated by calcineurin. Decreased cytokine production leads in turn to decreased T-cell activation and migration. Cyclosporine is often effective for refractory psoriasis and must be administered systemically (orally). Efforts to develop a topical formulation have failed, principally because cyclosporine is a large (>1,200 KDa) molecule, limiting its absorption. Because of potentially serious side effects, cyclosporine therapy is titrated upward until remission is induced and then downward to the lowest effective maintenance dose. Therapy is initiated at 3 mg/kg/day; if no response is obtained in one month, the dose may be titrated upward to 4 mg/kg/day; 5 mg/kg/day is the maximum dose. Following the induction of remission, the dose is adjusted downward by 0.5 mg/kg/day every two weeks to reach the lowest effective maintenance dose. Topical agents may be used to help maintain remission. Relapse

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occurs two to four months after cyclosporine discontinuation in virtually all patients. A 16-week trial involving 181 patients with severe psoriasis evaluated the ability of cyclosporine to induce and maintain disease remission (Shupack J, 1997). After 16 weeks of treatment with an initial dose of 5 mg/kg/day, 86% of patients achieved 70% or greater reduction in body surface area involvement. Responders were randomized to receive cyclosporine or placebo daily for six months; 42% of patients who received 3 mg/kg cyclosporine, compared with 84% in the placebo group, relapsed at the end of the six-month maintenance period. Disease relapse was marked by a 50% or greater return in skin surface involvement relative to baseline. The median time to relapse was six weeks in both the placebo and 1.5 mg/kg groups and was delayed in the 3.0 mg/kg group; 58% of patients that received the 3.0 mg/kg dose of cyclosporine remained relapse-free at end of this six-month maintenance period, compared with 16% of patients who received placebo. The efÞcacy of cyclosporine and methotrexate were compared in a randomized, controlled trial involving 88 patients with moderate to severe psoriasis (Heydendael MD, 2003). Patients were treated for 16 weeks and followed for an additional 36 weeks after treatment was discontinued. PASI scores were determined at baseline and monthly thereafter by trained assessors unaware of the treatment assignments. After 16 weeks of treatment, the reduction in PASI scores from baseline was 64% in the methotrexate group and 72% in the cyclosporine group. Almost complete remission (deÞned as a reduction in baseline score for the PASI index of more than 90%) was achieved in 40% of the patients in the methotrexate group and 33% of patients in the cyclosporine group. Partial remission (deÞned as a reduction in baseline score for the PASI index of more than 75%) was achieved in 60% of patients in the methotrexate group and 71% of patients in the cyclosporine group. The time to reach these degrees of remission was not signiÞcantly different in either group, with maximum degrees of clearance reached at weeks 12–16. Methotrexate and cyclosporine therefore appear to have comparable efÞcacy for treating moderate to severe psoriasis. Alterations in renal function occur in approximately 25% of cyclosporinetreated patients, necessitating blood monitoring for kidney dysfunction. Liver toxicity is also a potential side effect, and cyclosporine therapy is therefore contraindicated in patients with liver disease. GI side effects may include nausea, vomiting, anorexia, and diarrhea. Headache, sensory disturbances in the extremities, musculoskeletal pain, ßu-like symptoms, and upper-respiratory-tract infections are common side effects. The hematologic and metabolic disturbances associated with cyclosporine treatment, including alterations in blood coagulation and lipid levels, are more relevant to transplant patients than to psoriatics. Blood pressure may increase by up to 13 mm Hg systolic and 5 mm Hg diastolic. Study authors have reported that the risk of developing skin cancer may be elevated with cyclosporine treatment, but increased risk caused by concomitant UVA light treatments may have been a confounding variable. Dermatologic side effects of cyclosporine include hypertrichosis, acne eruptions, keratosis pilaris,

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and sebaceous hyperplasia. Adverse effects occur with the same frequency in all available cyclosporine preparations. T-Cell Modulating Agents Overview. T-cell modulating agents represent the Þrst generation of biological agents available to treat psoriasis. The systemic agents alefacept (Biogen Idec’s Amevive) and efalizumab (Xoma/Genentech’s Raptiva, previously known as Xanelim) received FDA approval in 2003, and efalizumab was recommended for approval in the European Union in June 2004. T-cell modulators are a signiÞcant addition to the arsenal of agents available to treat moderate to severe psoriasis, particularly in patients who have failed other therapies. Mechanism of Action. Cell-adhesion molecules (CAMs) play integral roles in all phases of inßammatory and immune responses, including recruiting inßammatory and immune cells from the circulation, retaining these cells within tissues, and, by regulating downstream signaling events, activating them to become effector cells and causing their subsequent proliferation. Selectins, integrins, and immunoglobulin (Ig) superfamily proteins mediate these activities through well-characterized cell-cell and cell-matrix interactions. Because psoriasis pathophysiology is thought to be largely a T-cell-mediated disease, several drugs in development attempt to interfere with CAM and accessory molecule interactions involved in T-cell activation. Targets include LFA-3, LFA-1 (CD11a), ICAM-1, and ICAM-3. Alefacept. Alefacept (Biogen Idec’s Amevive, formerly known as LFA3TIP) was introduced in the United States in 2003, but its launch in Europe will be delayed for some time because of a European Medicines Agency (EMEA) request for additional study data. Amevive is currently available only as a branded drug, and it is expected to retain market exclusivity for the next decade. Alefacept is a soluble fusion protein composed of the Þrst extracellular receptor-binding domain of the LFA-3 protein linked to the hinge domain of human IgG1. LFA-3 is present on a wide variety of cells, including antigenpresenting cells (APCs). Its ligand, the CD2 molecule, is expressed by T cells, thymocytes, and natural killer (NK) cells. The LFA-3/CD2 interaction facilitates adhesion of T-helper cells to APCs, activating a signal transduction cascade important for T-cell activation and proliferation. It was initially theorized that alefacept would block T-cell/APC interactions by binding to the CD2 molecule on T cells, thereby inhibiting T-cell activation. However, studies have revealed that the in vivo effects of alefacept appear to result from a bridge formed between CD2+ T cells and accessory cells bearing Fc receptors (such as macrophages and NK cells). The LFA-3 domain of alefacept binds to CD2 on T cells, whereas the CH2 domain (an Fc fragment) binds to Fc receptors on accessory cells. The formation of this intercellular bridge is believed to induce lysis of peripheral CD2+ T cells by directing NK-cell-mediated

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cytolysis (Majeau GR, 1994). Clinical evidence indicates that memory-effector T cells, believed to play a critical role in the development of psoriasis, decline in number in patients who are responsive to therapy with alefacept (Lowe N, 2002[a]). A Phase III trial of alefacept compared the efÞcacy of a single 12-week course of alefacept or two consecutive 12-week courses of alefacept with placebo. A total of 553 patients with moderate to severe psoriasis were randomized to receive alefacept (7.5 mg once weekly by 30-second IV bolus) for both courses of therapy (course 1), alefacept for the Þrst course and placebo for the second course (course 2), or placebo for the Þrst course and alefacept for the second course (course 3) (Krueger GG, 2002[b]; Krueger GG, 2003). Among patients who received course 1, 40% achieved 75% or greater improvement in PASI score (an improvement from 14% at week 14, compared with 4% among placebo-treated patients), and 71% achieved 50% or greater improvement in PASI score at any point during the study (Krueger GG, 2002[b]; Lebwohl M, 2002[a]). A “clear” or “almost clear” PGA status occurred more frequently among patients who received course 1 (32%) than course 2 or 3 (23% combined). These results suggest that two consecutive courses of alefacept prior to a disease ßare-up may provide greater beneÞt than re-treatment following a disease ßare-up (compare response rates of this trial with the open-label trial just described [Lowe N, 2002(a)]). Although the effect of continuous courses of alefacept is unknown, these data suggest that this agent may have a role as a maintenance therapy. Alefacept administered by intramuscular (IM) injection is also approved for the treatment of psoriasis. A multicenter and multinational dose-ranging Phase III trial evaluated the effect of IM administration of alefacept in 507 patients with chronic plaque psoriasis (Lebwohl M, 2003[a]). Patients received onceweekly IM injections of placebo or alefacept (10 mg or 15 mg) for 12 weeks; researchers Þrst measured efÞcacy at 14 weeks. Consistent with the Phase III trial of IV alefacept, maximum improvements in PASI score following one course of therapy were observed at week 18. Patients who received a 15 mg dose responded the most: 21% and 42% of these patients achieved 75% and 50% improvements in PASI scores, respectively, compared with 5% and 18%, respectively, among the placebo-treated patients. In the 15 mg group, 74% of patients who experienced a 75% or greater decline in PASI score maintained at least a 50% reduction in PASI during the 12-week follow up period after treatment was discontinued. Of those patients who experienced a 50–74% decline in PASI score, 79% maintained at least a 25% decline over the same follow-up period. Because the IM formulation permits self-administration by patients, it is likely to be favored over the IV formulation, which requires an ofÞce visit. Alefacept has been remarkably well tolerated, and none of the trials showed any evidence of increased cytokine release, capillary leak syndrome, disease rebounding, or rapid ßaring-up of disease following termination of therapy. Adverse events were mild and generally occurred at a similar rate in both the drug-treated and placebo-treated patients, as did patient withdrawal rates. In addition, alefacept was not found to elicit a signiÞcant antidrug antibody response

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in either the IV- or IM-dosing Phase III trials. A total of 22 patients developed anti-alefacept antibodies: 8 of the 553 patients in the IV dosing study and 14 of the 339 in the IM dosing study (Krueger GG, 2002[c]). These antibodies were transient in 14 of 22 cases; titers were generally low and did not increase with increasing exposure to alefacept. Additional studies are required to determine if the difference between the IV- and IM-dosing trial is signiÞcant. Limitations of alefacept include a likely high price, and, as with all psoriasis therapies, the eventual recurrence of psoriatic lesions at some point after discontinuation of treatment. Efalizumab. Efalizumab (Xoma/Genentech’s Raptiva, previously known as Xanelim) launched in the United States in 2003 and received marketing approval in Europe in 2004. It is currently marketed as a branded drug and is expected to retain market exclusivity for the next ten years. Efalizumab is a humanized monoclonal antibody (MAb) that targets CD11a (LFA-1). CD11a (the β chain) associates with CD18 (the α chain) to form the heterodimeric integrin LFA-1, which is expressed on the surfaces of T-helper cells and can interact with ICAM-1, -2, and -3. LFA-1/ICAM interactions are critical for T-cell proliferation and migration, and their disruption is hypothesized to inhibit T-cell function, providing clinical beneÞt in psoriasis. Results from an in vitro study suggest that efalizumab is effective in inhibiting transendothelial migration of T cells and suggest that it may decrease the accumulation of T cells in the skin (Lowe NJ, 2002[b]). A randomized, double-blind, placebo-controlled study of 556 patients with moderate to severe psoriasis explored efÞcacy and improvements in quality of life for patients treated with efalizumab (Gordon KB, 2003). Study participants received a 12-week course of subcutaneous efalizamab or placebo equivalent. The primary study end point was a 75% improvement as measured by the PASI index; secondary end points were improvement on the overall Dermatology Quality of Life Index (DLQI), Itching Visual Analogue Scale (VAS), and Psoriasis Symptom Assessment (PSA) at 12 weeks compared to baseline. An improvement of 75% or better on the PASI index was seen in 27% of the efalizumab-treated group versus 4% in the placebo group. Efalizumab-treated patients also showed greater mean percentages of improvement than placebo-treated patients on all secondary end points. Efalizumab was well tolerated, with adverse events including headache, chills, fever, myalgia, and pain. A Phase III, randomized, placebo-controlled, double-blind study of 597 patients in the United States explored increased doses of efalizumab and longer courses of treatment (Lebwohl M, 2003[b]). Patients received once-weekly subcutaneous (SC) injections of placebo or efalizumab (0.7 mg/kg for the Þrst week followed by 1 mg/kg or 2 mg/kg of efalizumab for the remaining 11 weeks). The primary end point was a 75% or greater improvement in PASI score, which was measured at various time points in a 12- or 24-week course of treatment and during a 12-week period after discontinuing treatment. SigniÞcant improvements in PASI were observed in both treatment groups relative to placebo after

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two to four doses (Lebwohl M, 2003[b]). After 12 weeks of therapy, 28% of patients who received 2 mg/kg of efalizumab and 22% of patients who received 1 mg/kg of efalizumab achieved 75% or greater improvements in PASI, compared with 5% of patients who received placebo. A subset of 434 subjects with varied responses to initial treatment with efalizumab underwent re-randomization and were followed for another 12-week course of treatment. Patients who initially achieved 75% or greater improvements in PASI maintained high rates of PASI improvement during the second 12-week period. Similar results were observed in the CLEAR (Clinical Experience Acquired with Raptiva) trial, an international, Phase III, randomized, double-blind, placebocontrolled study involving 793 patients (Sterry W, 2004). Patients with moderate to severe chronic plaque psoriasis received subcutaneous injections of 1mg/kg efalizumab or placebo once weekly for 12 weeks. The primary end point of the study was the percentage of patients achieving a PASI 75 improvement. At week 12 of the study, 31% of patients treated with efalizumab and 4% of patients treated with placebo had achieved a PASI 75 improvement. The mean percentage of PASI improvement in the efalizumab group was 48%, compared with an average 9% improvement in the placebo group. Adverse events included headache and inßuenza-like illness. No severe adverse events were reported. The drug generally has been well tolerated, and clinical trials have revealed no signiÞcant safety concerns. The most common side effects were mild headache and low-grade fever. Acute adverse events (AEs), occurring within two days of efalizumab injection, happened more frequently than in the placebo-only group with the Þrst two doses, and serious AEs were rare (2.4% among efalizumabtreated patients and 1% among placebo-treated patients). However, a recent hematologic analysis revealed that efalizumab-treated patients developed elevated lymphocyte counts when compared with placebo-treated patients; additional studies are required to determine the signiÞcance of this observation (Koo JY, 2002; Gordon KB, 2003). In an FDA brieÞng document on Raptiva released on September 9, 2003, concern was aired that some patients treated with efalizumab experience serious disease relapses, or “rebound,” upon withdrawal of the drug. Rebound is deÞned as reaching 125% of baseline PASI score within 12 weeks of discontinuation. In the clinical trials submitted to the FDA for review, 13.8% of patients treated with efalizumab experienced rebound ßares, compared with 11.1% of placebo. Some of these ßares may reßect a short washout period during clinical trials. TNF-α Inhibitors Overview. The central role of inßammation and TH 1 cytokines in the development of psoriasis is underscored by the emergence of agents targeting tumor necrosis factor alpha (TNF-α). Amgen/Wyeth’s etanercept (Enbrel) is the only TNF-α inhibitor currently approved for the treatment of psoriasis, though other agents targeting the same mechanism are currently in development (see “Emerging Therapies”).

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Mechanism of Action. TNF-α is a primary mediator of inßammation. In normal immune responses, TNF-α is produced in response to microbial infection and tissue injury. Increased expression of TNF-α provokes the recruitment of leukocytes from the circulation into tissue and increased production of chemokines and other proinßammatory cytokines. In patients with psoriasis, higher levels of TNF-α are observed in psoriatic lesions than in clinically normal skin. Levels of TNF-α produced by peripheral blood mononuclear cells in patients with active psoriasis correlate with disease severity, and elevated levels of TNF-α in psoriatic skin blister ßuids are signiÞcantly correlated with higher PASI scores. TNF-α inhibitors bind to both soluble and membrane-bound forms of TNF-α and inhibit their reaction with cell surface TNF-α receptors, preventing TNF-α from exerting effects on immune cells and keratinocytes. Etanercept. Amgen/Wyeth’s etanercept (Enbrel) is marketed for the treatment of RA and PsA in the United States and Europe and is preregistered for RA in Japan. Etanercept is marketed outside North America by Wyeth. A portion of global sales of etanercept must be paid as royalties to Genentech and Serono International because of their intellectual property rights to TNF-related technologies. Etanercept received FDA approval for the treatment of moderate to severe chronic plaque psoriasis in April 2004, and in August 2004 it received a recommendation for approval in Europe for the treatment of moderate to severe psoriasis in patients who have failed to respond to other therapies. Etanercept is currently marketed as a branded drug and is expected to retain market exclusivity during the next ten years. Etanercept is a soluble TNF-α receptor fusion protein consisting of part of the soluble p75 TNF-α receptor fused with the Fc fragment of human IgG1 . The TNF-α receptor component of etanercept binds both soluble and membranebound TNF-α with higher afÞnity than does the natural TNF-α receptor. A randomized, double-blind Phase III study of 672 patients examined the efÞcacy and safety of etanercept over a 24-week study period (Leonardi CL, 2003). Patients received placebo, low-dose etanercept (25 mg once weekly), mediumdose etanercept (25 mg twice weekly), or high-dose etanercept (50 mg twice weekly). After 12 weeks, patients in the placebo group began twice-weekly treatment with 25 mg of etanercept. The primary endpoint in the study was the percentage of patients achieving PASI 75. The mean improvement in PASI score and patient quality of life as assessed by the Dermatology Life Quality Index (DLQI) were also assessed. In the three treatment groups, 14% of patients treated with low-dose etanercept, 34% of patients treated with mediumdose, and 49% of patients treated with high-dose met the primary endpoint of a 75% or greater improvement in PASI scores, compared with 4% of patients in the placebo group. The mean improvement in PASI scores were 40.9 ± 2.4, 52.6 ± 2.4, and 64.2 ± 2.4, for the low-dose, medium-dose, and high-dose treatment groups, respectively, compared to an average improvement of 14.0 ± 2.6 with placebo. Etanercept was well tolerated in this study, with the most common adverse events being injection site reaction, headache, and upper-respiratory

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infection. Activation of latent tuberculosis infection is a concern with the TNF-α inhibitors inßiximab and adalimumab, but no cases of tuberculosis or opportunistic infections were reported during the study, and all laboratory abnormalities were of mild to moderate intensity. Because etanercept was approved for the treatment of psoriatic arthritis (PsA) in the United States and Europe in 2002, patients with both PsA and psoriasis were treated with etanercept prior to FDA approval for psoriasis. Also, many patients received off-label prescriptions for psoriasis even when they had no joint involvement, particularly in the United States. Therefore, etanercept was rapidly adopted by many physicians prior to regulatory approval for psoriasis. Etanercept has been prescribed for multiple indications, including rheumatoid arthritis, ankylosing spondylitis, and psoriatic arthritis, for Þve years, and longterm safety data indicate that it is well tolerated by patients who require ongoing treatment for chronic conditions. Patients treated with etanercept do not develop neutralizing antibodies to the molecule, and the therapeutic effect is therefore not expected to wane with chronic administration. As a biological agent, etanercept is among the most expensive agents available for the treatment of psoriasis. While Þve-year data have provided some reassurance regarding long-term, chronic administration of etanercept, many physicians remain concerned about the potential for malignancies arising in patients whose immune systems have been suppressed for extended periods. Infections will also remain a concern for any agent that causes long-term immunosuppression. Phototherapy and Photosensitizers Overview. Ultraviolet (UV) light phototherapy is used to treat localized lesions that do not respond to topical treatment and to treat widespread moderate to severe psoriasis. UV light is often combined with topical or systemic agents to reduce the dose of phototherapy. UV light exerts anti-inßammatory and immunosuppressive actions and induces apoptosis in proliferating cells. Two types of UV light are used: UVA and UVB. UVB is approximately 1,000 times more powerful than UVA, does not require the use of adjunct compounds, and is supplanting UVA as the Þrst choice of phototherapy. While UVB does not require the use of a photosensitizer, it is often used in combination with systemic and topical agents, including methotrexate, acitretin, calcipotriene, and coal tar. Mechanisms of Action. UVB and UVA phototherapies have similar mechanisms of action. However, because UVB radiation is more superÞcially absorbed, its effects are restricted to superÞcial keratinocytes and Langerhans’ cells. UVA radiation, which penetrates more deeply, affects superÞcial keratinocytes, Langerhans’ cells, dermal Þbroblasts, and dendritic and endothelial cells. Both phototherapies induce the expression of naturally anti-inßammatory mediators (e.g., IL-10 and the neuropeptide alpha-melanocyte-stimulating hormone [α-MSH]) and inhibit the expression of the TH 1 cytokine, IFN-γ . Both UVB and UVA radiation decrease the expression of receptors for the inßammatory cytokines IL-1 and TNF-α and of the intercellular adhesion molecule (ICAM)-1 on epidermal

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keratinocytes—the target for lymphocyte functional antigen (LFA)-expressing T cells in psoriasis. Recent data suggest that narrow-band UVB therapy may also induce T-cell apoptosis. UVB Radiation. UVB radiation may be used as monotherapy or in combination with anthralin or coal tar (the Goekerman regimen), both anthralin and coal tar (the Ingram regimen), retinoids, or other agents. Most research indicates that, unlike UVA, UVB radiation does not increase the risk of skin cancer, including melanoma. When used alone, both broadband (290–320 nanometer [nm]) and narrow-band (311–312 nm) UVB phototherapies have been shown to be effective for psoriasis. However, narrow-band UVB phototherapy has been shown to be superior to broadband for clearing psoriatic plaques, and it causes fewer side effects. Patients usually require 20–40 UVB radiation treatments at an average of three per week to obtain a full response (usually for three or more weeks). The initial treatment is followed by maintenance therapy twice weekly for at least one month and by once-weekly therapy for four months to prevent recurrence. The cumulative dose of UVB can be reduced by combining UVB therapy with systemic agents. The combination of UVB with methotrexate, for example, appears to be synergistic (Paul BS, 1982). One of the most clinically effective applications of UVB in psoriasis involves a combination of UVB and an oral retinoid (reUVB). In a randomized, doubleblind comparison of UVB therapy alone and UVB therapy with acitretin, 34 patients were assigned to two treatment groups (Lowe NJ, 1991). One group received UVB and placebo; the other received UVB and 50 mg per day acitretin. A third group of 16 severity-matched patients was treated in parallel with 50 mg per day of acitretin. Each group was treated for 12 weeks and evaluated every 4 weeks. The psoriasis grade scores used in the study decreased most significantly in the combination-treated group, an average of a 75% decline, versus a 35% decline in the UVB-only group and a 42% decline in the acitretin-only group. Although UVB phototherapy is believed to be largely free of cancer risk (except for applying it to the male genitalia), one case of melanoma in human tissue caused by UVB radiation has been reported. However, many physicians believe that UVB light may in fact be protective because the low doses used thicken but do not burn the skin. The primary limitation of phototherapy treatment is the requirement for multiple weekly visits to a treatment center, a requirement many patients are not able to reconcile with work schedules or family life. UVA Radiation with a Photosensitizer (PUVA). UVA treatment does require the concomitant use of a photosensitizing agent. UV lamps used for UVA phototherapy typically emit light at either 352 nm or 370 nm. The most common form of UVA phototherapy includes pretreatment with the photosensitizing agent psoralen and is known by the acronym PUVA. The most widely used psoralen formulation is 8-methoxypsoralen (8-MOP/methoxsalen/xanthotoxin; ICN Pharmaceuticals’ Oxsoralen), which is a naturally occurring plant product. Newer

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psoralen formulations with fewer side effects include 4, 5, 8-trimethoxypsoralen (ICN Pharmaceuticals’ Trisoralen) and 5-MOP (bergapten). Several mechanisms of action may underlie the effectiveness of PUVA therapy in psoriasis. Psoralens intercalate between DNA base pairs and, upon activation by photons in the UV spectrum, react with the pyrimidine bases to crosslink the DNA strands and inhibit DNA replication. This mechanism is thought to explain at least some of PUVA’s effectiveness in reducing the epidermal hyperproliferation of psoriasis. Psoralens also affect RNA and proteins—either directly or through free-radical generation—and this activity may in part explain their therapeutic effects in psoriasis and other dermatologic disorders. PUVA therapy decreases the concentration of several cell types in the skin, including CD4+ and CD8+ T cells as well as cutaneous antigen-presenting Langerhans’ cells, and decreases the production and release of IL-1β, IL-6, IL-8, and TNF-α. Psoralen formulations are administered before treatment with UVA light therapy as an oral formulation (two hours prior to light treatment), topical cream formulation, or placed in a bath preparation. When large areas of skin are treated, systemic levels of topical psoralens (as a 0.15% emulsion or solution) approach those present after oral administration. Psoralens applied by bath are readily absorbed into the skin yet result in low systemic concentrations. ICN Pharmaceuticals’ Trisoralen is frequently used in Scandinavia for bath PUVA. Delayed phototoxic erythema (skin reddening) occurs to varying degrees after PUVA therapy. Because a strong dose-response relationship exists between erythema and the dosage of psoralen and UVA energy, it is a major criterion for adjusting the dosage for future treatments. Treatments require eight hours and are administered no more than two to three times per week. Patients usually require a total of 30 PUVA treatments. Sunburn, blistering, and itching are the obvious potential side effects of PUVA, but they can be avoided with careful administration. Oral 8-MOP causes malaise and nausea in 20–30% of treated patients and vomiting in 10%. Overdoses of PUVA can cause swelling, intense pruritus, and a stinging sensation. PUVA therapy can also damage skin and accelerate aging of the skin. Psoralens can induce mutations in DNA, and PUVA signiÞcantly increases the risk for squamouscell carcinoma and slightly increases the risk for basal-cell skin malignancies (both of which are nearly always curable). A higher risk is associated with receiving more than 200 PUVA treatments, a history of skin cancer, radiation therapy or immunosuppressant drug therapy, and light skin tone. The risk of melanoma is also elevated in patients treated with PUVA, and it is cited as the main reason why physicians are decreasing use of PUVA and increasing use of UVB. Oral retinoids are sometimes used in conjunction with PUVA phototherapy to signiÞcantly lower the doses of radiation and of the oral retinoid. The mechanism of synergy between retinoids and UV radiation is not understood, but relatively low does of acitretin (10–25 mg daily, compared to 50 mg when given as monotherapy) accelerate the response to UV monotherapy, reducing

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the number and duration of sessions need to achieve clearance. A randomized, double-blind study of 60 patients compared PUVA plus placebo with PUVA plus acitretin (Tanew A, 1991). Marked or complete clearing occurred in 80% of the patients treated with PUVA plus placebo and in 96% of the patients treated with PUVA plus acitretin. In this study and a similar comparison, the addition of retinoids reduced the dose of PUVA needed for clearance by more than 40% (Tanew A, 1991; Saurat JH, 1988). Other agents that are combined with UVA phototherapy include glucocorticoids, anthralin, calcipotriene, and coal tar. PUVA cannot, however, be combined with cyclosporine due to increased risk of squamous cell cancers. The combination of methotrexate and PUVA may also increase the risk of squamous cell carcinomas, but it is an effective combination therapy in patients whose psoriasis is refractory to other treatments. Coal Tar. Coal tar is obtained by heating crude bituminous coal in the absence of oxygen. Although coal tar preparations have been used for the treatment of psoriasis for hundreds of years, they contain many organic products and it is not known which particular compound(s) are active or how they work in treating psoriasis. Coal tar is anti-inßammatory, decreases epidermal cell mitosis/development of scales, and reduces the production of sebum by sebaceous glands. Various preparations are available in different vehicles. OTC and prescription strengths are used clinically as monotherapy or in combination regimens. They are particularly useful in treating mild to moderate psoriasis when itching is severe and in guttate psoriasis. Coal tar is a photosensitizer and is sometimes used in conjunction with UV light monotherapy (without psoralen). It is also used with other topical therapies (usually corticosteroids). Coal tar is not recommended for pustular or erythrodermic pustular psoriasis because it may cause irritation, inducing Koebner’s phenomenon. Potential side effects of this therapy include folliculitis, contact dermatitis, and phototoxicity. Patient acceptance is a major problem because coal tar preparations are often greasy and messy, have an unpleasant odor, and stain clothing. EMERGING THERAPIES Psoriasis has attracted signiÞcant development over the past several years as biological agents, particularly the TNF-α inhibitors, emerge as therapeutic options for patients with severe disease. New immunosuppressants and fumaric acid esters also represent potential treatment options for patients with moderate or even mild disease. However, in recent years development has been discontinued for several drugs that reached Phase II or Phase III trials for this indication, including the therapeutic vaccine PVAC, oprelevkin, and ilodecakin, because of concerns about safety or failure to demonstrate adequate efÞcacy. Table 9 summarizes emerging therapies in late-stage development for psoriasis. The discussion of emerging therapies here is organized according to their mechanism of action.

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TABLE 9. Emerging Therapies in Development for Psoriasis Compound

Development Phase

Marketing Company

TNF-α inhibitors Infliximab United States Europe Japan

III III II

Johnson & Johnson/Centocor/Tanabe Johnson & Johnson/Centocor Tanabe

Adalimumab United States Europe Japan

III PC —

Abbott Laboratories Cambridge Antibody Technology —

Immunosuppressants Topical pimecrolimus United States Europe Japan

— IIa —

— Novartis —

Oral pimecrolimus United States Europe Japan

II II —

Novartis Novartis —

Topical tacrolimus United States Europe Japan

III II PC

Fujisawa Fujisawa Fujisawa

Retinoids Oral tazarotene United States Europe Japan

PR III —

Allergan Allergan —

Fumaric acid esters BG-12 United States Europe Japan

II IIIa —

Biogen-Idec Biogen-Idec —

PPAR-gamma agonists Rosiglitazone United States Europe Japan

III — —

GlaxoSmithKline — —

a European clinical trials conducted in Switzerland.

D = Discontinued; PC = Preclinical (including discovery); PR = Preregistered; R = Registered; S = Suspended.

TNF-α Inhibitors Overview. Tumor necrosis factor-alpha (TNF-α) inhibitors are the most prominent class of drugs in development for the treatment of psoriasis. Within the next Þve years, as many as four TNF-α inhibitors will be competing for a share of the psoriasis market. Two TNF-α inhibitors are in Phase III development

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for the treatment of psoriasis. Inßiximab (Johnson & Johnson/Centocor/Tanabe’s Remicade) and adalimumab (Abbott Laboratories/Cambridge Antibody Technology’s Humira) are approved for other indications, including rheumatoid arthritis and Crohn’s disease. Two TNF-α inhibitors that are not yet approved for any indication are also in development for psoriasis. CDP-870 (from Celltech) is in preclinical development for psoriasis, but will not be discussed in detail here because of the scarcity of clinical data speciÞc to psoriasis. The fourth, etanercept, is approved for the treatment of psoriasis in the United States and Europe. A Þfth TNF-α inhibitor, Serono’s onercept, was in Phase III trials for psoriasis, but development was discontinued when two patients treated with the drug developed sepsis. Mechanism of Action. TNF-α is a primary mediator of inßammation. In normal immune responses, TNF-α is produced in response to microbial infection and tissue injury. Increased expression of TNF-α provokes the recruitment of leukocytes from the circulation into tissue and increased production of chemokines and other proinßammatory cytokines. In patients with psoriasis, higher levels of TNFα are observed in psoriatic lesions than in clinically normal skin. Levels of TNF-α produced by peripheral blood mononuclear cells in patients with active psoriasis correlate with disease severity, and levels of TNF-α in psoriatic skin blister ßuids are signiÞcantly correlated with Psoriasis Area and Severity Index (PASI) scores. TNF-α inhibitors bind to both soluble and membrane-bound forms of TNF-α and inhibit their reaction with cell surface TNF-α receptors, preventing TNF-α from exerting effects on immune cells and keratinocytes. Infliximab. Inßiximab (Centocor/Johnson & Johnson’s Remicade) is a mousehuman chimeric monoclonal antibody (MAb) targeting TNF-α. It has been available in the United States since August 1998 for the treatment of Crohn’s disease and since November 1999 for rheumatoid arthritis (RA). In 2000, ScheringPlough launched inßiximab in Europe for Crohn’s disease and RA. In Japan, where it has been licensed to Tanabe Seiyaku, it is marketed for Crohn’s disease and RA. Inßiximab is currently in Phase III trials for psoriasis in the United States and Europe and in Phase II in Japan. Inßiximab consists of 75% human protein, comprising effector functions (elimination of TNF-α-positive cells and inhibition of upregulation of adhesion molecules), and 25% mouse protein, which contains the TNF-α binding sites. Inßiximab binds both soluble and membrane-bound TNF-α. Human antichimeric antibodies—reportedly as high as 30% (twice the rate of antibody production induced by etanercept)—have been observed, but with no clinically signiÞcant effects. Inßiximab is administered by IV infusion (over a two-hour period, in a physician’s ofÞce) every six to eight weeks. Inßiximab is already used off-label for psoriasis and psoriatic arthritis (PsA). Several small studies of its effectiveness for treating PsA and psoriasis are ongoing. Indeed, retrospective analysis of 31 patients with PsA and skin lesions who were treated with inßiximab at zero, two, and six weeks and every eight weeks

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thereafter revealed that 30 out of 31 patients experienced signiÞcant improvements in both skin lesions and affected joints, and only 1 patient failed to respond to therapy (Fleishmann RM, 2002). A small, randomized, controlled trial involving 33 patients with moderate to severe plaque psoriasis suggests that inßiximab may be efÞcacious for psoriasis as a monotherapy. Patients received placebo or inßiximab (5 or 10 mg/kg) at weeks 0, 2, and 6 and were evaluated at week 10. Inßiximab-treated patients achieved a median of 90% improvement in PASI score, compared with 11% among the placebo-treated group. All responders in the 5 mg/kg group and 64% of the 10 mg/kg group were rated either excellent (75–99% clearing with marked improvement) or clear on the Physician’s Global Assessment (PGA). In addition, markers of keratinocyte activation and inßammation declined relative to week 0 (Gottlieb AB, 2002[c]). Inßiximab was well tolerated; three patients, one from each treatment group, withdrew from the trial and were considered to be nonresponders. In an open-label follow-up study of these patients, the responders (19/22 patients) remained relapse-free for as long as ten months; however, one of two placebo-treated “responders” also remained relapse-free for nine months (Gottlieb AB, 2002[d]). Data from a larger Phase II study were presented at the annual meeting of the American Academy of Dermatology in February 2004 and at the annual meeting of the Society of Investigative Dermatology in April 2004 (Gottlieb AB, 2003; Gottlieb AB, 2004). In this randomized, placebo-controlled trial, 249 patients with moderate to severe psoriasis were treated with placebo or with infusions of 3 mg/kg or 5 mg/kg of inßiximab at weeks 0, 2, and 6. Patients were evaluated at week 10, and the primary end point of the study was a 75% improvement in PASI score (PASI 75). At week 10, 72% of patients receiving the lower dose of inßiximab and 88% of patients receiving the higher dose achieved PASI 75, compared with 6% of patients receiving placebo. Patients were followed through week 26 of the study to evaluate adverse events. Infusion reactions, a commonly noted adverse event, were observed in 4.1–10.7% of patients at various points in the study, versus 0–2% for placebo. The most common adverse events included upper respiratory infections, headaches, pruritus, sinusitis, pain, and arthralgia (Gottlieb AB, 2003; Gottlieb AB, 2004). Four potentially related serious adverse events occurred through week 40 of the study. In the lower-dose group, one patient developed squamous cell carcinoma and one developed cholecystitis and cholelithiasis. In the higher-dose group, one patient developed diverticulitis and one developed sepsis and pyelonephritis. These adverse events raised concerns about infections and malignancies arising from inßiximab-induced immunosuppression. Blood was drawn from participants through week 46 of the study to assess the development of antibodies against inßiximab as well as antinuclear antibodies (Gottlieb AB, 2004). The overall incidence of antibodies was 23.3%, comparable to similar patient populations in other clinical trials of inßiximab. Of participants who received inßiximab, 24% were newly positive for antinuclear antibodies during the study, with 13.5% positive at the Þnal visit.

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Overall, the Phase II data presented to date indicate that inßiximab effectively clears plaques in most patients, but the data fail to lay to rest concerns about inßiximab’s safety. Waning efÞcacy over time also remains a concern due to the development of anti-inßiximab antibodies. The package insert for inßiximab includes a boxed warning about tuberculosis and other opportunistic infections. The warning also states that patients should be evaluated for latent tuberculosis infection prior to treatment with inßiximab; latent tuberculosis infections have become active in a small number of patients treated with inßiximab. In a presentation at the Society of Investigative Dermatology in April 2004, the speaker noted that patients develop neutralizing antibodies against the murine sequences of inßiximab unless they receive a concomitant course of methotrexate when treatment with inßiximab is initiated (Gottlieb AB, 2004). Patients can be weaned from methotrexate at a later date. Inßiximab is delivered by a four-hour intravenous infusion requiring ofÞce space and staff that are not commonly found in dermatology practices. It is not clear from clinical trial data presented to date that inßiximab offers signiÞcant advantages in efÞcacy over the TNF-α inhibitor adalimumab (Abbott/Cambridge Antibody Technology’s Humira). Furthermore, dosage creep and waning efÞcacy are signiÞcant concerns for long-term administration. Adalimumab. The fully human monoclonal anti-TNF-α antibody adalimumab (D2E7, Humira) was developed by Abbott (formerly Knoll) in collaboration with Cambridge Antibody Technology. As a human monoclonal antibody, adalimumab is unique among monoclonal antibodies in this class. Adalimumab is currently marketed in the United States and in Europe for RA. Adalimumab is in Phase III clinical trials for psoriasis in the United States and in the discovery phase for psoriasis in Europe. Development of adalimumab for this indication has not commenced in Japan. Adalimumab’s mode of action is unique in the class of TNF-α inhibitors. While the monoclonal antibody comprises a TNF-α-binding site like other members of its class, its effector region promotes the lysis of TNF-expressing cells in the presence of complement. Adalimumab has a longer half-life than etanercept and therefore can be administered less frequently. Because it is derived from human sequences, it is likely to be minimally immunogenic. Promising preliminary results of the Phase II U.S. trial were presented at the annual meeting of the American Academy of Dermatology in February 2004 and at the Society of Investigative Dermatology Meeting in April 2004 (Gordon KB, 2004; Gottlieb AB, 2004; Peterson L, 2004). A total of 142 patients with moderate to severe chronic plaque psoriasis and no history of treatment with TNF-α antagonists were enrolled for a 12-week, double-blind study comparing two dosage regimens of adalimumab with placebo. In one treatment group, patients received 80 mg adalimumab at week 1, followed by 40 mg every other week from week 1 onward. The second group received 80 mg at weeks 0 and 1, and 40 mg weekly beginning at week 2. Placebo was administered weekly

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to patients in the placebo group. The primary end point of the study was the percentage of patients achieving PASI 75 at week 12 of the study. In the weekly treatment group, 80% of patients achieved PASI 75, compared with 53% of patients treated every other week and 4% of patients receiving placebo. The response to adalimumab was rapid, with a signiÞcantly greater mean percentage change in PASI score becoming evident as early as one week after the initial dose. According to adalimumab’s product label, the most frequently observed side effects of adalimumab in placebo-controlled clinical trials were mild injection-site reaction, hypertension, headache, pruritus, and rashes. The incidence of serious infections was 0.04 per patient-year in adalimumab-treated patients compared with 0.02 per patient-year with placebo; serious infections included pneumonia, septic arthritis, and erysipelas. The product label for adalimumab acknowledges that malignancies have been observed in recipients of TNF inhibitors, including adalimumab. Among 2,468 patients treated with adalimumab for a median of 24 months during clinical studies for RA, 0.4% of patients developed lymphomas. This occurrence is higher than that of the general population, but the rate of lymphoma may be higher in patients with highly active RA. Adalimumab’s label also includes a boxed warning cautioning physicians and patients about the risk of tuberculosis infection. The warning recommends that patients should be tested—and possibly treated—for latent tuberculosis infection prior to receiving adalimumab. Immunosuppressants Overview. Immunosuppressants are among the most effective agents for treating psoriasis, but those that are currently available are linked to renal toxicity and other serious adverse effects. Several new immunosuppressants that have demonstrated efÞcacy in other indications, such as renal and liver transplantation and graft-versus-host disease, are in development for psoriasis. These include tacrolimus (Fujisawa’s Prograf) and ISA-247 (Isotechnika/Roche). Isotechnika has completed Phase II trials of ISA-247 in Canada, and the drug is in Phase II trials in the United States for transplant rejection. Safety data for this agent derived from clinical trials of renal transplantation patients appear promising, but it will not be discussed here because of the scarcity of available data speciÞc to the psoriasis indication. Novartis’s pimecrolimus is in clinical trials for the psoriasis indication in topical formulation, but no new development has been announced for the indication since 1999. Pimecrolimus also reached late-phase clinical trials in an oral formulation, but Novartis announced in November 2005 that clinical trials of the drug had been suspended. Mechanism of Action. Immunosuppressants such as cyclosporine, tacrolimus, and pimecrolimus bind to cytoplasmic receptors, which are isomerases known as immunophilins. The drug/immunophilin complex exerts its primary immunosuppressive activity by binding to and inhibiting calcineurin, a protein that

620

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plays an essential role in calcium-dependent signal transduction in T cells. This signal transduction pathway activates cytokine gene expression, among others. Decreased cytokine production concomitantly decreases T-cell (TH 1 and TH 2) activation, including IL-2 production, growth, migration, and cytotoxic T-cell degranulation. Reduced cytokine secretion also affects the functions of keratinocytes, antigen-presenting cells, and neutrophils. Calcineurin is involved in T-cell-derived IL-2, IL-4, GM-CSF, TNF-α, and IFN-γ production. Therapy with cyclosporine or tacrolimus reduces the synthesis of all of these cytokines. See “Pathophysiology” for more information on the role of cytokines in psoriasis. Formulation. Topical and oral immunosuppressants are in development for psoriasis. Existing immunosuppressants, most notably cyclosporine, have not successfully been developed as topical agents. Cyclosporine is a high molecular weight compound that exhibits poor skin penetration, and it is therefore unsuitable for treating psoriatic plaques. Pimecrolimus is currently marketed in topical form only. Tacrolimus is marketed in both topical and oral forms for indications other than psoriasis. Pimecrolimus. The topical formulation of Novartis’ pimecrolimus (Elidel) is currently indicated for atopic dermatitis in the United States and Europe and is also in Phase II trials in Europe for psoriasis. The European trials have progressed slowly; no new development has been reported since 1999. Like other ascomycin derivatives, pimecrolimus is an immunophilin ligand. Its cognate receptor is the cytosolic immunophilin macrophilin-12. The pimecrolimus/macrophilin-12 complex prevents the synthesis of inßammatory cytokines by inhibiting calcineurin, much like other ascomycin derivatives (Gupta AK, 2003). In a ten-patient, double-blind trial reported in 1998, both 0.3% and 1% topical pimecrolimus, used under occlusion (e.g., occlusive tape), were effective in treating psoriasis. After two weeks of therapy, patients treated with 0.3% and 1% pimecrolimus exhibited 60% and 80% reductions, respectively, in microplaque score, compared with 18% for patients treated with the vehicle alone (i.e., placebo). Drug-treated patients exhibited a greater than 90% reduction in microplaque score—results comparable to those obtained with 0.05% clobetasol propionate, a high-potency corticosteroid. A galenic formulation of pimecrolimus has also been developed, and preliminary results suggest that it is effective at a 1% concentration for psoriasis. These results suggest that topical pimecrolimus is as effective as topical corticosteroids (Mrowietz U, 1998). Tacrolimus. Another immunosuppressant, tacrolimus (Fujisawa’s Prograf) (Figure 7), is indicated for atopic dermatitis. The topical formulation is currently in Phase III clinical trials in the United States and in Phase II trials in Europe. The oral formulation, however, is not presently in active development for psoriasis. A pilot study of 70 psoriasis patients comparing topical tacrolimus ointment with calcipotriene and with placebo found no statistically signiÞcant difference between the efÞcacy of tacrolimus and placebo ointment (Zonneveld IM, 1998).

EMERGING THERAPIES

621

HO

H3CO

CH3 H3C HO

O

O N O O

O

H3C

HO H3C

H3C O

OCH3 OCH3 FIGURE 7. Structure of tacrolimus.

However, the authors of that study noted that the high molecular weight of tacrolimus might prevent penetration into psoriatic skin. This theory is bolstered by evidence that systemic tacrolimus is effective for treating psoriasis (European FK506 Multicentre Psoriasis Study Group, 1996). Therefore, additional studies were undertaken exploring the potential of tacrolimus for treating the face and intertriginous areas (e.g., the groin and other skin folds) and combining tacrolimus with salicylic acid to promote penetration of psoriatic skin. An open-label clinical trial of 21 patients with psoriasis examined the efÞcacy of tacrolimus for treatment of the face and intertriginous areas, as assessed by the investigator’s evaluation of signs and symptoms of disease (Freeman AK, 2003). Assessments of adverse events were also made throughout the study to evaluate the safety of tacrolimus ointment. Few adverse events were noted, and no signs of skin atrophy, telangiectasia (an abnormal dilatation of capillary vessels and arterioles), or striae (stripes or lines in the skin)—complications associated with topical corticosteroids—were noted during the study. Most patients showed signiÞcant improvement compared with baseline. The authors acknowledged the limitations of the study design and noted that more extensive randomized, controlled trials were needed to conÞrm the observations. A right-left comparison study of 30 subjects compared 6% salicylic acid gel plus vehicle with 6% salicylic gel plus 0.1% tacrolimus ointment (Carroll CM, 2004). The primary outcome measure was the difference in change from baseline to end of treatment in a sum score of erythema, scale, and thickness scores. Lesions treated with tacrolimus on one side of the body were compared with lesions treated with placebo on the other side of the body. Twenty-four subjects completed the eight-week study. Adverse events were limited and included a stinging sensation on the side of the body treated with tacrolimus reported by four subjects. Greater improvement was observed with tacrolimus plus salicylic acid in the target plaque at weeks 1, 2, and 8 of treatment, but not at week 4.

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While topical tacrolimus causes relatively few adverse effects, it is more expensive than many topical agents, particularly generic topical corticosteroids. Topical corticosteroids are currently the most prescribed agents for psoriasis of the face and intertriginous areas, the most likely sites of application for topical tacrolimus. Nevertheless, topical tacrolimus is likely to be viewed as safer for treating sensitive and/or visible areas than topical corticosteroids, particularly given that it does not appear to cause the same side effects as topical steroids, such as atrophy, striae, and telangiectasia. Tacrolimus offers an improved treatment option for patients with facial lesions, who currently rely on topical corticosteroids. Topical immunosuppressants appear to promote clearance with fewer side effects than corticosteroids and are likely to be used for this purpose. Relatively few patients develop facial plaques, however, so only a small number of patients are likely to beneÞt. Retinoids Overview. Oral retinoids have been a mainstay of treatment for psoriasis for the past two decades. New retinoids, however, face substantial hurdles to approval due to increased concerns about fetal exposure to these teratogenic agents. Retinoids with improved selectivity for particular receptor classes and with shorter retention times in the body are being developed for psoriasis. At least two oral retinoids, tazarotene (Allergan’s Tazoral) and bexarotene (Ligand’s Targretin), are in development for this indication. Because of the limited availability of clinical trial data speciÞc to psoriasis, bexarotene will not be discussed in detail here. Mechanism of Action. Retinoids bind to speciÞc receptors called retinoic acid receptors (RARs) in the cell nucleus, affecting the transcription of genes involved in keratinocyte differentiation, cell proliferation, and inßammation. Clinical evidence and in vitro studies demonstrate that retinoids downregulate transcription of proinßammatory cytokines, halt proliferation of cultured cells, and alter expression of indicators of keratinocyte differentiation. Oral Tazarotene. Allergan recently submitted a new drug application (NDA) to the U.S. Food and Drug Administration (FDA) for oral tazarotene (Tazoral). Phase III trials of oral tazarotene are also under way in Europe. Tazarotene is a third-generation retinoid that selectively binds to retinoic acid receptors (RARs)-beta and -gamma for the treatment of moderate to severe psoriasis. Binding to its cognate RARs downregulates transcription of proinßammatory cytokines and suppresses keratinocyte proliferation. Because it activates neither RAR-α nor retinoid X receptors (RXRs), oral tazarotene is not associated with some of the side effects that occur with other oral retinoids, such as hypertriglyceridemia, hypothyroidism, and hepatoxicity (Duvic M, 2004). The efÞcacy and safety of oral tazarotene were examined in two multicenter, double-blind, randomized, placebo-controlled Phase III studies (Koo JY, 2004). In these studies, a total of 706 patients with moderate to severe psoriasis were enrolled and evaluated (Kang S, 2004). Greater than 50% global improvement and changes in Overall Lesional Assessment (OLA) scores were used as end points to

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measure the effects of tazarotene. After a three-month period, 54% of the patients treated with oral tazarotene achieved at least 50% global improvement (compared with 15% of the patients treated with placebo). Of those who experienced at least 50% global improvement, 80% maintained their condition for at least one additional three-month period without treatment. Among tazarotene-treated patients, 28% experienced a two-grade improvement in OLA (on a six-grade scale), compared with 6% of the patients treated with placebo; 81% of these tazarotene-treated patients were able to maintain an improved OLA score for at least three months post-treatment. Results also showed a measurable reduction in lesional elevation, scaling, and erythema on all parts of the body but especially on difÞcult-to-treat lesions on the knees and elbows (Koo JY, 2004; Kang S, 2004). The safety of oral tazarotene has been examined in Þve studies involving a total of 1,100 patients. The combined results of the Þve studies were presented at the annual meeting of the American Academy of Dermatology in February 2004 (Walker PS, 2004). Adverse events that occurred at higher frequency among tazarotene-treated patients than among placebo-treated patients were dry lips (66% versus 17%), dry skin (24% versus 15%), headache (19% versus 12%), and several other adverse events that occurred at lower frequency. None was serious in nature. Other adverse effects typically observed among patients treated with oral retinoids occurred at levels comparable to placebo. These included elevated serum triglyceride and cholesterol levels, abnormal liver function tests, eye dryness, shedding of skin (desquamation), and alopecia. Oral tazarotene has the potential to replace the current oral retinoid therapy, acitretin (Roche’s Soriatane). Because pivotal clinical trials for tazarotene and acitretin used different end points, however, it is difÞcult to compare their efÞcacy. Tazarotene has a more selective mechanism of action and causes less elevated lipid levels and liver toxicity than acitretin. Furthermore, investigation of the pharmacokinetic proÞle of tazarotene demonstrated that the drug has a half-life of 7 to 12 hours and is completely eliminated within 11 days. In contrast, acitretin elimination can take up to four weeks, and consumption of even small amounts of alcohol can extend retention considerably (Yu D, 2004). Therefore, women of reproductive age must avoid pregnancy for three years after discontinuing therapy with acitretin. No such long-term restriction is necessary for tazarotene, but fetal exposure must nevertheless be avoided while taking the drug and for one month after discontinuing therapy. The launch of oral tazarotene suffered a substantial setback in July 2004 when the FDA panel evaluating the drug voted against approving oral tazarotene for the treatment of moderate to severe psoriasis. In a joint meeting of the Dermatologic and Ophthalmic Drugs Advisory Committee and the Drug Safety and Risk Management Advisory Committee on July 12, 2004, the joint panel focused heavily on the program proposed by Allergan to prevent fetal exposure to tazarotene. However, the panel also mentioned concerns about efÞcacy, particularly among patients with very severe disease, and several members felt that more data were needed (particularly longer studies) to assess the drug’s efÞcacy. Furthermore,

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several members of the panel expressed limited conÞdence regarding the instruments used to assess efÞcacy. In a conference call broadcast on July 12, 2004, Allergan indicated that it will continue to pursue approval for this indication and will work with the FDA to resolve the issues identiÞed by the committee. Fumaric Acid Esters Overview. Fumaric acid esters have been prescribed in Germany for several decades, but they have only recently attracted attention in other markets. However, fumaric acid esters are not currently approved for the treatment of psoriasis in markets outside of Germany. Agents currently available in Germany are mixtures of dimethylfumarate and monoethylfumarate salts. A single agent belonging to this class, Biogen-Idec/Fumapharm’s BG-12, is in Phase II clinical trials in the United States and Phase III clinical trials in Europe. Mechanism of Action. Fumaric acid esters, and dimethylfumarate in particular, appear to downregulate TH 1 cytokines and modulate cytokine expression toward a TH 2 proÞle (Loewe R, 2002). Dimethylfumarate and its hydrolysis product methylhydrogen fumarate both appear to inhibit keratinocyte proliferation and monocyte differentiation into dendritic cells. Dimethylfumarate also induces apoptosis in activated human T cells (Treumer F, 2003). BG-12. Biogen Idec acquired the rights to develop and market a secondgeneration fumaric acid derivative from Fumapharm AG in October 2003. Phase II trials have been completed for this indication and were presented at the 2004 annual meeting of the European Academy of Dermatology and Venereology (Langner A, 2004). A double-blind, placebo-controlled, dose-Þnding study of 144 patients measured improvements in PASI score, the Physician’s Clinical Global Impression, Patient’s Global Assessment, and quality of life (using the Skindex-29 instrument) over a 12-week period. Patients received placebo or 120 mg, 360 mg, or 720 mg of fumaric acid ester. At week 12, the median percentage reductions from baseline PASI score were 6% for the placebo group and 31%, 52%, and 71% for the 120 mg, 360 mg, and 720 mg doses, respectively. PASI 75 reductions were observed in 11%, 8%, 25%, and 42% of the placebo, 120 mg, 360 mg, and 720 mg groups, respectively. PASI 50 reductions were observed in 14%, 39%, 50%, and 64% of the placebo, 120 mg, 360 mg, and 720 mg groups. The response to BG-12 was rapid, with evidence of clearing appearing two weeks after beginning therapy. Gastrointestinal side effects, the most commonly noted adverse effect in patients receiving mixed fumaric acid esters, were observed in less than 5% of patients in each group. Adverse events included ßushing, minor elevations of liver enzymes, and respiratory infections (Langner A, 2004). PPAR Agonists Overview. Peroxisome proliferator-activated receptor (PPAR) agonists are wellestablished and widely marketed drugs for diabetes, but their potential value

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FIGURE 8. Structure of rosiglitazone.

for treating psoriasis has only recently been noted. The initial motivation for exploring the utility of PPAR agonists in the treatment of psoriasis was the serendipitous observation that patients affected with both diabetes and psoriasis noticed improvements in their skin condition upon initiating treatment with PPAR agonists for diabetes. While preclinical studies exist exploring the effect of various PPAR agonists on proliferating keratinocytes and on inßammation, only rosiglitazone (GlaxoSmithKline’s Avandia) is known to be in clinical development for psoriasis. Mechanism of Action. PPARs are members of the nuclear hormone receptor superfamily. The family comprises ligand-activated transcription factors that are related to retinoid, steroid, and thyroid hormone receptors. The PPAR subfamily of ligand-activated transcription factors includes three isotypes (PPAR-α, PPAR-β/γ , and PPAR-γ ), which are all expressed in keratinocytes. PPAR-β/γ is expressed at the highest levels of the three subtypes in human epidermis, though PPAR-α and PPAR-γ increase upon differentiation. In psoriatic epidermis, all three subtypes show altered patterns of expression compared with normal epidermis. Rosiglitazone. Rosiglitazone (GlaxoSmithKline’s Avandia) (Figure 8), a PPAR-γ agonist currently approved for the treatment of type II diabetes, is in Phase III clinical trials for psoriasis in the United States. Preclinical data indicate that rosiglitazone inhibits keratinocyte proliferation and motility in cell culture (Bhagavathula N, 2004). Rosiglitazone also reduces acute inßammation in animal models (Cuzzocrea S, 2004). While the theoretical basis for treating psoriasis with PPAR-γ agonists is well documented, no Phase II or Phase III clinical trials have been published in peer-reviewed journals or presented at scientiÞc conferences to date. Clinical data in the public domain are limited to the results of a small pilot study of eight patients followed for 30 days, in which no clinically signiÞcant improvement was seen in patients taking rosiglitazone compared with patients treated with placebo (Kuenzli S, 2003). In the diabetes market, rosiglitazone is generally regarded as safe, with notable adverse effects being weight gain and ßuid retention. REFERENCES Abel EA. Psoriasis. In: Dale DC, Federman DG, eds. ScientiÞc American Medicine. New York: ScientiÞc American Inc.; 1994.

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Amblard P. Psoriasis: diagnostic, evolution, prognostic, treatment. La R´evue du Praticien (Paris). 1993;43:229–231. Antoni C, et al. Successful treatment of psoriatic arthritis with inßiximab. Presented at the American College of Rheumatology 63rd Annual ScientiÞc Meeting; November 16, 1999; Boston. Abstract 1801. Antoni C, et al. Long-term open label treatment of psoriatic arthritis with inßiximab. 60th Annual Meeting of the American Academy of Dermatology; February 22–27, 2002; New Orleans. Abstract P538. Baron SE, et al. The use of complimentary medicine within a population of dermatological outpatients in Leeds, UK. 60th Annual Meeting of the American Academy of Dermatology; February 22–27, 2002; New Orleans. Abstract P271. Bell LM, et al. Incidence of psoriasis in Rochester, Minn., 1980–1983. Archives of Dermatology. 1991;127:1184–1187. Bhagavathula, N. Rosiglitazone inhibits proliferation, motility, and matrix metalloproteinase production in keratinocytes. Journal of Investigative Dermatology. 2004;122: 130–139. Bhushan M, et al. Levels of endothelial cell stimulating angiogenesis factor and vascular endothelial growth factor are elevated in psoriasis. British Journal of Dermatology. 1999;141:1054–1060. Camisa C. Psoriasis: a clinical update diagnosis and new therapies. Cleveland Clinical Journal of Medicine. 2000;67:105–119. Carroll CM, et al. Topical tacrolimus ointment in combination with 6% salicylic acid gel for the treatment of psoriasis: results of a randomized, double-blind, right to left clinical trial. 62nd Annual Meeting of the American Academy of Dermatology; February 6–11, 2004; Washington, D.C. Abstract P579. Christophers E. Psoriasis—epidemiology and clinical spectrum. Clinical and Experimental Dermatology. 2001;26:314–320. Cohen MR, et al. Baseline relationships between psoriasis and psoriatic arthritis: analysis of 221 patients with active psoriatic arthritis. Journal of Rheumatology. 1999;26: 1752–1755. Cuzzocrea S, et al. Rosiglitazone, a ligand of the peroxisome proliferators-activated receptor-gamma, reduces acute inßammation. European Journal of Pharmacology. 2004;483:79–93. de Jong EMGJ. The course of psoriasis. Clinics in Dermatology. 1997;15:687–692. Diaz BV, et al. Regulation of vascular endothelial growth factor expression in human keratinocytes by retinoids. Journal of Biological Chemistry. 2000;275:642–650. Dubertret L, et al. EfÞcacy and safety of calcipotriol (MC 903) ointment in psoriasis vulgaris. Journal of the American Academy of Dermatology. 1992;27:983–988. Dummer W, et al. Pharmacodynamic effects of subcutaneous (SC) administration of efalizumab (anti-CD11a). 60th Annual Meeting of the American Academy of Dermatology; February 22–27, 2002; New Orleans. Abstract P547. Duvic M, et al. Oral retinoids in the treatment of psoriasis. 62nd Annual Meeting of the American Academy of Dermatology; February 6–11, 2004; Washington, D.C. Abstract P599. Elder JT, et al. Epidemiology and the genetics of psoriasis. Journal of Investigative Dermatology. 1994;102:24S–27S.

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Gordon KB, et al. Efalizumab for patients with moderate to severe plaque psoriasis: a randomized controlled trial. Journal of the American Medical Association. 2003;290: 3073–3080. Gottlieb A, et al. Subcutaneous efalizumab (anti-CD11a) is effective in the treatment of moderate to severe plaque psoriasis pooled results of two Phase III clinical trials. 60th Annual Meeting of the American Academy of Dermatology; February 22–27, 2002; New Orleans. Abstract P576. [a] Gottlieb AB, et al. EfÞcacy of Enbrel in patients with psoriasis. 60th Annual Meeting of the American Academy of Dermatology; February 22–27, 2002; New Orleans. Abstract 159. [b] Gottlieb AB, et al. Inßiximab monotherapy normalizes keratinocyte differentiation and decreases inßammation in skin biopsies from patients with moderate to severe plaque psoriasis. 60th Annual Meeting of the American Academy of Dermatology; February 22–27, 2002; New Orleans. Abstract P539. [c] Gottlieb AB, et al. Inßiximab prevents relapse of moderate to severe psoriasis in responding patients. 60th Annual Meeting of the American Academy of Dermatology; February 22–27, 2002; New Orleans. Abstract P540. [d] Gottlieb AB, et al. The efÞcacy and safety of inßiximab induction treatment in subjects with severe plaque-type psoriasis. 9th International Psoriasis Symposium; June 17–22, 2003; Poster. Gottlieb AB, et al. Evaluating therapeutic strategies for psoriatic disease and the use of biologics. 65th Annual Meeting of the Society for Investigative Dermatology; April 28-May 3, 2004; Providence, RI. Seminar. Gottlieb SL, et al. Response of psoriasis to a lymphocyte-selective toxin (DAB389IL-2) suggests a primary immune, but not keratinocyte, pathogenic basis. Nature Medicine. 1995;1:442–447. Greaves MW, Weinstein GD. Treatment of psoriasis. New England Journal of Medicine. 1995;332:581–588. GrifÞths C, et al. Treatment with oral pimecrolimus signiÞcant improves psoriasis with a clear dose-response effect. Joint Meeting of the European Society for Dermatological Research, Japanese Society for Investigative Dermatology, and Society for Investigative Dermatology; April 30-May 4, 2003; Miami Beach, FL. Abstract 0391. Guedjonsson JE, et al. HLA-Cw6-positive and HLA-Cw6-negative patients with psoriasis vulgaris have distinct clinical features. Journal of Investigative Dermatology. 2002;118:362–365. Gulliver W, et al. The effect of intramuscular alefacept on the individual components of the psoriasis area and severity index. 63rd Annual Meeting of the Society for Investigative Dermatology; May 15–18, 2002; Los Angeles. Abstract P822. Gupta AK, et al. Pimecrolimus: a review. Journal of the European Academy of Dermatology and Venereology. 2003;17:493–503. Henseler T. The genetics of psoriasis. Journal of the American Academy of Dermatology. 1997;37(2 Pt. 3):S1–S11. Hewett D, et al. IdentiÞcation of a psoriasis susceptibility candidate gene by linkage disequilibrium mapping with a localized single nucleotide polymorphism map. Genomics. 2002;79:305–314. Heydendael et al. Methotrexate versus cyclosporine in moderate-to-severe chronic plaque psoriasis. New England Journal of Medicine. 2003;349:658–665.

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The International Psoriasis Genetics Consortium. The International Psoriasis Genetics Study: assessing linkage to 14 candidate susceptibility loci in a cohort of 942 affected sib pairs. American Journal of Human Genetics. 2003;73:430–437. Johnson MT, Roberts J. Prevalence, disability, and health care for psoriasis among persons 1–74 years: United States. Advance data from vital and health statistics; no. 47. Hyattsville, MD: National Center for Health Statistics; 1979. Kang S, et al. Oral tazarotene and difÞcult-to-treat psoriatic lesions. 62nd Annual Meeting of the American Academy of Dermatology; February 6–11, 2004; Washington, D.C. Abstract P597. Katz, HI, et al. Intermittent corticosteroid maintenance of psoriasis: a double-blind multicenter trial of augmented betamethasone dipropionate ointment in a pulse dose treatment regimen. Dermatologica. 1991;183:269–274. Kehren JC, et al. Genomic analysis of blood cells from psoriatic patients following treatment with oral pimecrolimus (SZD ASM 981). 60th Annual Meeting of the American Academy of Dermatology; February 22–27, 2002; New Orleans. Abstract P101. Koo J. Population-based epidemiologic study of psoriasis with emphasis on quality of life assessment. Dermatologic Clinics. 1996;14:485–496. Koo JY, et al. Efalizumab (anti-CD11a) is safe and well tolerated in the treatment of moderate to severe plaque psoriasis: pooled results of two Phase III clinical trials. 60th Annual Meeting of the American Academy of Dermatology; February 22–27, 2002; New Orleans. Abstract P548. Koo JY et al. Duration of clinical improvement with oral tazarotene in plaque psoriasis. 62nd Annual Meeting of the American Academy of Dermatology; February 6–11, 2004; Washington, D.C. Abstract P598. Kragballe K, et al. A new combination of calcipotriol and betamethasone dipropionate is effective used once daily in psoriasis vulgaris. 60th Annual Meeting of the American Academy of Dermatology; February 22–27, 2002; New Orleans. Abstract P536. Krueger GG, Duvic M. Epidemiology of psoriasis: clinical issues. Journal of Investigative Dermatology. 1994;102:14S–18S. Krueger GG, et al. Duration of response to alefacept therapy in patients with chronic plaque psoriasis. 60th Annual Meeting of the American Academy of Dermatology; February 22–27, 2002; New Orleans. Abstract P584. [a] Krueger GG, et al. EfÞcacy of two courses of intravenous alefacept in patients with chronic plaque psoriasis. 63rd Annual Meeting of the Society for Investigative Dermatology; May 15–18, 2002; Los Angeles. Abstract 819. [b] Krueger GG, et al. Alefacept (human LFA-3/IgG1) is well tolerated and non-immunogenic: results of two randomized, placebo-controlled Phase III trials. 60th Annual Meeting of the American Academy of Dermatology; February 22–27, 2002; New Orleans. Abstract P583. [c] Krueger GG, et al. Clinical response to alefacept: results of a phase 3 study of intravenous administration of alefacept in patients with chronic plaque psoriasis. Journal of the European Academy of Dermatology and Venereology. 2003;17 Suppl 2:17–24. Kuenzli S. et al. Effect of topical PPAR beta/delta and PPAR gamma antagonists on plaque psoriasis. A pilot study. Dermatology. 2003;206:252–256. Langner A. et al. Oral fumarate for the treatment of severe forms of psoriasis: results of a phase II clinical study. 2nd Spring Symposium of the European Academy of

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Dermatology and Venereology. April 29-May 1, 2004; Budapest, Hungary. Abstract PS296. Lebwohl M. et al. Tazarotene in combination with topical corticosteroids. Journal of the American Academy of Dermatology. 1998;39S:139–143. Lebwohl M, et al. Calcipotriene ointment and halobetasol ointment in the long-term treatment of psoriasis: effects on the duration of treatment. Journal of the American Academy of Dermatology. 1998;39:447–450. Lebwohl M, et al. Results of a multiple-course, randomized, Phase III study of alefacept (human LFA-3/IgG1) in patients with chronic plaque psoriasis. 60th Annual Meeting of the American Academy of Dermatology; February 22–27, 2002; New Orleans. Abstract P580. [a] Lebwohl M, et al. Etanercept improved psoriasis activity in patients with psoriatic arthritis: results of a Phase 3 multicenter clinical trial. 60th Annual Meeting of the American Academy of Dermatology; February 22–27, 2002; New Orleans. Abstract P561. [b] Lebwohl M, et al. An international, randomized, double-blind, placebo-controlled Phase III trial of intramuscular alefacept in patients with chronic plaque psoriasis. Archives of Dermatology. 2003;139:719–727. [a] Lebwohl M, et al. A novel targeted T-cell modulator, efalizumab, for plaque psoriasis. New England Journal of Medicine. 2003;349:2004–2013. [b] Lebwohl M, et al. Combination therapy to treat moderate to severe psoriasis. Journal of the American Academy of Dermatology. 2004:50;416–430. Leonardi C, et al. Pilot study of topical ISIS 2302, an antisense oligodeoxynucleotide, in patients with plaque tope psoriasis vulgaris. 60th Annual Meeting of the American Academy of Dermatology; February 22–27, 2002; New Orleans. Abstract P28. Leonardi CL, et al. Etanercept as monotherapy in patients with psoriasis. New England Journal of Medicine. 2003;349:2014–2022. Leonardi CL, et al. The safety of efalizumab in patients with moderate to severe plaque psoriasis: summary of clinical trial experience. 62nd Annual Meeting of the American Academy of Dermatology; February 6–11, 2004; Washington, D.C. Abstract P613. Loewe R, et al. Dimethylfumarate inhibits TNF-induced nuclear entry of NF-kappa B/p65 in human endothelial cells. Journal of Immunology. 2002;168:4781–4787. Lowe N, et al. Results of repeat courses of alefacept therapy for the treatment of chronic plaque psoriasis. 60th Annual Meeting of the American Academy of Dermatology; February 22–27, 2002; New Orleans. Abstract P578. [a] Lowe NJ, et al. Acitretin plus UVB therapy for psoriasis. Comparisons with placebo plus UVB and acitretin alone. Journal of the American Academy of Dermatology. 1991;24:591–594. Lowe NJ, et al. Efalizumab (anti-CD11a) inhibits trans-endothelial migration of T cells. 63rd Annual Meeting of the Society for Investigative Dermatology; May 15–18, 2002; Los Angeles. Abstract 710. [b] Majeau GR, et al. Mechanism of lymphocyte function-associated molecule 3-Ig fusion proteins inhibition of T cell responses. Structure/function analysis in vitro and in human CD2 transgenic mice. Journal of Immunology. 1994;152:2753–2767. Mease PJ, et al. Etanercept in the treatment of psoriatic arthritis and psoriasis: a randomized trial. Lancet. 2000;356:385–390.

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Rheumatoid Arthritis

ETIOLOGY AND PATHOPHYSIOLOGY Introduction Rheumatoid arthritis (RA) (International ClassiÞcation of Diseases, Ninth Revision [ICD-9], code 714, and Tenth Revision [ICD-10], code M06.9) is characterized by chronic joint inßammation that typically leads to debilitating tissue damage, further compounded by joint deformation. Although the precise etiology remains elusive, experts view RA as a disease of autoimmune origin, spurred by environmental triggers acting on a genetically predetermined host. However, few genetic, and no environmental, factors have yet been identiÞed deÞnitively. This section reviews the disease course and complications of RA as well as the potential role of select genetic and environmental risk factors in the initiation and evolution of RA pathophysiology. Pathophysiology Immune Response. RA is widely considered an autoimmune disease. The prominence of the T-cell inÞltrate in the synovium (the inner lining of synovial joints) suggests that these cells are key participants. A 1998 study reported that a subset of CD4+ (helper) T cells in RA patients, but not human leukocyte antigen (HLA)-DR-matched controls, displayed increased expression of the cell survival protein bcl-2, which blocks apoptosis (Schirmer M, 1998). The researchers speculated that this increase may favor the clonal expansion of autoreactive T cells in RA patients. Several additional studies have focused on the expression of the Fas (a member of the TNF receptor family) and its ligand in RA. Fas regulates apoptosis (cell death) in the immune system through interaction with Fas ligand. Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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In addition, synovial cells express elevated levels of bcl-2, likely induced by pro-inßammatory cytokines and contributing to pannus (an inßammatory exudate overlying the layer of synovial cells lining the inside of a joint) formation. Infusion of Fas ligand by gene transfer techniques or by injection of cross-linking Fas antibodies is being explored in animal models of arthritis. In a recently reported study in a murine model, upon treatment with anti-Fas in vitro, cell death of Þbroblasts was reduced and the expression of pro-inßammatory cytokines, tumor necrosis factor-alpha (TNF-α), and interleukin-6 (IL-6) increased, pointing to a role of Fas in joint destruction (Hoang TR, 2004). Disease Course. RA typically displays an insidious onset spanning weeks or months. Symptoms may begin with general fatigue, morning stiffness, and pain. They may progress to localized, symmetrical joint inßammation, most commonly in the hands, wrists, and feet. Systemic effects of RA may appear later in the disease (as discussed in the section “Extra-articular and Systemic Manifestations”). Importantly, no clinical markers currently exist that can deÞnitively diagnose RA in all patients. RA progression is classiÞed into four stages. Stage I marks the onset of the earliest clinical manifestation—synovial inßammation (synovitis), which is characterized by an increase in synovial ßuid volume containing abnormally elevated quantities of Þbrin, immune complexes, complement components, proteases, impaired protease inhibitors, and the pro-inßammatory cytokines interleukin-1beta (IL-1β), IL-6, and TNF-α. Inßammation of the synovial microvasculature and increased capillary permeability result in an inßux of B cells, T cells, macrophages, and neutrophils. In addition, cyclooxygenase and lipoxygenase gene expression increases, resulting in greater prostaglandin and leukotriene production. All these events result in the induction and maintenance of an inßammatory state in the joint. Stage II of RA is marked by synovial hypertrophy and cellular proliferation, resulting in invasion of the joint cavity. Erosion of the articular cartilage ensues because the avascular, cartilage-producing chondrocyte layer is denied access to the nutrient-rich synovial ßuid, resulting in chondrocyte death. The synovium becomes further inundated with activated Þbroblasts and immune cells, exacerbating the inßammatory state of the joint. Endothelial cells also may be observed in the synovium because of ongoing neovascularization. Stage III of RA is characterized by pannus formation. Consisting of numerous Þbroblasts, small blood vessels, and mononuclear cells, this vascular granulation tissue spreads to cover the articular cartilage. In response to this chronic inßammatory state, large amounts of tissue- and cartilage-degrading enzymes, including collagenases and stromelysins, are produced. Eventually, subchondral bone is exposed and demineralized by osteoclasts, which are activated by the high levels of IL-1β, TNF-α, and prostaglandin E2 . Signs of articular deformity may be observed at this stage. The inßammatory process begins to diminish by stage IV, when erosion of articular cartilage and bone is extremely advanced, and Þbrous or bony ankylosis severely limits the function of affected joints. Nodulosis (the formation of

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subcutaneous rheumatoid nodules) occurs in 20–30% of patients, particularly in those strongly seropositive for rheumatoid factor (RF; see the later section “Pathogenesis of Joint Damage”). These nodules, which are seldom symptomatic, may be caused by focal vasculitis and consist of a central necrotic core, a middle zone of palisading macrophages, and an outer ring of granulation tissue. RA patients can also be classiÞed into four categories according to the Steinbrocker Functional Class system, which enables a determination of the percentage of patients presenting by severity of disease (Beckman JC, 1992; Sany J, 1998): • • • •

Class I (early) consists of patients with no restriction of ability to perform normal activities (approximately 24% of prevalent RA cases). Class II (moderate) encompasses patients who have moderate physical restriction but are nevertheless able to perform normal activities (50%). Class III (severe) comprises patients with marked restriction; they are unable to perform most tasks related to occupation or self-care (25%). Class IV (terminal) consists of RA patients incapacitated by the disease and, in some cases, conÞned to bed or a wheelchair (1%).

Extra-articular and Systemic Manifestations. Research suggests that the term rheumatoid disease more properly deÞnes RA, particularly because of systemic involvement. Rheumatoid nodules may develop in the posterior portion of the brain, pleura, or meninges, with potentially serious complications. Cardiopulmonary manifestations include pleuritis and pericarditis with effusions resembling synovial ßuid. These events are difÞcult to diagnose, but asymptomatic pericarditis is observed in 40–50% of autopsied RA patients. Lung parenchymal involvement may present as either interstitial pneumonitis or pulmonary Þbrosis. Renal abnormalities such as proteinuria may occur in RA patients, often as a complication of drug therapy. Rheumatoid vasculitis can affect any organ system and is typically observed in patients who have aggressive disease and are strongly RF-positive. Approximately 15–20% of RA patients develop Sj¨ogren’s syndrome, which is characterized by kerato-conjunctivitis sicca (typical signs being hyperemia of the conjunctiva, lacrimal deÞciency, thickening of the corneal epithelium, itching and burning of the eye, and often reduced vision). Other ocular manifestations affecting less than 1% of patients include scleritis, episcleritis (an inßammatory condition of the connective tissue between the conjunctiva and sclera), and ocular nodules. Osteoporosis is a common and often underdiagnosed complication of RA. Felty’s syndrome may occur late in the disease course, involving neutropenia, thrombocytopenia, and splenomegaly. Pathogenesis of Joint Damage. Although the cause of RA remains elusive, the destructive features of the disease have been extensively documented and described. The following sections review each key element of disease pathogenesis. They include the inÞltration of lymphocytes and macrophages into synovial tissue, hyperplasia of synovial cells, and the enhanced expression of multiple inßammatory mediators, such as cytokines and degradative enzymes.

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FIGURE 1. Pathological changes in the joints of patients with rheumatoid arthritis.

Leukocyte Infiltration in the Joints. As depicted in the left side of (Figure 1), the healthy joint consists of cartilage layered upon bone that is encased by the synovial membrane, or synovium. By contrast, in the joint of an RA patient (Figure 1, right side), the tissue beneath the synovial lining becomes vascularized and inÞltrated with T cells, B cells, and macrophages. Leukocytes also accumulate in the synovial ßuid, leading to cartilage destruction and bone damage. B cells present in the synovial ßuid characteristically produce large quantities of immunoglobulin G (IgG) or IgM autoantibodies directed against the constant region of other IgG antibodies. These autoreactive antibodies are termed rheumatoid factors (RFs) and form immune complexes that activate the complement cascade and phagocytic activity of macrophages. Both CD4+ and CD8+ (cytotoxic) T cells are present in the synovial ßuid. Researchers note that the CD4+ subset appears to exist in an activated state, presumably owing to the abundance of IL-2 present in the synovial ßuid, which also promotes the initiation of an immune response involving major histocompatibility complex (MHC) class II molecules.

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Synovial Cell Hyperplasia and Tissue Destruction. Researchers believe that synovial tissue macrophages and proliferating Þbroblasts are major contributors to cartilage and bone destruction because both cell populations produce degradative enzymes, such as matrix metalloproteinases (MMPs). Synovial Þbroblasts produce collagenase and cathepsins as well, and they demonstrate proliferative and invasive properties similar to those of transformed tumor cells. Monocytes and macrophages may produce large quantities of pro-inßammatory cytokines, including TNF-α and IL-1β; IL-8, a potent stimulator of angiogenesis; and monocyte chemoattractant protein-1 (MCP-1). This overabundance of proinßammatory cytokines results in persistent inßammation. The clinical success of the TNF-α inhibitor etanercept (Amgen/Wyeth/Takeda’s Enbrel) has resulted in an increased interest in the role of cytokines in RA. Table 1 lists some commonly detected cytokines in the synovial tissue and ßuid of RA patients; Table 2 describes the pathophysiological effects of key cytokines. “Current Therapies” discusses the role of TNF-α in RA pathology. Cytokines in Rheumatoid Arthritis. The RA synovium contains increased levels of cytokines, prominent among which are TNF-α and IL-1. Many other pro-inßammatory cytokines, including IL-6 and IL-18, are also present, as well as anti-inßammatory cytokines such as IL-10, IL-13, and transforming growth factor-beta (TGF-β), suggesting that an imbalance in favor of pro-inßammatory mediators may be a central pathogenic mechanism in RA. An array of cytokines with pro-inßammatory activity in RA synovitis have been described that offer therapeutic potential, including IL-6, IL-12, IL-15, and IL-18. Research has demonstrated that the macrophage-produced cytokine IL-1β plays a dominant role in cartilage degradation and joint invasion by synoviocytes. IL-1 receptor antagonist (IL-1ra) expression correlates with the induction of IL-1β. SpeciÞcally, IL-1ra neutralizes the ability of IL-1β to induce inßammation by binding to IL-1β receptors and inhibiting cellular activation. IL-10, a T helper (Th)2-type cytokine, also has anti-inßammatory properties; it suppresses synthesis of the pro-inßammatory Th1 cytokines IL-1β and TNF-α. Elevated levels of IL-10 are detected in the synovial ßuid of active RA patients, and two laboratories have reported that adenoviral transfer of the vIL-10 gene prevents arthritis development in animal models (Whalen JD, 1999, Lechman ER, 1999). The results of these studies suggest that IL-1ra and IL-10 buffer the pro-inßammatory effects of TNF-α and IL-1β. IL-6, a pleiotropic cytokine, has a wide range of effects, including stimulating B cells to differentiate into plasma cells in order to produce immunoglobulin and stimulating T lymphocytes to differentiate into cytotoxic T cells. Serum IL-6 concentrations have been shown to correlate with disease activity in active RA (Dasgupta B, 1992). IL-12, and the more recently discovered IL-23 and IL-27, comprise a family of structurally related cytokines that regulate cell-mediated immune responses and Th1-type inßammatory reactions. Inhibition of IL-15 and IL-18 represent additional attractive approaches that could block Th1 differentiation, inßammatory mediator production, or TNF-α expression. IL-15 is produced

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TABLE 1. Select Cytokines Detected in the Synovial Tissue of Patients with Rheumatoid Arthritis

Cytokine TNF-α IL-1β IL-2 IL-6

IL-8 IL-10

IL-12 IL-15 IL-17 IL-18

Primary Sources

Targets Activated

Monocytes, macrophages Monocytes, macrophages T lymphocytes Monocytes, macrophages, fibroblasts Monocytes, macrophages Monocytes, macrophages, T lymphocytes Monocytes, macrophages Fibroblasts, monocytes, macrophages T lymphocytes T lymphocytes

Macrophages, fibroblasts Fibroblasts, macrophages T lymphocytes, NK cells Lymphocytes

Degree of Degree of Effect Abundance in on Inflammation Synovial Tissue or Tissue or Fluid Damage High

High

High

High

Medium Medium

Medium Medium

Neutrophils

Medium

Medium

Lymphocytes

Medium

None

Lymphocytes

Low

Medium

Lymphocytes

Low

Medium

Low Medium

Medium Medium

Low Medium

Medium None

Medium

Medium

Fibroblasts Monocytes, macrophages, T lymphocytes, NK cells IFN-γ T lymphocytes NK cells TGF-β Fibroblasts, monocytes, Lymphocytes, fibroblasts macrophages, T lymphocytes GM-CSF Monocytes, Neutrophils, macrophages, macrophages T lymphocytes

GM-CSF = Granulocyte-macrophage colony-stimulating factor; IFN = Interferon; IL = Interleukin; NK cell = Natural killer cell; TGF = Transforming growth factor; TNF = Tumor necrosis factor.

by endothelial cells and is a potent T-cell chemoattractant. It also activates cell adhesion molecule expression on T cells. IL-17, produced by activated CD4+ T cells, is highly expressed in the diseased joint and stimulates formation of bone-resorbing osteoclasts. The pleiotropic cytokine IL-18 acts as a proinßammatory cytokine by inducing TNF-α, interferon-gamma (IFN-γ ), IL-8, IL-2, and prostaglandin production by macrophages and/or T cells, and elevated levels have been detected within the joints of RA patients. Angiogenesis and Rheumatoid Arthritis. Angiogenesis occurs within the synovium early in the pathogenesis of RA. These new vessels support the growing synovial tissue, express chemokines that attract additional cells (including

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TABLE 2. Select Cytokines Involved in the Pathogenesis of Rheumatoid Arthritis Mechanism Synovial tissue inflammation

Cytokines IFN-γ , IL-1β, TNF-α IL-8 IL-1β, IL-2, IL-6, TNF-α IFN-γ , IL-1β, IL-2, IL-6, TNF-α

Synovial fluid inflammation

Synovial proliferation Cartilage and bone damage Systemic manifestations

GM-CSF, IFN-γ , TNF-α GM-CSF, IFN-γ , IL-2 IFN-γ , IL-1β, TNF-α IL-8, TNF-α GM-CSF, IL-8 TNF-α IL-1β, TGF-β TNF-α, TGF-β IL-1β, TNF-α

Role of Cytokine Increased adherence of postcapillary venules Chemotaxis of T cells T-cell activation and proliferation B-cell differentiation and antibody formation Increased expression of HLA antigens Macrophage activation Increased adherence of postcapillary venules Chemotactic for PMN Activation of PMN Fibroblast growth

IL-1β, TNF-α

Neovascularization Activation of chondrocytes, fibroblasts, and osteoblasts/osteoclasts Fever, constitutional symptoms

IL-1β, IL-6, TNF-α

Acute-phase reactants

GM-CSF = Granulocyte-macrophage colony-stimulating factor. HLA = Human leukocyte antigen. IFN = Interferon. IL = Interleukin. PMN = Polymorphonuclear leukocyte. TGF = Transforming growth factor. TNF = Tumor necrosis factor.

leukocytes), and provide an additional source of inßammatory mediators to the synovium. In 1998, researchers proposed that the rheumatoid synovium can be viewed as a rapidly dividing tumorlike structure (Fassbender HG, 1998), a view later endorsed by other researchers. Thus, inhibiting angiogenesis in RA may limit the inßux of inßammatory cytokines and inhibit synovial proliferation. Prognosis. The clinical prognosis of RA varies signiÞcantly. Although a small number of patients display complete remission, and as many as 20–30% experience periods of partial remission, the vast majority suffer chronically. RA patients who are RF positive are more likely to experience a worse disease course. The susceptibility epitope might also inßuence the severity of disease. Patients with HLA-DR1 or HLA-DR4 molecules (see the following section “Genetic Risk Factors”) are at greater risk of developing extra-articular and erosive disease, while individuals who inherit two shared-epitope-containing HLA-DR molecules suffer particularly aggressive disease (Weyand CM,1992). Most patients eventually develop some form of chronic disability, associated with severe pain and progressive functional decline.

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Etiology Genetic Risk Factors. RA occurs in patients’ relatives 2.3 times more frequently than in the general population, suggesting that genetics plays a role in disease development. This suggestion is supported by comparisons of monozygotic twins (derived from one ovum) and dizygotic twins. Studies suggest that the concordance of RA in monozygotic twins is 15.4% versus only 3.6% for dizygotic twins. This concordance extrapolates to an incidence of 12.0 cases per 1,000 per year for monozygotic and 2.2 for dizygotic twins (Silman AJ, 1993; Jawaheer D, 1994). Numerous studies have found a correlation between RA susceptibility and the expression of speciÞc alleles encoding components of the MHC. The MHC is located on the short arm of chromosome 6 and is the only genetic region that has been consistently associated with RA. Much of the MHC comprises the human leukocyte antigen (HLA) genes, which encode an individual’s tissue type and participate in antigen presentation. Two different classes (I and II) of MHC may be expressed on the cell surface for the purposes of antigen presentation. The strongest link to RA within the MHC is the class II HLA region, in particular the HLA-DR locus, and precisely the third hypervariable region of DRβ chains, known as the susceptibility epitope. This epitope is found in multiple HLA-DR genes, including DR1, DR4, and DR14 . Expression of the HLA-DR4 allele has been found in as many as 70% of white and Japanese RA patients, compared with 28% of unaffected individuals. Expression of the HLA-DR1 variant has been associated with RA in Israeli Jews and Asian Indians, and expression of the HLA-Dw16 variant has been associated with RA in the Yakima (Native American) population. Additional HLA-DR alleles—including HLA-Dw4, HLA-Dw14, and HLA-Dw15 —have also been correlated with RA incidence in speciÞc populations (Jaraquemada D, 1986; Ronningen KS, 1992; Lai NS, 1995). (See the earlier “Immune Response” section.) Although HLA-DR alleles show a consistent correlation with RA susceptibility, many researchers believe that RA is polygenic in origin and that subtle differences in the quantitative and qualitative expression of many genes result in a cumulative effect on RA susceptibility. Indeed, the concordance among monozygotic twins with RA is higher than that among siblings bearing identical copies of MHC risk alleles, suggesting a role for additional genes. The North American Rheumatoid Arthritis Consortium (NARAC) was established in 1997 to identify additional genes associated with RA susceptibility using genomic analysis of 1,000 sibling pairs. In a recent analysis, researchers pooled data from 512 families. While conÞrming the association between RA and HLA genes, the study found that genetic regions on chromosomes 1 and 18 are likely to contain genes involved in RA (Jawaheer D, 2003). Another study suggests a role for a single nucleotide polymorphism (SNP) in a gene (PTPN22 ) that encodes for an enzyme known to be a negative regulator involved in controlling T-cell activation. Where the SNP is present, regulation by the enzyme appears inefÞcient such that T cells and other immune cells are hyperresponsive, leading to increased inßammation and tissue damage. The variation is present in approximately 28%

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of individuals with RA compared with 17% of the general population (Begovich AB, 2004). SNP mapping exercises are planned while further studies will determine the actual contribution of such genes to RA susceptibility and their utility as therapeutic targets. Environmental Risk Factors. Despite compelling evidence linking genetic risk factors to the occurrence of RA, long-term concordance rates among monozygotic twins do not exceed 20%, suggesting a role for nongenetic risk factors. Indeed, typically, only one member of an identical twin pair is afßicted with RA. However, while environmental factors appear to interact with, and trigger, RA in genetically predisposed individuals, none has been identiÞed deÞnitively as causative agents. Infectious agents present clear candidates, because many are associated with arthritic disorders. For example, HIV may precipitate an arthritic illness, while infection with Mycobacteria or one of several other pathogens can produce a transient synovitis similar to that characteristic of RA. Persistent Epstein-Barr virus (EBV) has been implicated in a number of studies as having a role in the development of RA, either acting alone or in concert with other viruses such as cytomegalovirus (CMV) or parvovirus B19 (Mehraein Y, 2004). However, no single pathogen or pathogen type is routinely detected in RA patients, and the disease does not occur in clusters or demonstrate seasonal variation. Although some RA cases may develop as a result of infection, the theory that such infection deÞnes the etiology of RA has become less accepted. Some researchers have proposed that RA and other autoimmune diseases result from cross-reactivity of a pathogenic epitope with self-antigens, also known as molecular mimicry. Attention has focused recently on the role of heat-shock proteins such as Mycobacterium tuberculosis hsp65. This protein is highly homologous to the human form, and an immune response against the former could trigger an autoimmune response against the latter (Durai M, 2004). The identiÞcation of such implicated disease-regulating determinants offers the prospect of a potential novel approach for immunotherapy in RA. However, no speciÞc autoantigen (or cross-reactive “self-protein”) has been identiÞed consistently in RA patients. Hormonal Risk Factors. RA is two to three times more common in women than men, with incidence in women steadily increasing with age until menopause. Although RA incidence is rare among younger men compared with younger women, it rises sharply among elderly males, eventually approaching incidence rates in women. Indeed, male RA patients frequently present with decreased testosterone levels, which may suppress normal immunity. Both male and female RA patients exhibit low levels of the female androgen dehydroepiandrosterone (DHEA), and reductions in levels of the steroid appear to precede RA onset. However, DHEA supplementation does not appear to affect RA progression (Giltay EJ, 1998). In the female RA population, disease onset and ßare-ups are most frequent at menopause and in the postpartum period, when estrogen and/or progesterone

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levels fall. In contrast, RA usually remits during pregnancy, likely as a result of the increased production of corticosteroids, estrogens, and progesterones (a deÞciency of these hormones promotes cellular immunity, which is characteristic of RA). Exposure to oral contraceptives—but not estrogen replacement therapy—has been shown to reduce the risk of development of RA (Doran MF, 2004), and the former has been attributed to the decline in RA incidence in women during recent decades. Breast-feeding for more than 12 months has also been shown to lower RA risk (Karlson EW, 2004). Despite these observations, the idea that sex hormones are etiological factors in RA remains contentious, and there are no consistent data on the long-term effects of pregnancy or any protective role for oral contraceptives (Merlino LA, 2003). CURRENT THERAPIES Table 3 lists the current therapies used for the treatment of rheumatoid arthritis (RA). Patients diagnosed with RA typically receive two major forms of therapy. Disease-modifying antirheumatic drugs (DMARDs) are started with a view to halting the destructive course of RA, while anti-inßammatory drugs are prescribed to provide symptomatic relief. This section is organized from the perspective of rheumatologists, who consider the mainstay of treatment to be disease modiÞcation, supported by symptomatic treatment. It Þrst discusses DMARDs, including conventional agents and the more recently introduced biologics. It then discusses symptomatic treatments for RA, including traditional nonsteroidal anti-inßammatory drugs (NSAIDs; including the preferential cyclooxygenase [COX]-2 inhibitors), selective COX-2 inhibitors, and corticosteroids. It also reviews key clinical trials of these agents. In many such trials, researchers measured patient outcomes using the 1987 American College of Rheumatology (ACR) 20, 50, or 70 response criteria, which refer to the percentage of reduction in the signs and symptoms of RA. For example, an ACR 20 response is deÞned as a 20% or greater improvement in the tender and swollen joint count and three of the following Þve outcome measures: physician’s global assessment of disease activity; patient’s global assessment of disease activity; patient’s pain assessment; patient’s disability assessment, typically determined using the health assessment questionnaire (HAQ); and acute-phase reactant levels as measured by C-reactive protein (CRP) or erythrocyte sedimentation rate (ESR). Conventional Disease-Modifying Antirheumatic Drugs Overview. The conventional DMARD class includes a wide range of agents as diverse as antimalarials, cytotoxics, and immunosuppressants. Unlike antiinßammatories, which provide symptomatic relief without altering the natural course of the disease, DMARDs lack a direct analgesic effect but have the potential to slow or prevent joint damage in RA patients (ACR, 2002). It may take months of treatment for the clinical beneÞts of conventional DMARDs to become apparent, and some clinicians and researchers use the term slow-acting antirheumatic drugs (SAARDs) to indicate that these agents take longer than

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TABLE 3. Current Therapies Used for Rheumatoid Arthritis Agent

Company/Brand

Dose

Conventional DMARDs Methotrexate Stada’s Rheumatrex, 7.5–25 mg in a single generics weekly dose Leflunomide Sanofi-Aventis’s Arava 10–20 mg daily Sulfasalazine

Hydroxychloroquine/ chloroquine Biological agents Etanercept

Infliximab

Adalimumab

Anakinra Traditional NSAIDs Diclofenac

Pfizer’s Azulfidine/ Salozopyridine, generics Sanofi-Aventis’s Plaquenil, generics

2,000 mg daily

400 mg daily

Amgen/Wyeth/ Takeda’s Enbrel

Either 25 mg twice weekly or 50 mg once weekly (SC injection, self-administered) Centocor/Schering300 mg every 6-8 weeks Plough/Tanabe (IV perfusion, Seiyaku’s Remicade administered by a health care professional) Abbott/Eisai’s Humira 40 mg every other week (SC injection, self-administered) Amgen’s Kineret 100 mg daily (SC injection, self-administered)

Novartis’s Voltaren/ Voltarol, generics Naproxen Roche’s Naprosyn/ Anaprox/Proxen, Bayer’s Aleve, generics Meloxicam Boehringer Ingelheim/ Abbott/Daiichi’s Mobic Selective COX-2 inhibitors Celecoxib Pfizer/Astellas’s Celebrex Valdecoxib Pfizer/Astellas’s Bextra Etoricoxib Merck’s Arcoxia Corticosteroids Prednisone Pfizer’s Deltasone, generics Methylprednisolone Pfizer’s Depo-Medrol, acetate generics

150 mg daily 1,000 mg daily

Availability US, F, G, I, S, UK, J US, F, G, I, S, UK, J US, F, G, I, S, UK, J US, F, G, I, S, UK US, F, G, I, S, UK

US, F, G, I, S, UK, J

US, F, G, I, S, UK US, F, G, I, S, UK US, F, G, I, S, UK, J US, F, G, I, S, UK, J

10–15 mg daily

US, F, G, I, S, UK, J

400 mg daily

US, F, G, I, S, UK US, G, I, UK I, S, UK

10–20 mg daily 60-120 mg daily 5-25 mg daily (oral)

US, F, G, I, S, UK 40 mg per injection (prn no US, F, G, I, S, more than 3–4 times per UK, J year)

COX-2 = Cyclooxygenase-2; DMARDs = Disease-modifying antirheumatic drugs; IV = Intravenous; NSAIDs = Nonsteroidal anti-inflammatory drugs; prn = As needed; SC = Subcutaneous. US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan.

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anti-inßammatories to achieve an effect. However, DMARDs is the more widely used terminology. In 2002, the ACR published revised guidelines for the treatment of RA. These revised guidelines advocate more aggressive use of DMARDs earlier in the RA treatment regimen in an effort to limit joint damage and minimize loss of joint function and disability. As a result, the use of DMARDs, which were once reserved for treating severe or late-stage RA, has become widespread and is now the cornerstone of RA treatment. Numerous conventional DMARDs are employed in the treatment of RA. The discussion here is limited to the agents that are most commonly employed in current practice. Older DMARDs such as cyclosporine (Novartis’s Neoral, generics), penicillamine (Merck’s Cuprimine, generics), and azathioprine (AZA) (GlaxoSmithKline’s Imuran, generics) are now only infrequently employed in RA. Thus, these drugs are not discussed individually here. Bucillamine (Santen’s Rimatil, generics), actarit (Nippon Shinyaku’s Orcl, Mitsubishi Pharma’s Mover), and lobenzarit (Chugai’s Carfenil) are three conventional DMARDs used in Japan, but these agents are not available in any other region under study and hold only a small fraction of patient share in the RA market, so these agents are not discussed in detail. Mechanism of Action. Conventional DMARDs include a wide range of agents, such as antimalarials, cytotoxics, and immunosuppressants, each with a distinct mechanism of action. In general, agents in this class are anti-inßammatory and/or antiproliferative, inhibiting various molecules and cell types that play a role in the inßammatory cascade. Most have demonstrated the ability to slow the rate of progression of joint erosion and disability to varying degrees. Methotrexate. Originally developed as a treatment for cancer, methotrexate (Stada’s Rheumatrex, generics) (Figure 2) was not widely used in RA therapy until the 1980s. It has since become the leading DMARD in most countries because it offers relatively rapid onset of action (within three to eight weeks), low cost, and arguably the best balance between efÞcacy and tolerability among conventional DMARDs. Although methotrexate’s precise pharmacology is unclear, it is known to inhibit the action of several regulatory enzymes in the folic acid metabolic pathway and to alter leukocyte trafÞcking (Cronstein BN, 1997). Methotrexate is a H2N

N

N CH3

N

N N H N

NH2 O

COOH COOH

FIGURE 2. Structure of methotrexate.

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RHEUMATOID ARTHRITIS

folic acid analogue that interferes with DNA synthesis. Long-term use of this drug at low doses inhibits thymidylate, purine, and methionine production, resulting in the accumulation of a potent anti-inßammatory molecule, adenosine. Methotrexate inhibits cellular proliferation and decreases the formation of antibodies and inßammatory mediators such as cytokines and eicosanoids. Most patients treated with methotrexate exhibit improvement in both subjective and objective measures of disease activity. In a Þve-year study evaluating methotrexate’s long-term efÞcacy among 123 patients, 64% continued taking this agent (Weinblatt ME, 1994). Only 7% discontinued treatment because of lack of efÞcacy, while another 7% withdrew because of adverse effects. Patients who continued therapy exhibited a 70% improvement in the joint pain/tenderness and the joint swelling indices. A ten-year study that enrolled 256 patients who were refractory to treatment with other DMARDs demonstrated that patients who failed to beneÞt from methotrexate therapy were generally those with a poor prognosis (Krause D, 2000). Methotrexate is also the most popular DMARD for use in combination therapy with other conventional DMARDs or with biologics. Recent clinical trials have shown that methotrexate used in combination with a biological agent increases efÞcacy without signiÞcantly increasing adverse effects. In the one-year Trial of Etanercept and Methotrexate with Radiographic Patient Outcomes (TEMPO), 682 patients with active RA were randomly allocated to treatment with etanercept (Amgen/Wyeth/Takeda’s Enbrel) 25 mg (subcutaneously twice a week), oral methotrexate (>20 mg weekly), or the combination. The primary efÞcacy end point was the numeric index of the ACR response (ACR-N) over the Þrst 24 weeks. ACR-N area under the curve (AUC) at 24 weeks was greater for the combination group compared with etanercept or methotrexate alone (18.3% versus 14.7% and 12.2%, respectively) (Klareskog L, 2004). Similarly, in a oneyear study in combination with inßiximab (Centocor [a subsidiary of Johnson & Johnson]/Schering-Plough/Tanabe Seiyaku’s Remicade), 1,049 RA patients were randomly assigned in a ratio of 4:5:5 to three treatment groups: methotrexate with placebo, methotrexate with 3 mg/kg inßiximab, and methotrexate with 6 mg/kg inßiximab. Methotrexate dosages were rapidly escalated to 20 mg/week, and inßiximab or placebo infusions were given at weeks 0, 2, and 6, and every eight weeks thereafter. At week 54, the median percentage of ACR-N improvement was higher for the methotrexate plus 3 mg/kg inßiximab and methotrexate plus 6 mg/kg inßiximab groups than for the methotrexate plus placebo group (38.9% and 46.7% versus 26.4%, respectively) (St Clair EW, 2004). The maintenance dose of oral methotrexate ranges from 7.5 mg (generally 10 mg) to 20.0 mg (25 mg in some practices) in a single, weekly dose. Intramuscular injections are sometimes used to improve bioavailability and to achieve higher dosages. A 2004 study demonstrated that methotrexate given intramuscularly showed improved clinical efÞcacy with fewer side effects than when given orally (Wegrzyn J, 2004). Methotrexate is, however, teratogenic and can cause severe toxicities, including myelosuppression, pneumonitis, nephrotoxicity,

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CF3

O N N O

H CH3

FIGURE 3. Structure of leflunomide.

and hepatotoxicity (Ostensen M, 1998; van Ede AE, 1998). As a result, regular liver function tests, full blood counts, and monitoring of renal function are recommended. Leflunomide. Launched in the United States in 1998, leßunomide (SanoÞAventis’s Arava) (Figure 3) was the Þrst drug to be approved by the FDA for reducing the signs and symptoms of active, adult RA and for retarding structural joint damage—a decision based on the strength of Phase III trial X-ray evidence. In June 2003, the FDA approved additional labeling for leßunomide for improvement in physical function in RA patients. After a lengthy approval process, the agent was launched for use in adults with RA in Europe in 1999 and in Japan in 2003. Patents expire in Europe in 2004 and in 2006 in the United States; the Japanese patent for leßunomide has already expired. Leßunomide is an orally administered isoxazole pro-drug with antiproliferative and anti-inßammatory properties, the latter attributable to direct inhibition of COX-2. The drug’s active metabolite (A77-1726) is believed to block immune-cell proliferation by inhibiting dihydroorotate dehydrogenase, a ratelimiting enzyme involved in de novo pyrimidine synthesis, which is required for lymphocyte turnover. It may also inhibit lymphocyte tyrosine kinases and reduce responsiveness to interleukin (IL)-2 (Fox RI, 1998). Onset of action is as little as four weeks, giving it a signiÞcant advantage over many conventional DMARDs. Its recommended dosage is 100 mg once daily for three days and 10–20 mg once daily thereafter. The relative efÞcacy of leßunomide and methotrexate is controversial, but leßunomide is generally considered to have efÞcacy comparable to that of methotrexate. A double-blind, parallel-group Phase III trial of 999 RA patients, randomized to receive either 10–15 mg weekly methotrexate or 20 mg daily leßunomide (following a 100 mg daily loading dose for three days), found a signiÞcant difference in efÞcacy in favor of methotrexate at one year—50.5% of patients in the leßunomide group were ACR 20 responders compared with 64.8% in the methotrexate group. In fact, the ACR response rate and improvements in all efÞcacy variables with methotrexate were signiÞcantly greater than with leßunomide. However, radiographically assessed disease progression was not statistically different between the two treatments. After two years in 612 patients continuing this trial, ACR 20 response rates were similar with both leßunomide and methotrexate (64.3% and 71.7%). Furthermore, a second, two-year Phase

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III comparative study found that leßunomide efÞcacy and safety were superior to methotrexate (Cohen S, 2001). In this double-blind trial, 235 patients were randomized to receive leßunomide (20 mg daily, loading dose 100 mg for three days) or methotrexate (8–17 mg) weekly. At the end of the study period, 79%, 56%, and 26% of the leßunomide-treated patients achieved ACR 20, ACR 50, and ACR 70 responses, respectively, compared with 67%, 43%, and 20% for methotrexate. Leßunomide was also signiÞcantly superior to methotrexate in improving physical function over the 24 months of treatment in this study. Leßunomide is considered to have a similar or slightly better safety proÞle as compared with methotrexate. Serious adverse events were reported in 1.6% of the leßunomide-treated patients and 3.7% of the methotrexate-treated patients in the 2001 (Cohen) study. In addition to being teratogenic, the most common drugrelated adverse events associated with leßunomide in these clinical trials were diarrhea, liver enzyme abnormalities, rash, reversible alopecia, and hypertension. In an open-label extension study in 96 patients, diarrhea and nausea were less frequent in patients who did not receive a loading dose, with no apparent decrease in efÞcacy (Kremer J, 2004). In a mandatory post-marketing surveillance program in all leßunomide-treated RA patients in Japan (n = 3, 658), interstitial lung disease (ILD) was reported in 0.8%. Twenty-nine cases of interstitial pneumonitis were reported, 11 of which proved fatal. However, the causality link between leßunomide use and the reported ILD was frequently confounded by pre-existing pulmonary disease and previous or concomitant use of other DMARDs. These events are being evaluated by SanoÞ-Aventis, which has recommended that pulmonary status be assessed prior to the initiation of leßunomide, together with close monitoring of patients during treatment. Leßunomide is frequently employed in combination with other DMARDs. According to the opinion obtained at an International Expert Panel Meeting held in Paris in May 2003, 61% of the Expert Panel would use leßunomide with methotrexate, 71% with sulfasalazine (PÞzer’s AzulÞdine/Salozopyridine, generics), 43% with inßiximab, 38% with anakinra (Amgen’s Kineret), 33% with adalimumab (Abbott/Eisai’s Humira), and 19% with etanercept (Kalden JR, 2004). Sulfasalazine. Sulfasalazine (PÞzer’s AzulÞdine/Salozopyridine, generics) (Figure 4) is available in all the markets under study, including Japan, where it is called “salazosulfapyridine.” It is a conjugate of salicylic acid, belonging to the sulfonamide drug class. The drug is metabolized to two key components: sulfapyridine, which has antibacterial activity, and 5-aminosalicylic acid, which is an anti-inßammatory (Rains CP, 1995). Although sulfasalazine suppresses disease activity in RA, its mode of action as a disease modiÞer remains unknown. Sulfasalazine was shown to have superior efÞcacy compared with placebo in a 48-week, double-blind, prospective study (Hannonen P, 1993). The study enrolled 80 patients with early RA, and study participants were randomized to receive either 2 g sulfasalazine daily or placebo; patients were also allowed concomitant NSAID and corticosteroid therapy. At the end of the 48-week study,

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COOH

N NHSO2

OH

N

N

FIGURE 4. Structure of sulfasalazine.

patients treated with sulfasalazine were shown to have statistically signiÞcant improvements compared with placebo on three of the four measures of efÞcacy: joint tenderness (measured by a modiÞed Ritchie articular index; score range 0–90), number of swollen joints, and patient’s global assessment of disease activity (measured by a 5-point scale); the difference in improvement between the two treatment groups for the physician’s global assessment of disease activity was not signiÞcant. Improvements on these efÞcacy measures were greater for rheumatoid factor seronegative (RF-) early RA patients compared with seropositive (RF+) early RA patients. RF− and RF+ patients taking sulfasalazine had mean improvements of 5.46 and 1.68 points, respectively, on the Ritchie articular index. Corresponding mean changes for the placebo group were a 1.69-point improvement for RF- patients and a 0.24-point worsening for RF+ patients. The mean number of swollen joints improved by 4.39 and 1.77 for RF- and RF+ patients taking sulfasalazine, respectively, while RF- and RF+ patients taking placebo had a 0.23 improvement and a 0.76 worsening, respectively, in the number of swollen joints. Lastly, mean improvements in the global assessment of disease activity were 0.92 and 0.23 for RF- and RF+ patients taking sulfasalazine, respectively. RF- and RF+ patients taking placebo had corresponding improvements of 0.08 and 0.04, respectively. Sulfasalazine is used alone for mild disease and in combination with other conventional DMARDs, principally methotrexate, in moderate RA when the latter has proven inadequate as monotherapy. Once highly popular in Europe, sulfasalazine has been displaced as the Þrst-choice DMARD by methotrexate, primarily because the latter is seen as a more effective agent. The dosage of sulfasalazine is 500 mg twice daily taken orally, with increases of up to 2–3 g per day in divided doses. Onset of action requires three to eight weeks, with approximately 70% of patients demonstrating a satisfactory clinical response within one year (Rains CP, 1995). The agent’s use is constrained by its sideeffect proÞle. Gastrointestinal symptoms, the most common side effect, are often resolved with dose attenuation, but some rare but serious hematological reactions necessitate regular blood tests. Occasionally, hepatotoxicity, pneumonitis, and myelosuppression occur (Scott DL, 1988). Hydroxychloroquine/Chloroquine. Originally developed as antimalarials, these agents are employed principally in treating mild RA. Because of its superior side-effect proÞle, hydroxychloroquine (SanoÞ-Aventis’ Plaquenil, generics) (Figure 5) is more popular than chloroquine (generics), its parent compound.

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FIGURE 5. Structure of hydroxychloroquine.

Researchers believe that hydroxychloroquine and chloroquine inhibit antigen presentation and block receptor-mediated endocytosis, granulocyte migration, the action of phospholipase A2 , and the release of cytokine IL-1β (Fox RI, 1993). The efÞcacy of hydroxychloroquine was compared with that of placebo in a six-week double-blind, randomized trial involving 126 patients with early RA (Clark P, 1993). Patients were randomized to receive either 400 mg daily hydroxychloroquine or placebo. Treatment with hydroxychloroquine was associated with clinically and statistically signiÞcant improvements over placebo on four efÞcacy measures used in the study: joint score (sum of scores for joint swelling [58 joints graded on a 0–3 point scale] and joint tenderness [60 joints graded on a 0–3 point scale])—the primary end point of the trial; patients’ assessment of joint pain (using a 10-cm visual analogue scale [VAS]); grip strength (measured by a mercury-column sphygmomanometer in mm Hg); and ESR (measured in mm/hr). Mean improvements for joint score, patient’s assessment of pain, grip strength, and ESR were 18.5 points, 25.8 mm, 31.9 mm HG, and 5.6 mm/hr, respectively, for the group treated with hydroxychloroquine; the corresponding improvements for the placebo group were 11.9 points, 6.5 mm, 10.7 mm Hg, and 4.3 mm/hr. Hydroxychloroquine was well tolerated in this study with no difference in the overall incidence of adverse events between the active and placebo groups, although the incidence of headache was signiÞcantly higher in the treated group. Prescribed in oral daily dosages of 200 mg to 400 mg, antimalarials are employed as an alternative to sulfasalazine in early disease. Support for the use of hydroxychloroquine in the treatment of early RA came in a recent study that demonstrated that, after two years, a majority of early RA patients (56/94) were controlled on hydroxychloroquine or were in remission (Matteson EL, 2004). Much of current prescribing of antimalarials is, however, in dual or triple combinations with other conventional DMARDs, particularly methotrexate and sulfasalazine (Jobanputra P, 2004). Nausea or dizziness may accompany use, although the most serious side effect is irreversible macular damage (Jones SK, 1999), so regular ophthalmologic exams are recommended. Biological Agents Overview. Biological DMARDs for the treatment of RA include the tumor necrosis factor-alpha (TNF-α) inhibitors etanercept, inßiximab, and adalimumab,

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FIGURE 6. Role of select cytokines in rheumatoid arthritis pathogenesis.

as well as the IL-1β receptor antagonist anakinra. TNF-α is a potent proinßammatory cytokine. Its role in RA has been demonstrated in transgenic mouse models in which TNF-α overexpression has led to joint destruction and aggressive synovitis (Figure 6). Among RA patients, the degree of synovial TNF-α expression correlates with the degree of synovitis and bone erosion. In addition to TNF-α, many other proinßammatory cytokines—including IL-1β —are involved in the pathogenesis of RA. Current biological agents antagonize the destructive role of such cytokines in the disease. Mechanism of Action. Receptors for TNF-α are found on the surface of most cells, including mononuclear cells and cells in the synovium. Two distinct types of TNF-α receptors have been identiÞed: type I (p55) and type II (p75). Cleavage of membrane-bound TNF-α receptors yields soluble receptors that retain ligandbinding ability but cannot activate cells. TNF-α inhibitors reduce free, bioactive TNF-α by emulating the physiological role played by soluble TNF-α receptors, which modulate the amount of circulating, bioactive TNF-α by binding

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to the cytokine before it can activate cell-surface receptors on mononuclear cells. Because TNF-α plays an important role in the eradication of neoplastic cells, its suppression is not without hazards—particularly as a long-term strategy. Concerns have been raised as to whether chronic immunosuppression leads to opportunistic infection, malignancies, or other complications (Alldred A, 2001; Lee JH, 2002). In October 2004, the FDA recommended a warning concerning malignancy be added to the labeling for all TNF-α inhibitors stating that in controlled studies, more cases of lymphoma have been observed among patients receiving TNF-α inhibitors than among control group patients. In practice, rheumatologists trade the beneÞts of anti-TNF-α therapy in treating this serious disease with persistent worry over possible long-term effects. Such concerns remain particularly pronounced when initiating lifelong treatment in young patients. Inßammation induces IL-1 production in macrophages, in turn mediating various inßammatory and immunological responses including cartilage degeneration, stimulation of bone resorption, and joint invasion by synoviocytes. Anakinra, the Þrst interleukin-modulating therapy indicated for RA, competitively inhibits the activity of IL-1β, thus limiting the degrading effects of the cytokine. Etanercept. Launched in 1998 in the United States, etanercept (Amgen/ Wyeth/Takeda’s Enbrel) was the Þrst TNF-α inhibitor approved for the treatment of RA. It was initially indicated for moderate to severe RA patients unresponsive to one or more conventional DMARDs or for use in combination with methotrexate for patients unresponsive to methotrexate alone. It has since been granted orphan drug approval for use in juvenile RA patients. In June 2000, the FDA expanded the agent’s label to include reducing symptoms and delaying structural damage in patients with moderate to severe, active RA at an early stage. In July 2003, the FDA approved a further indication extension for use in improving physical function in patients with moderate to severe, active RA, while in October 2003, approval was also gained for a 50 mg once-weekly dosage for adults (compared with the then-standard dosage of 25 mg twice weekly). This development was followed by the approval in September 2004 of a 50 mg preÞlled syringe formulation in the United States. In the same month, etanercept became the only FDA-approved biologic to induce a major clinical response in RA patients, deÞned as achieving an ACR 70 response for six consecutive months. This agent is marketed by Amgen (formerly Immunex) in conjunction with Wyeth in the United States and by the latter in Europe. It was approved by the European Agency for the Evaluation of Medicinal Products (EMEA) for use in adult and juvenile RA patients in the 15 European Union countries in February 2000 and was launched later that year. Japanese approval was granted to copromoters Wyeth and Takeda in January 2005, and the drug launched in March 2005. Patents expire in 2010 in Japan, in 2014 in the United States, and in 2015 in Europe. Etanercept is a soluble TNF-α receptor fusion protein, consisting of a portion of the soluble p75 TNF-α receptor fused to the Fc fragment of human

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immunoglobulin G (IgG)1. The TNF-α receptor component binds free TNF-α protein, and the antibody component can mediate effector functions such as initiation of the complement system and phagocytosis. Etanercept was bioengineered to have a higher binding afÞnity for TNF-α than the natural receptor. Moreover, the presence of an Fc fragment extends the agent’s half-life approximately Þvefold to eightfold in vivo. TNF-α inhibitors tend to have a faster onset of action than conventional DMARDs, and etanercept’s effects become apparent within two to three weeks. A two-year, double-blind trial designed to compare clinical and radiographic outcomes in early, aggressive RA showed that etanercept is superior to methotrexate in reducing disease activity, arresting structural damage, and decreasing disability (Genovese MC, 2002[a]). The trial enrolled 632 patients who had suffered from RA for up to three years. Patients were randomized to receive etanercept (10 or 25 mg) twice weekly or methotrexate (mean dose 19 mg) weekly. The primary end points were ACR 20 response and X-ray progression as measured according to the Sharp score. (The Sharp score is the sum of a measure of joint space narrowing and a measure of bone erosion; increases in Sharp scores indicate a worsening of disease.) Although all three treatments effectively reduced the signs and symptoms of RA, signiÞcantly more patients in the etanercept 25 mg group achieved an ACR 20 response (72% versus 59% with methotrexate). Patients receiving etanercept 25 mg also experienced the most dramatic reduction in radiographic progression. The mean increase in Sharp score was 1.3 units in the etanercept 25 mg group compared with 3.2 units in the methotrexate group. Approximately 15% of patients in the etanercept groups withdrew because of adverse events versus 21% in the methotrexate group. Injection site reactions were the most common adverse event in the etanercept-treated group. Etanercept has also been investigated in combination with methotrexate in the ongoing, two-year TEMPO study of 686 patients with RA. As assessed by the Disease Activity Score (DAS) at 52 weeks, signiÞcantly more patients (35%) receiving the combination of etanercept and methotrexate achieved clinical remission, compared with 16% on etanercept alone and 13% on methotrexate alone (Klareskog L, 2004). No radiographic progression of joint damage, as assessed by Sharp score, was seen at two years in 74.2% of patients taking the combination therapy, compared with 65.5% and 59.2% of etanercept and methotrexate alone, respectively. Patients treated with the combination experienced a 56% mean improvement in HAQ scores from baseline, compared with 39% with etanercept alone and 36% with methotrexate alone (Amgen, press release, October 18, 2004). In August 2001, Immunex (acquired by Amgen in 2002) and Wyeth announced the initiation of the largest clinical trial to date to evaluate the impact of etanercept in U.S. RA patients. The trial, known as the Rheumatoid Arthritis DMARD Intervention and Utilization Study (RADIUS), will compare the safety, efÞcacy, and treatment patterns of 5,000 RA patients treated with etanercept in the second phase of the study with those of 5,000 patients treated with a variety of other DMARDs in the Þrst phase of the study. In July 2003, Amgen announced the

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completion of enrollment of 5,000 patients at more than 400 study sites for the second phase of the study. Data will be collected for at least Þve years. Studies have shown that etanercept may cause hematological reactions and demyelinating disorders, the latter in patients with preexisting central nervous system demyelinating disease (Alldred A, 2001; Mohan N, 2001). In addition, several patients developed serious infections during postmarketing surveillance, resulting in the inclusion of a warning on etanercept’s label advising caution in patients with a history of recurring infections or predisposition to infection. Observed event rates among 2,054 clinical trial participants eligible to enroll in extension studies with a combined etanercept exposure of 7,382 patient-years were presented at the 2004 ACR Annual Congress. Rates of serious adverse events, serious infections, opportunistic infections, and deaths did not increase with more than seven years of treatment. However, the incidence of lymphoma was higher than that expected in the general population (67 versus 61 cases), and the authors called for additional studies to determine whether this Þnding reßects inherent risk in RA or is related to etanercept treatment (Moreland LW, 2004[b]). Infliximab. Inßiximab (Centocor [a subsidiary of Johnson&Johnson]/ScheringPlough/Tanabe Seiyaku’s Remicade) was the second TNF-α inhibitor to make a major impact on the treatment of RA. It is marketed in the United States by the originator, Centocor; by licensee Schering-Plough in Europe; and by Tanabe Seiyaku in Japan, where it has been the only TNF-α inhibitor available until the 2005 launch of etanercept. Initially launched in the United States in 1998 for the short-term treatment of Crohn’s disease, inßiximab received FDA approval in 1999 for use in combination with methotrexate to treat adult RA patients refractory to methotrexate alone. In December 2000, labeling was expanded for use of the same regimen in inhibiting the progression of structural damage. In February 2002, supplemental FDA approval was gained for improving physical function in patients with moderate to severe, active RA. In September 2004, the FDA approved expanded labeling for inßiximab in combination with methotrexate as a Þrst-line regimen for moderate to severe RA, eliminating the requirement that patients must Þrst fail to respond to methotrexate alone. Inßiximab received centralized marketing authorization from the EMEA in June 2000 for use in combination with methotrexate for the reduction of signs and symptoms in RA patients with active disease showing inadequate response to previous DMARDs, including methotrexate. In June 2004, approval was granted in Europe as Þrst-line therapy for the treatment of early RA. Additional labeling was approved in Europe in July 2004 for ankylosing spondylitis, while in September 2004, the EMEA approved further labeling for inßiximab in combination with methotrexate for the treatment of active and progressive psoriatic arthritis in patients responding inadequately to DMARDs. In December 2004, approval was gained in the United States for the treatment of psoriatic arthritis in patients with active disease. Inßiximab was launched in Japan for Crohn’s

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disease in May 2002 and for RA in July 2003. Patents expire in 2012 in Europe and Japan and in 2016 in the United States. Inßiximab is a chimeric monoclonal antibody composed of 75% human and 25% mouse protein. The drug exerts its therapeutic effect by binding soluble and membrane-bound TNF-α, thereby blocking its ability to activate the inßammatory cascade. The mouse portion of the antibody contains the TNF-α binding sites, and the human portion is responsible for effector function, including IgGstimulated elimination of TNF-α positive cells by both antibody-dependent cellular cytotoxicity and complement-dependent mechanisms (Feldmann M, 2001). Inßiximab may also inhibit the upregulation of cell-surface adhesion proteins—a process that plays an important role in mediating inßammation. The development of human antichimeric antibodies has been reported in as many as 50% of patients across various clinical studies, or more than twice the rate associated with etanercept use (Moreland LW, 2001). No clinically signiÞcant effects have been observed as a result of these antibodies, although debate regarding their importance is ongoing. Inßiximab must always be used in conjunction with methotrexate because doing so appears to help reduce antibody production. The inclusion of the murine component leads some physicians to prefer other anti-TNF-α agents, as they are perceived to carry less of an immunological risk. The approval of inßiximab for improving physical function was based on data from a Phase III trial, the Anti-TNF-α Trial in Rheumatoid Arthritis with Concomitant Therapy (ATTRACT) (Wong JB, 2002). Study participants had active RA for an average of 8.4 years, had failed to respond to a median of three DMARDs, and had previously experienced an insufÞcient response to methotrexate alone. In this multicenter trial conducted at 34 centers in the United States and Europe, 428 patients were randomized to receive inßiximab (3 mg/kg or 10 mg/kg) or placebo for one year. All received concomitant methotrexate (10–15 mg weekly). Inßiximab was administered at zero, two, and six weeks, and additional doses were given every four or eight weeks thereafter. ACR 50 response criteria were achieved in 29, 27, 26, and 31% of patients receiving 3 mg/kg every four or eight weeks or 10 mg/kg every four or eight weeks, respectively, compared with 5% of patients receiving placebo plus methotrexate. In a second Phase III study, the ASPIRE trial (Active Controlled Study of Patients Receiving Inßiximab for Treatment of Rheumatoid Arthritis of Early Onset), a total of 1,004 patients with early RA were assigned to receive infusions of placebo or inßiximab at weeks 0, 2, and 6, and then every eight weeks thereafter to week 46. All patients received concomitant methotrexate. At the 2004 ACR Annual ScientiÞc Meeting, Centocor presented new Þndings from this study which formed the basis for the expanded indication as Þrst-line therapy in patients with moderate to severe, active RA. An erosion-free state was observed in 79% of patients who began the study with no joint erosions that were treated with inßiximab plus methotrexate for one year, compared to 58% of patients treated with methotrexate alone (Centocor, press release, October 18, 2004).

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Adverse events associated with inßiximab are generally mild. They include headache, nausea, rash, coughing, sinusitis, dyspnea, urticaria, hypotension, and upper respiratory tract infection. Although the incidence of malignancies in clinical trials is not greater than with placebo, experts remain concerned about the agent’s long-term safety, particularly with regard to chronic immunosuppression. Inßiximab’s packaging has long included a disclaimer stating that patients should discontinue therapy if they develop a serious infection or sepsis. Following an FDA recommendation, Centocor also included a boxed warning on the package insert noting an increased risk of developing tuberculosis and other opportunistic infections. The warning was based on the occurrence of more than 80 cases of tuberculosis in inßiximab-treated patients worldwide. In August 2001, following the completion of clinical trials in congestive heart failure, Centocor/ScheringPlough/Tanabe Seiyaku issued a warning stating that inßiximab therapy should not be initiated in patients with concomitant congestive heart failure. The companies have discontinued trials in this indication because of adverse events. By August 2004, worldwide postmarketing experience revealed hematological events in patients receiving inßiximab, including leukopenia, neutropenia, thrombocytopenia, and pancytopenia—some with fatal outcomes. Accordingly, a warning statement was added to inßiximab labeling. Inßiximab is administered in 3 mg/kg doses given as a two-hour infusion every eight weeks, following a standard induction regimen requiring treatment at weeks 0, 2, and 6. The regimen requires administration directly by caregivers in the clinic or ofÞce setting. Most patients who respond do so within two to three weeks. Adalimumab. Like other TNF-α inhibitors, adalimumab (Abbott/Eisai’s Humira) blocks the activity of TNF-α by binding to this cytokine. Discovered by Cambridge Antibody Technology and developed by Abbott Laboratories, adalimumab is the Þrst fully human monoclonal anti-TNF-α antibody to be marketed. Abbott and GTC Biotherapeutics have established a strategic partnership in which the latter will produce adalimumab in transgenic goats. Adalimumab was launched in the United States for RA in January 2003. It is approved for reducing the signs and symptoms of RA and inhibiting the progression of structural damage in adults with moderate to severe, active RA who have had insufÞcient response to one or more DMARDs. In August 2004, the labeling was expanded to include improvement in physical function for adult patients with moderate to severe, active RA. The product was launched in September 2003 in Germany and the United Kingdom, followed by other European countries in subsequent months. In Europe, adalimumab is approved for the treatment of moderate to severe, active adult RA following inadequate response to DMARDs, including methotrexate. In May 2004, Abbott announced that the EMEA had granted a “positive opinion” for an RA label extension for reducing the rate of progression of joint damage as measured by X ray and improving physical function in adults when given in combination with methotrexate. In Japan, adalimumab is licensed to Eisai and is in Phase II development for RA. Patents expire in 2014 in the United States and in 2017 in Europe and Japan.

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Patients with chronic RA treated with adalimumab in combination with methotrexate demonstrated signiÞcant clinical improvements in Phase II and III clinical trials. In a 52-week, double-blind study, 619 patients with active RA were randomized to receive adalimumab (20 mg weekly or 40 mg every other week) plus methotrexate or placebo. A total of 467 patients (75.4%) completed the full course of treatment. At week 52, there was statistically signiÞcantly less radiographic progression, as measured by the change in Sharp score, in the patients receiving adalimumab either 40 mg (mean change 0.1) or 20 mg weekly (0.8) compared with placebo (2.7). ACR 20 responses were achieved by 59% and 55% of patients taking adalimumab 40 mg and 20 mg, respectively, versus 24% of patients taking placebo. Physical function, as measured by HAQ scores, demonstrated signiÞcant improvement with adalimumab 40 mg (mean change in HAQ score −0.59) and 20 mg (−0.61) compared with placebo (−0.25) (Keystone EC, 2004[b]). To evaluate adalimumab immunotherapy in RA where previous DMARD treatment had failed, 544 patients were randomized to monotherapy versus placebo. After 26 weeks, patients treated with adalimumab 20 mg every other week, 20 mg weekly, 40 mg every other week, and 40 mg weekly had a statistically significantly better ACR 20 response rates than those treated with placebo (35.8%, 39.3%, 46.0%, and 53.4%, respectively, versus 19.1%) (van de Putte LB, 2004). The most frequently observed adverse events in clinical trials of adalimumab were injection site reactions, upper respiratory infection, injection site pain, headache, hypertension, rash, and sinusitis. In November 2004, Abbott issued revised warnings including serious infections with the combined use of adalimumab and anakinra, hypersensitivity reactions including anaphylaxis, hematologic events including pancytopenia, and aplastic anemia. The product label also acknowledges that malignancies have been observed in patients treated with adalimumab and that there may be an increased risk of lymphoma with TNF-α inhibitor therapy. To ensure maximum efÞcacy, adalimumab is typically given in combination with methotrexate. Adalimumab can also be given as monotherapy in cases of intolerance to methotrexate or when continued treatment with methotrexate is inappropriate. Because it is a fully human antibody, this agent is expected to cause less immunogenicity than the chimeric monoclonal antibody inßiximab, and adalimumab may eventually replace inßiximab for this reason. Adalimumab, given as twice-monthly 40 mg subcutaneous injections, is also more convenient than either inßiximab or etanercept. However, adalimumab may have greater difÞculty displacing etanercept, a drug already favored by many rheumatologists. Adalimumab’s reduced immunogenicity is unlikely to confer a signiÞcant advantage over etanercept because this problem occurs only infrequently following long-term etanercept use. The most signiÞcant barrier to adalimumab displacing etanercept is physician comfort and experience with etanercept. Therefore, adalimumab will likely be used as a second-line TNF-α inhibitor, following treatment failure with etanercept.

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In an attempt to tip the scale in favor of the use of adalimumab over etanercept in the U.S. RA market, Abbott announced the start of the Humira Medicare Assistance Program in January 2003. This drug access program will provide adalimumab to Medicare-eligible senior citizens in the United States. Only inßiximab is currently reimbursed by Medicare, given that self-administered therapies, like etanercept and adalimumab, are not covered. Starting in 2006, however, the Centers for Medicare and Medicaid Services—a federal agency within the U.S. Department of Health and Human Services—is providing access to selfadministered prescription medicines through the Medicare replacement drug demonstration program. Adalimumab will already be well established in the elderly U.S. population when the restrictions on the reimbursement of self-administered drugs by Medicare recipients are relaxed, giving it a potential advantage over etanercept. Anakinra. Anakinra (Amgen’s Kineret) is an IL-1β receptor antagonist that competitively inhibits the activity of IL-1β, a macrophage-produced cytokine that plays a dominant role in cartilage degeneration and joint invasion by synoviocytes. The drug was launched in the United States in December 2001 for the treatment of RA in patients with moderate to severe disease failing to respond to one or more DMARDs. In March 2002, anakinra was approved in Europe for the treatment of RA in combination with methotrexate in patients with an inadequate response to methotrexate alone. By June 2002, the agent was launched in Germany, the United Kingdom, and a number of smaller European markets. Amgen and the University of Colorado Foundation have been granted a U.S. patent for the IL-1β inhibitor sequences and the methods for producing them, thereby securing rights to this and future IL-1β inhibitory molecules. Patents expire in 2009 in France, Germany, and Japan; in 2014 elsewhere in Europe; and in 2017 in the United States. A 24-week, multicenter study designed to evaluate the safety and efÞcacy of the agent in 506 patients with moderate to severe active RA showed that anakinra combined with methotrexate is superior to methotrexate alone (Cohen SB, 2004). Patients received subcutaneous injections of anakinra 100 mg/day or placebo in a single daily subcutaneous injection. All patients received concomitant methotrexate (10–25 mg weekly), and the primary end point was an ACR 20 response at 24 weeks. SigniÞcantly greater proportions of patients treated with anakinra compared with placebo achieved ACR 20 (38% versus 22%), ACR 50 (17% versus 8%), and ACR 70 (6% versus 2%) responses. Data presented at the 66th annual ACR meeting in 2002 demonstrated that anakinra inhibits bone and joint damage in RA patients (Shergy W, 2002). In the study, 906 patients were randomized to receive anakinra (100 mg) or placebo for 52 weeks. All patients received concomitant methotrexate (10–25 mg weekly), and the primary end point was X-ray progression as measured according to the Sharp score. At the end of the study period, 50% of the anakinra-treated patients showed no disease progression compared with 42% for placebo. Adverse events, events leading to withdrawal, and infectious events were similar in the two treatment groups. Based on the results of this study, Amgen gained additional

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labeling for slowing the progression of structural damage in moderate to severe, active RA. In a study to examine the safety of anakinra when added to background DMARD therapy, patients with RA were randomly assigned in a 4:1 allocation ratio to treatment with anakinra 100 mg or placebo administered daily by injection. Anakinra patients (n = 1, 116) showed no difference in the incidence of upper respiratory infections or overall serious adverse events compared with placebo patients (n = 283). The anakinra group had more injection site reactions (72.6% versus 32.9%), and a small increase in serious infections (2.1% versus 0.4%) was observed (Tesser J, 2004). An increased incidence of adverse events such as infections were reported with anti-TNF-α and IL-1 receptor antagonist combination therapy trials in RA patients, prompting the FDA’s Arthritis Advisory Committee to release a statement in August 2001 (at the time of anakinra’s approval) advising against such therapy. Anakinra requires daily administration by 100 mg subcutaneous injection because of its rapid clearance, which is seen as a drawback in comparison with the TNF-α inhibitors. This agent has assumed a niche role in patients refractory to conventional DMARDs and the TNF-α inhibitors. Traditional Nonsteroidal Anti-Inflammatory Drugs Overview. Generally, the Þrst course of therapy for RA initiated by primary care physicians has been nonsteroidal anti-inßammatory drugs (NSAIDs), which are mainly administered to help relieve pain and inßammation. Their analgesic effect is felt quickly and lasts just a few hours, whereas their anti-inßammatory properties become apparent only after several days of repeated administration. However, NSAIDs are unable to halt the progression of RA or prevent joint damage caused by the disease. This symptomatic treatment approach covers the use of both older NSAIDs and the more recently introduced selective (COX-2) inhibitors. The older NSAIDs are often referred to as nonselective relative to the highly selective COX-2 inhibitors, but these agents actually demonstrate a spectrum of selectivity for the COX enzymes, ranging from preferential COX-1 inhibition to preferential COX-2 inhibition. In line with industry convention, the discussion here groups the older, less selective agents (including the preferential COX-2 inhibitors) together under the heading of traditional NSAIDs and considers them separately from the selective COX-2 inhibitors. Numerous traditional NSAIDs—many of which are available as inexpensive over-the-counter (OTC) preparations—are available to relieve the pain and inßammation associated with RA. However, treatment of moderate to severe pain generally requires dosing that is available only by prescription. Because of the large number of traditional NSAIDs available on the market, this section discusses only three of the most popular agents used in the treatment of RA—diclofenac (Novartis’ Voltaren/Voltarol, generics), naproxen (Roche’s Naprosyn/Anaprox/Proxen, Bayer’s Aleve, generics), and meloxicam (Boehringer Ingelheim/Abbott/Daiichi’s Mobic)—as representatives of this class.

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Mechanism of Action. Although the mechanism of NSAID action is not fully understood, these drugs are known to block the arachidonic acid pathway at an early stage. Arachidonic acid is the major fatty acid incorporated into cell membranes, and its metabolites serve as precursors to the synthesis of prostaglandins. These inßammatory mediators, especially prostaglandin E2 (PGE2 ), cause inßammation by increasing the vascular permeability of blood vessels. Elevated expression of prostaglandins has been detected in the synovium of arthritic joints. The COX-1 and COX-2 enzymes initially convert arachidonic acid into intermediary cyclic endoperoxides, which, in turn, are converted into prostaglandins. Traditional NSAIDs inhibit the activity of one or both of the COX enzymes, thereby preventing the production of prostaglandins (see Figure 7). Although NSAIDs are highly effective at relieving symptoms of pain and inßammation, their use is complicated by serious side effects. Researchers now recognize that both the desired therapeutic effects and the main side effects of NSAIDs are attributable largely to the suppression of PGE2 . In addition to mediating inßammation, PGE2 , as well as other prostaglandins, has a protective role in the gastrointestinal (GI) tract and kidneys. The most common and disturbing adverse events associated with traditional NSAIDs are impaired renal function and GI complications, including GI hemorrhage, ulcers, perforations, and obstructions. In a large study evaluating treatment outcomes of RA patients receiving NSAID therapy for an average of 2.5 years, 15% of patients experienced

FIGURE 7. The role of tumor necrosis factor-alpha in the pathogenesis of rheumatoid arthritis.

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an NSAID-induced GI side effect (Singh G, 1996). To mitigate the suppression of PGE2 , some physicians prescribe proton pump inhibitors or misoprostol (PÞzer’s Cytotec) along with NSAID therapy. Proton pump inhibitors block the production of acid in the GI tract, minimizing the formation of ulcers, and misoprostol is a synthetic PGE1 analogue that protects the lining of the stomach by mimicking the action of naturally occurring prostaglandins. To facilitate dosing convenience, combination products combining a traditional NSAID with either a proton pump inhibitor or misoprostol are now available. For example, TAP Pharmaceutical Products (a joint venture between Abbott and Takeda) markets Prevacid Naprapac, a combination of the proton pump inhibitor lansoprazole (TAP’s Prevacid) and naproxen, while PÞzer markets Arthrotec, a combination of misoprostol with diclofenac. Diclofenac. Diclofenac (Novartis’s Voltaren/Voltarol, generics) (Figure 8) is a phenylacetic acid derivative. In addition to inhibiting the COX enzymes, diclofenac decreases the availability of arachidonic acid and inhibits production of leukotrienes. It is a rapidly metabolized NSAID with a plasma half-life of less than two hours, requiring multiple daily doses. The efÞcacy of diclofenac was compared with that of placebo in a two-week double-blind trial of 44 RA patients (Doreen MS, 1978). Patients were given either placebo or 75 mg diclofenac daily during the Þrst week; diclofenac doses were increased to 75–150 mg daily during the second week of the study. Patients were also allowed to take acetaminophen as a rescue analgesic. After the Þrst week of the study, 50% of patients treated with diclofenac reported an improvement in their pain, compared with 24% of placebo recipients; this difference was statistically signiÞcant. After the second week of treatment, 60% and 33% of diclofenac- and placebo-treated patients, respectively, reported improvement in pain; this difference was not statistically signiÞcant. However, acetaminophen consumption was signiÞcantly lower for patients treated with diclofenac compared with those receiving placebo. At the end of the 14-day treatment period, improvements in grip strength (measured by the sum of three readings for each hand using a grip bag inßated to 30 mm Hg) and articular index of joint tenderness (measured using the Ritchie scale with a maximum score of 69) were signiÞcantly superior to those of the placebo group. The diclofenac group demonstrated

COOH NH Cl

Cl 3' 4'

FIGURE 8. Structure of diclofenac.

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a 15 mm Hg increase in grip strength and an improvement of 3.5 on the articular index, compared with a 40 mm Hg worsening on grip strength and an improvement of 0.8 on the articular index for the placebo-treated group. Duration of morning stiffness decreased by 20 minutes for patients treated with diclofenac and increased 18 minutes for placebo recipients at the end of the study period. Six patients in the diclofenac group reported minor GI side effects and/or headaches and dizziness, compared with ten patients treated with placebo. Like other NSAIDs, diclofenac’s side effects are predominantly GI-related, including dyspepsia, epigastric pain, vomiting, and diarrhea (Singh G, 1996). Hepatotoxicity is also a concern. According to 180 cases reviewed by the FDA between 1988 and 1991, diclofenac poses a risk of liver damage to all patients, particularly women (Banks AT, 1995). Naproxen. Naproxen (Roche’s Naprosyn/Anaprox/Proxen, Bayer’s Aleve, generics) (Figure 9), a naphthaleneacetic acid derivative, is a long-acting traditional NSAID available OTC and by prescription. The OTC preparations of this drug are all sodium salts (naproxen sodium), but prescription formulations are either naproxen (Naprosyn) or naproxen sodium (Anaprox). The sodium salt was developed to enable faster drug dissolution, thus speeding absorption and onset of action. A controlled-release formulation of naproxen sodium (Wyeth/Elan’s Naprelan, generics) is also available. Like other traditional NSAIDs, naproxen inhibits the activity of the COX enzymes, thereby preventing the production of prostaglandins. By inhibiting prostaglandin production, naproxen is able to reduce pain and inßammation. In a placebo-controlled, double-blind study, 1,149 RA patients were randomized to receive 500 mg naproxen twice daily, 100 mg, 200 mg, or 400 mg celecoxib (PÞzer/Astellas’s Celebrex, a selective COX-2 inhibitor, discussed later in this section) twice daily, or placebo for 12 weeks (Simon LS, 1999). Patients were allowed to maintain stable doses of oral corticosteroids or DMARDs during the course of the trial. EfÞcacy measures included the percentage of patients achieving an ACR 20 response, a reduction of two or more grades from baseline on the Þve-point patient’s and physician’s global assessment scales, and mean improvements over baseline on the patients’ assessment of arthritis pain (100 mm VAS). Results for these four efÞcacy measures for the Þve treatment groups are reported in the following order: placebo, 500 mg naproxen twice daily, 100 mg celecoxib twice daily, 200 mg celecoxib twice daily, and 400 mg celecoxib twice

CO2H O FIGURE 9. Structure of naproxen.

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daily. Response rates using the ACR 20 criteria were 29%, 36%, 40%, 44%, and 39%, respectively. The percentage of patients demonstrating an improvement on the patient’s global assessment were 16%, 19%, 22%, 30%, and 25%, respectively. The percentage of patients demonstrating an improvement on the physician’s global assessment were 15%, 20%, 21%, 30%, and 25%, respectively. Lastly, the various treatment groups demonstrated improvements of 9.3 mm, 16.9 mm, 16.9 mm, 20.7 mm, and 18.1 mm, respectively, on the patient’s assessment of arthritis pain. The naproxen group’s ACR 20 response rate and improvement in arthritis pain were signiÞcantly superior to those of the placebo group. However, the naproxen group’s improvement on the patient’s and physician’s global assessments over placebo was not statistically signiÞcant. In this trial, adverse events were reported by 65% of patients treated with naproxen and 55% of placebo recipients. The most commonly reported adverse events for the naproxen group were GI tract disturbances (31% of patients), headache (16%), and upper respiratory tract infections (11%), compared with 19%, 23%, and 9% of placebo-treated patients, respectively. One of naproxen’s advantages over other traditional NSAIDs is its 12- to 15-hour plasma half-life, which allows for fewer daily doses. Naproxen is contraindicated in patients taking anticoagulants such as warfarin because naproxen prevents these anticoagulants from binding to plasma proteins, resulting in toxic blood levels of these agents. Recent controversy over the cardiovascular safety of naproxen was spurred by the suspension of the Alzheimer’s Disease Antiinßammatory Prevention Trial (ADAPT) due to concerns over the safety of celecoxib and preliminary Þndings that naproxen increased the risk of cardiovascular and cerebrovascular adverse events. However, the vast majority of data show that there is no relationship between the use of naproxen and an increased risk of myocardial infarction or stroke, although recent data suggest that naproxen may not have the cardioprotective effect it was previously thought to have (Graham DJ, 2005). Meloxicam. Meloxicam (Boehringer Ingelheim/Abbott/Daiichi’s Mobic) has been available in Europe since 1996. It was approved and launched in mid 2000 in the United States for osteoarthritis (OA), where it is co-promoted by Boehringer Ingelheim and Abbott. In July 2004, the FDA expanded meloxicam’s label by approving the drug to treat signs and symptoms of RA. The drug launched in Japan in February 2001, and was comarketed by Nippon Boehringer Ingelheim and Daiichi Pharmaceutical. However, Daiichi took over full Japanese marketing rights for meloxicam in May 2004. Meloxicam lost European patent protection at the end of 2003 and U.S. patent protection in 2005; the Japanese patent for meloxicam has already expired. Meloxicam has the strongest preference for inhibition of COX-2 of the traditional NSAIDs. The drug preferentially inhibits COX-2 over COX-1, so the inhibition of COX-2-stimulated proinßammatory prostaglandin production is greater than the inhibition of the production of COX-1-stimulated prostaglandins, which regulate renal function, blood ßow, platelet activity, and the protection of the

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mucous membrane along the GI tract. As a result, meloxicam was the Þrst NSAID to be marketed speciÞcally as a COX-2 inhibitor. Since then, debate has ensued over whether meloxicam is, in fact, a selective COX-2 inhibitor. The development of truly selective COX-2 agents (discussed later in “Selective COX2 Inhibitors”) has squelched that claim for meloxicam, and it is now referred to as a preferential COX-2 inhibitor. The efÞcacy of meloxicam (7.5 mg or 15 mg daily) has been demonstrated in double-blind trials of more than 5,000 RA and OA patients to be signiÞcantly greater than placebo and comparable to other traditional NSAIDs, such as diclofenac (100 mg daily slow release) and naproxen (750–1000 mg daily) (Barner A, 1996). One double-blind, randomized, placebo-controlled trial of 468 RA patients found that 15 mg meloxicam administered for 21 days was signiÞcantly superior to placebo in three of the four primary end points: patient’s and physician’s assessments of disease activity and reduction in the number of tender joints (Lemmel EM, 1997). A lower dose of meloxicam (7.5 mg) used in this study was also signiÞcantly better than placebo in the patient’s assessment of disease activity and reduction in the number of tender joints. In a 12-week, randomized, double-blind, double-dummy, parallel-group study of 894 RA patients, meloxicam (7.5 mg and 22.5 mg) was signiÞcantly superior to placebo in all Þve primary efÞcacy end points (patient’s and physician’s global assessments, patient’s assessment of pain, tender joint count, and swollen joint count) (Furst DE, 2002). In the same trial, 75 mg diclofenac administered twice daily signiÞcantly improved four of the primary end points over placebo, excluding swollen joint count. A six-month, double-blind, parallel-group study of 700 RA patients that compared 7.5 mg meloxicam with 750 mg naproxen showed no signiÞcant difference between the two treatment groups in the four primary efÞcacy end points (patient’s and physician’s global assessments, tender joint count, and swollen joint count) and in eight of the ten secondary end points (Wojtulewski JA, 1996). There is also evidence that meloxicam produces signiÞcantly fewer GI side effects than other traditional NSAIDs (Barner A, 1996). Meloxicam had fewer GI adverse events than naproxen (30.3% versus 44.7%) in the six-month, doubleblind study of 700 RA patients discussed previously (Wojtulewski JA, 1996). Furthermore, signiÞcantly more patients discontinued treatment because of GI adverse events in the naproxen group as compared with the meloxicam group. However, the 12-week, placebo-controlled, dose-response study of 894 RA patients comparing meloxicam (7.5–22.5 mg daily) and diclofenac (75 mg twice daily) found that the rate of GI events for all of the treatment groups did not differ signiÞcantly from each other or from placebo (Furst DE, 2002). Researchers and physicians were hopeful that these results validated the theory that preferential COX-2 inhibitors were safer than other traditional NSAIDs. However, GI disturbances, including dyspepsia, nausea and vomiting, abdominal pain, and diarrhea, remain common side effects of meloxicam treatment (Hawkey C, 1998; Dequeker J, 1998). In addition, the U.K. Medicines Control Agency and the Committee on Safety of Medicines received 733 reports of 1,339 suspected

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adverse drug reactions during postmarketing surveillance of meloxicam’s Þrst two years on the market in the United Kingdom. Gastric perforation, ulcers, or bleeding accounted for 18% of the reports, and Þve patients died as a result of GI side effects. In response to these adverse drug reactions, packaging for meloxicam was updated to include warnings about potentially serious GI and dermatologic side effects. Still, current concerns regarding the cardiovascular safety of selective COX-2 inhibitors (discussed in the next section) have boosted meloxicam’s popularity as physicians seek safer therapies to replace selective COX-2 agents. Selective Cyclooxgenase-2 Inhibitors Overview. As a result of the GI side effects that occur with traditional NSAIDs, considerable effort has been expended in developing new agents with less GI toxicity. In 1991, researchers achieved a major breakthrough when they discovered that two distinct isoforms of COX exist. COX-1, or constitutive cyclooxygenase, is present in cells under normal physiological conditions and stimulates the synthesis of prostaglandins that help regulate renal function, blood ßow, and platelet activity and help protect the mucous membrane along the GI tract. COX-2, or inducible cyclooxygenase, is expressed only under pathological conditions. Its production is induced by proinßammatory cytokines, mitogens, or endotoxins, and it stimulates the production of prostaglandins that drive the inßammatory process. Researchers believe that inhibition of COX-2 by NSAIDs is responsible for their analgesic and anti-inßammatory properties, while inhibition of COX-1 causes GI side effects by decreasing the production of gastroprotective prostaglandins. Unlike traditional NSAIDs, agents in this class inhibit COX-2 while only minimally affecting COX-1’s beneÞcial effects. Figure 10 illustrates the structural differences between COX-1 and COX-2 and how nonspeciÞc and speciÞc inhibition of these enzymes occurs. Drugs targeting COX-2, such as meloxicam and etodolac (Wyeth’s Lodine, generics), have been available since the mid 1990s. However, these agents are no longer considered to be true COX-2 inhibitors because they are only modestly selective for the inducible COX isoform. Thus, such agents are now considered to be preferential COX-2 inhibitors. The Þrst two agents to be considered truly selective COX-2 inhibitors, celecoxib (PÞzer/Astellas’s Celebrex) and rofecoxib (Merck’s Vioxx), were launched in 1999. These Þrst-generation selective COX-2 inhibitors demonstrated their ability to reduce GI side effects compared with traditional NSAIDs, but still cause some degree of GI upset. Thus, research and development efforts have focused on developing second-generation COX-2 inhibitors with even higher degrees of selectivity for COX-2. In 2002, two second-generation selective COX-2 inhibitors, valdecoxib (PÞzer/Astellas’s Bextra) and etoricoxib (Merck’s Arcoxia), entered the market. All these agents are indicated for treatment of RA pain. Selective COX-2 inhibitors boast a GI side-effect proÞle that is safer than that of traditional NSAIDs, but some experts have expressed concern over an

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FIGURE 10. Effect of nonsteroidal anti-inflammatory drugs and corticosteroids on the arachidonic acid pathway.

increased risk of cardiovascular and cerebrovascular adverse events associated with the selective agents. This concern stems from their potential to diminish vascular prostacyclin (PGI2 ) production; PGI2 acts as a vasodilator and inhibits aggregation of platelets (Mukherjee D, 2001). It was argued that by inhibiting the production of PGI2 , selective COX-2 inhibitors may alter the natural balance between prothrombotic thromboxane A2 and antithrombotic PGI2 . This altered balance could lead to an increase in thrombotic vascular events (Mukherjee D, 2001).

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Data showing that rofecoxib increases the risk of cardiovascular and cerebrovascular adverse events led to Merck’s voluntary worldwide withdrawal of this agent in September 2004. Upon rofecoxib’s launch in the United States in 1999, drug safety data did not demonstrate an increased risk of heart attack or stroke, although data from six-week studies suggested that thromboembolic events may be more frequent in patients receiving rofecoxib as compared with placebo (Villalba ML, 1998). Concerns over the cardiovascular safety of rofecoxib heightened with the results of the Vioxx GI Outcomes Research (VIGOR) clinical trial of 8,076 RA patients, which was designed to examine whether rofecoxib (50 mg daily) had fewer GI events than naproxen (500 mg twice daily), a traditional NSAID. Both drugs showed similar efÞcacy in this study, but rofecoxib had signiÞcantly fewer GI events and signiÞcantly more myocardial infarctions than naproxen (Bombardier C, 2000). The issue came to a head in August 2001, when a meta-analysis of more than 18,000 patients in four clinical trials, including VIGOR, concluded that the risk of cardiovascular events in patients treated with rofecoxib was signiÞcantly higher than placebo-treated patients (Mukherjee D, 2001). Merck responded later that year with its own meta-analysis of more than 28,000 patients from 23 rofecoxib clinical trials ranging from Phase IIb to Phase V showing that the relative risk of cardiovascular events is similar with rofecoxib and placebo (Konstam MA, 2001). As a result of these studies, the FDA revised the rofecoxib label in April 2002 to include language cautioning that the agent poses higher cardiovascular risks than a traditional NSAID but no greater risk than placebo. Most recently, interim data demonstrated a doubling in the risk of myocardial infarction and cerebrovascular accident for rofecoxib versus placebo at 18 months, leading to the suspension of the Adenomatous Polyp Prevention on Vioxx (APPROVe) trial and prompting Merck to withdraw the drug (Merck, press release, September 30, 2004; Bresalier RS, 2005). The withdrawal of rofecoxib has focused attention on the cardiovascular and cerebrovascular safety associated with the remaining selective COX-2 inhibitors. The FDA issued a public health advisory in December 2004 recommending limited use of COX-2 inhibitors pending further review of the accumulating data suggesting an increased cardiovascular risk with selective COX-2 inhibitor use. The participants in an FDA joint committee meeting on COX-2 inhibitor safety in February 2005 suggested that the increased risk of cardiovascular and cerebrovascular events associated with rofecoxib is a class-wide effect. However, current data suggest that the level of risk differs for each agent and that rofecoxib has the highest risk of all selective COX-2 inhibitors (Graham DJ, 2005). In the FDA meeting of the Arthritis Advisory Committee and the Drug Safety and Risk Management Advisory Committee in February 2005, all committee members agreed that available data support the conclusion that rofecoxib, celecoxib, and valdecoxib signiÞcantly increase the risk of cardiovascular events. However, a majority of the committee members concluded that the overall risk versus beneÞt proÞles for all three agents supports their marketing in the United States. Celecoxib received the most votes for a favorable risk-beneÞt assessment supporting continued U.S. marketing, 31 to 1, as compared with 17 to 13 for

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valdecoxib (2 members abstained) and 17 to 15 for rofecoxib. The FDA committee also suggested a black box warning be added to selective COX-2 inhibitor package inserts and that DTC advertising should be curtailed. The EMEA issued a contraindication in February 2005 for the use of selective COX-2 inhibitors in patients with ischemic heart disease or risk of stroke. Shortly after the FDA’s advisory committees ruled in favor of valdecoxib remaining on the market, the FDA took action itself in April 2005 by asking PÞzer to withdraw the agent from the market because of its unfavorable cost/beneÞt proÞle; the FDA’s decision reßects valdecoxib’s association with an increased risk of serious skin reactions in addition to CV and cerebrovascular events. The EMEA followed suit and also asked PÞzer to remove the drug from all European markets. As a result, PÞzer withdrew valdecoxib from the worldwide market. Celecoxib is currently the only selective COX-2 inhibitor on the U.S. market. Although Merck might decide to revive rofecoxib, the company is more likely to keep it off the market because of the drug’s negative image. Instead, Merck will likely focus its attention on its newer selective COX-2 agent, etoricoxib. Although the future of individual COX-2 therapies is somewhat murky, what is clear is that prescriptions for all agents in this class, regardless of the Þnal FDA decision, will be severely restricted to patients who cannot tolerate the GI side effects associated with traditional NSAID use and who are not at risk for cardiovascular problems. Mechanism of Action. Selective COX-2 inhibitors predominantly inhibit COX-2 while minimizing inhibition of COX-1 (see Figures 7 and 10). These agents block the activity of COX-2, which produces prostaglandins that drive the inßammatory process, without disrupting the beneÞcial effects of COX-1 prostaglandins in the GI tract. Celecoxib. Celecoxib (PÞzer/Astellas’s Celebrex) (Figure 11) received FDA approval for the treatment of OA and RA in December 1998 and was launched in Europe in 2000. PÞzer markets this agent in all the countries under study except Japan, where Astellas Pharma (formerly Yamanouchi Pharmaceuticals, which merged with Fujisawa Pharmaceuticals on April 1, 2005, to form Astellas Pharma) is guiding it through clinical development. A new drug application (NDA) for pain and other symptoms of RA and OA was submitted in Japan in December 2002, with approval expected in 2004, but the application was still under consideration at the time of the writing of this report. Celecoxib gained FDA approval for the management of acute pain and primary dysmenorrhea in October 2001, and it is approved in the United States and Europe as an adjunct treatment for familial adenomatous polyposis (FAP). These additions give celecoxib the broadest range of approved indications of any selective COX-2 inhibitor. The agent is also in development for a range of further indications such as pancreatic and prostate cancers. Patent coverage extends to the end of 2013 in the United States and to the end of 2014 in other countries under study. Celecoxib works by the same mechanism of action as all selective COX-2 inhibitors, as discussed in the “Mechanism of Action” section for the drug class.

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FIGURE 11. Structure of celecoxib (R1 = NH2 , R2 = CH3 ).

The numerous trials of celecoxib have been designed primarily to show that its efÞcacy in OA and RA is comparable to that of other NSAIDs and to highlight its improved GI safety. In three large clinical trials that cumulatively involved more than 2,000 patients, celecoxib at doses of 100, 200, and 400 mg twice daily was found to have signiÞcantly better efÞcacy than placebo and similar efÞcacy to traditional NSAIDs—namely, naproxen 500 mg twice daily or diclofenac slow release 75 mg twice daily (Emery P, 1999; Goldstein JL, 2001; Simon LS, 1999). These studies also showed a lower incidence of upper GI events as compared with traditional NSAIDs, suggesting that celecoxib has decreased GI toxicity. PÞzer designed the Celecoxib Long-Term Arthritis Safety Study (CLASS) in hopes of providing statistically signiÞcant evidence that celecoxib results in fewer GI adverse events than traditional NSAIDs, much as the VIGOR trial did for rofecoxib. The double-blind, randomized, one-year CLASS trial assigned 8,059 OA and RA patients to receive celecoxib 800 mg, ibuprofen 2,400 mg, or diclofenac 150 mg daily. The celecoxib dose used was four times that recommended for OA and twice the recommended highest RA dose. Patients were allowed to take concomitant aspirin for cardiovascular prophylaxis. The main outcome measures were the incidence of symptomatic upper GI ulcers and ulcer complications. Preliminary analysis at six months suggested that the celecoxib-treated group had signiÞcantly fewer upper GI incidents than the NSAID-treated groups (Silverstein FE, 2000). Based on these results, PÞzer petitioned the FDA for a revision of celecoxib’s labeling. However, the full data at one year showed that celecoxib did not differ from traditional NSAIDs in its effect on the predeÞned GI end points, although there may have been fewer GI events in celecoxib-treated patients who were not taking aspirin (Fitzgerald GA, 2003). The FDA concluded that CLASS did not show a safety advantage in upper GI events for celecoxib as compared with traditional NSAIDs and that the celecoxib label should continue to have the standard warning associated with all NSAIDs, including risks of GI ulceration, bleeding, and perforation. As mentioned in the overview of selective COX-2 inhibitors, there are concerns that celecoxib may be associated with an increased risk of cardiovascular adverse events. A meta-analysis of more than 18,000 patients in four clinical trials, including CLASS, concluded that the risk of cardiovascular events in patients treated with celecoxib was signiÞcantly higher than placebo-treated

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patients (Mukherjee D, 2001). In December 2004, the Data Safety Monitoring Board for the Prevention of Sporadic Colorectal Adenomas with Celecoxib (APC) trial recommended suspension of the trial based on interim data at 33 months indicating an increased risk of serious cardiovascular events, including myocardial infarction, stroke, and death, in patients treated with celecoxib (200 or 400 mg twice daily) as compared with placebo-treated patients. The frequency of adverse cardiovascular events in this trial was as follows: 7 of 679 patients in the placebo group, 16 of 685 patients in the 200 mg celecoxib group, and 23 of 671 patients in the 400 mg celecoxib group (corresponding to hazard ratios of 1.0, 2.3, and 3.4, respectively), suggesting that celecoxib use was associated with a dose-related increase in cardiovascular risk (Solomon SD, 2005). In response to the APC data, the Prevention of Colorectal Sporadic Adenomatous Polyps Trial (PreSAP) and ADAPT were also suspended despite the lack of an increased cardiovascular risk for celecoxib-treated patients over placebo in these trials. However, most epidemiological studies and clinical trials with up to one-year duration show no increased cardiovascular risk for celecoxib over placebo or traditional NSAIDs. Although celecoxib may increase the risk of adverse cardiovascular events, this risk is signiÞcantly lower than that of rofecoxib (Graham DJ, 2005). Pending further analysis of the trial data and the dosages employed, the FDA has advised physicians to evaluate the risk-beneÞt ratio of celecoxib in individual patients and, if continued use is considered appropriate, advises the use of the lowest effective dose for the shortest possible time (FDA Alert, December 2004). Elsewhere, regulatory authorities have recommended that celecoxib be reserved for patients at risk for GI complications and that it not be used in patients with established heart disease or those at high risk of cardiovascular or cerebrovascular events. Valdecoxib. In April 2002, PÞzer launched the Þrst second-generation selective COX-2 inhibitor, valdecoxib (Bextra) (Figure 12), in the United States for the treatment of OA, RA, and dysmenorrhea. In May 2003, valdecoxib received approval in Europe for treatment of the pain and inßammation associated with OA, RA, and primary dysmenorrhea and has since launched in the United Kingdom, Germany, and Italy. As of this writing, sales of valdecoxib remain suspended. In vitro studies by PÞzer showed valdecoxib to be the most potent and selective of the marketed COX-2 inhibitors, providing a basis for the observed potent

FIGURE 12. Structure of valdecoxib.

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analgesic and anti-inßammatory activity of the agent in humans (Gierse JK, 2004). Valdecoxib works by the same mechanism of action as all selective COX-2 inhibitors, as discussed in the “Mechanism of Action” section for the drug class. Valdecoxib has demonstrated efÞcacy similar to that of traditional NSAIDs and signiÞcantly superior to that of placebo in numerous clinical trials. In a 12-week double-blind study of 1,090 RA patients, study participants were randomized to receive 10, 20, or 40 mg valdecoxib once daily, naproxen 500 mg twice daily, or placebo. All three doses of valdecoxib provided similar relief of the signs and symptoms of RA superior to placebo and similar to naproxen as assessed by ACR 20 response (Bensen W, 2002). In another study of 726 adult-onset RA patients, valdecoxib (20 and 40 mg daily) was compared with diclofenac (75 mg slow release twice daily) at 26 weeks (Pavelka K, 2003). EfÞcacy was measured by the patient’s assessment of arthritis pain and the modiÞed HAQ. Both doses of valdecoxib were comparable to diclofenac in efÞcacy, with no signiÞcant differences observed between treatment groups. Further evidence of the efÞcacy of valdecoxib came from the results of a meta-analysis of nine studies involving a total of 5,726 RA and OA patients. Valdecoxib (10 or 20 mg) was found to have efÞcacy superior to placebo and equivalent to the maximum daily doses of traditional NSAIDs (Edwards JE, 2004). Like celecoxib, valdecoxib has some data supporting a lower incidence of GI adverse events, but deÞnitive evidence from a large-scale clinical trial is lacking. In the 12-week study of 1,090 RA patients discussed previously, the incidence of gastroduodenal ulcers was 6% in patients receiving valdecoxib 20 mg and 4% for valdecoxib 40 mg, which were both signiÞcantly lower than diclofenac-treated patients (16%) (Pavelka K, 2003). There were also signiÞcantly fewer discontinuations due to gastrointestinal adverse events (4% versus 8%) or endoscopic ulcers of 3 mm or more (5% versus 13%) with valdecoxib compared with traditional NSAIDs in the meta-analysis of 5,726 RA and OA patients (Edwards JE, 2004). However, the study authors cautioned that convincing evidence of reduced major gastrointestinal adverse events could not be addressed by the trials. In November 2002, valdecoxib’s label was updated with new warnings following postmarketing reports of serious adverse events, including toxic epidermal necrolysis, Stevens-Johnson syndrome, erythema multiforme, and anaphylactoid reactions. In December 2004, the previous warnings about the risk of lifethreatening skin reactions were strengthened with a boxed warning added to the product label advising that treatment be stopped at the Þrst appearance of skin rash, mucosal lesions, or any other sign of hypersensitivity. Like celecoxib, valdecoxib is contraindicated in patients with an allergy to sulfa-containing products. In Europe, the EMEA has issued a public statement warning that treatment with valdecoxib may result in serious skin reactions. In November 2004, following lay press criticism of the cardiovascular safety of valdecoxib, PÞzer responded by citing a peer-reviewed meta-analysis of data from 7,934 patients. In this study, the incidence of cardiovascular thrombotic events was determined by analyzing pooled valdecoxib (10–80 mg daily), nonselective NSAID (diclofenac 75 mg twice daily, ibuprofen 800 mg three times daily, or

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naproxen 500 mg twice daily), and placebo data from ten randomized OA and RA trials of 6–52 weeks duration. Short- and intermediate-term treatment with valdecoxib doses was not associated with an increased incidence of thrombotic events relative to nonselective NSAIDs or placebo in OA and RA patients in controlled clinical trials (White WB, 2004). However, two clinical trials that examined the potential use of valdecoxib for pain immediately following coronary artery bypass graft (CABG) surgery demonstrated an increased risk of adverse cardiovascular events in this high-risk population (Ott E, 2003; Nussmeier NA, 2005). The second of these studies involved 1,671 patients randomly assigned to receive intravenous parecoxib (PÞzer’s Dynastat) for at least three days followed by oral valdecoxib (20 mg every 12 hours) through day 10, intravenous placebo followed by oral valdecoxib, or placebo for ten days. (Parecoxib, approved for the treatment of acute pain in Europe, is the injectable, water-soluble pro-drug that is metabolically converted to valdecoxib in vivo.) Cardiovascular events, including myocardial infarction, cardiac arrest, stroke, and pulmonary embolism, were signiÞcantly more frequent in the group treated with parecoxib and valdecoxib as compared with placebo (2.0% versus 0.5%). The group treated with intravenous placebo followed by valdecoxib had more cardiovascular events than placebo (1.1%), but was not statistically different from the other two groups (Nussmeier NA, 2005). As a result of these studies, labeling in the United States and Europe was revised in December 2004 contraindicating valdecoxib in patients undergoing CABG surgery, and the EMEA issued a public statement regarding the drug’s cardiovascular risks. Although some studies suggest that valdecoxib increases the likelihood of cardiovascular events by as much as two- to threefold (Furberg CD, 2005), there are limited published data, and further studies are needed to evaluate the cardiovascular risk. Etoricoxib. Merck’s second-generation COX-2 inhibitor, etoricoxib (Arcoxia) (Figure 13), was approved in the United Kingdom in April 2002 for the treatment of symptomatic pain relief in OA, RA, acute gouty arthritis, chronic musculoskeletal pain, acute pain associated with dental surgery, and primary dysmenorrhea. By October 2002, etoricoxib had been approved in Europe under the Mutual Recognition Procedure, and it has since launched in Italy and Spain. The U.K. launch came on the heels of the company’s withdrawal of its initial NDA for approval in the United States, following the FDA’s request for additional data on etoricoxib’s cardiovascular safety. In December 2003, Merck submitted an expanded NDA, seeking indications for the treatment of OA, RA, chronic low back pain, acute pain, dysmenorrhea, acute gouty arthritis, and ankylosing spondylitis. In October 2004, the FDA issued an approvable letter. However, it informed Merck that before approval could be issued, additional safety and efÞcacy data for the drug are required. Patent coverage in all the markets under review extends to 2017. Etoricoxib works by the same mechanism of action as all selective COX-2 inhibitors, as discussed in the “Mechanism of Action” section for the drug class.

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FIGURE 13. Structure of etoricoxib.

Etoricoxib has demonstrated efÞcacy signiÞcantly superior to that of placebo and at least comparable to that of naproxen in reducing the signs and symptoms of RA. In a double-blind study involving 816 RA patients, the efÞcacy of etoricoxib was shown to be signiÞcantly better than that of either placebo or naproxen (Matsumoto AK, 2002). Participants were randomized to receive either 90 mg etoricoxib once daily, 500 mg naproxen twice daily, or placebo for 12 weeks. Patients were allowed to take concomitant oral corticosteroids or DMARDs during the trial. Primary efÞcacy measures in this study included improvements over baseline on the 100 mm patient global assessments of disease activity VAS, a four-point investigator global assessment of disease activity scale, the total number of tender joints, and the total number of swollen joints. The etoricoxib group demonstrated a mean 17 mm improvement on the patient’s global assessment of disease activity VAS, a mean 0.63 point improvement on the investigator global assessment of disease activity scale, a mean 6.3 joint reduction in the number of tender joints, and a mean 3.3 joint reduction in the number of swollen joints. Each of these improvements for the etoricoxib group was statistically signiÞcant compared with the placebo and naproxen groups. The percentage of patients who completed the trial and met the ACR 20 response criteria was 21%, 39%, and 53% for the placebo, naproxen, and etoricoxib groups, respectively. An identically designed study using the same primary and secondary end points in 891 RA patients found similar significant improvements over baseline scores for the etoricoxib group compared with placebo recipients (Collantes E, 2002). However, there was no significant difference in improvements between the etoricoxib and naproxen treatment groups. Etoricoxib has a low risk of GI adverse events owing to its higher selectivity for COX-2 than the Þrst-generation COX-2 inhibitors or traditional NSAIDs. In the study discussed in the previous paragraph, the incidence of drug-related adverse events did not differ signiÞcantly between treatment groups, though the incidence of GI-related adverse events (i.e., dyspepsia, heartburn, nausea) was slightly higher for the naproxen group compared with the placebo and etoricoxib groups (Matsumoto AK, 2002). A combined analysis of ten endoscopy

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trials of patients with OA, RA, or ankylosing spondylitis presented at the Annual European Congress of Rheumatology in June 2004 showed that treatment with etoricoxib was associated with 52% fewer conÞrmed upper GI perforations, ulcers, and bleeds compared with treatment with nonselective, traditional NSAIDs (Ramey DR, 2004). Further support for the improved GI tolerability of etoricoxib was supplied by the Etoricoxib Diclofenac Gastrointestinal Evaluation (EDGE) study, a double-blind trial in 7,111 OA patients treated for up to 16.5 months (Baraf HSB, 2004). Patients were randomized to receive etoricoxib 90 mg daily (n = 3, 593) or diclofenac 50 mg three times daily (n = 3, 518). At study baseline in both treatment groups, 4% of patients had a history of an upper GI event and 28% were taking low-dose aspirin. The discontinuation rate due to GI adverse events, the primary end point of this study, was signiÞcantly lower with etoricoxib than diclofenac, at 9.4 versus 19.2 events per 100 patient years, respectively, or a risk reduction of 50%. Although etoricoxib appears to be associated with fewer GI adverse events than traditional NSAIDs, other studies have shown that the GI safety proÞle is still signiÞcantly worse than placebo. For example, a 12-week, double-blind, randomized study in 680 patients found that the incidence of endoscopy-detected ulcers was 17% for ibuprofen (800 mg three times daily), 8% for etoricoxib (120 mg daily), and 2% for placebo, all of which were signiÞcantly different from each other (Hunt RH, 2003). Despite an improved GI safety proÞle, consideration of etoricoxib’s safety has been dominated by debate over potential cardiovascular complications that appear to affect the selective COX-2 inhibitors as a class. There has been little research on the cardiovascular safety of etoricoxib, but available data show no evidence for increased cardiovascular risk. There was no signiÞcant difference between etoricoxib and diclofenac in the incidence of overall, drug-related, serious adverse events in the EDGE trial (Baraf HSB, 2004). Furthermore, cardiovascular data from the study demonstrated that the rates of conÞrmed thrombotic cardiovascular events were similar for both etoricoxib and diclofenac. The relative risk of etoricoxib compared with diclofenac was 1.07 for events that occurred within 14 days of discontinuing study treatment and 1.02 for events that occurred within 28 days of discontinuing study treatment (a relative risk of 1.00 means equivalence between the two treatment groups). Fewer patients discontinued etoricoxib for any adverse event as compared with diclofenac, although signiÞcantly fewer patients taking diclofenac (0.7%) discontinued the study because of hypertensionrelated adverse events compared with patients taking etoricoxib (2.3%) (Merck, press release, October 2004). Two ongoing clinical trials may shed light on the debate over etoricoxib’s cardiovascular safety. The EDGE II trial is examining the GI tolerability of etoricoxib (90 mg daily) as compared with diclofenac (50 mg three times daily) in approximately 4,000 RA patients. The Multinational Etoricoxib Diclofenac Arthritis Long-Term (MEDAL) study was designed with cardiovascular safety as a primary end point, comparing diclofenac (50 mg three times daily) with etoricoxib (60 or 90 mg) in more than 23,000 RA and OA patients. Results from

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both trials are expected to become available in 2006. Until then, the EMEA advises limited use of all selective COX-2 inhibitors in patients with a risk of GI complications and without ischemic heart disease or a history of stroke. Corticosteroids Overview. Corticosteroids have long been employed in the treatment of RA and remain a key component of symptom management. They produce potent, rapid suppression of inßammation with consequent improvements in joint pain and swelling. Studies showing that corticosteroids can reduce the rate of joint damage in RA patients led to their initial classiÞcation as disease-modifying agents (Kirwan RJ, 1995; van Everdingen AA, 2002). However, rheumatologists caution that the disease-modifying properties of corticosteroids are limited, and they should not be administered as a monotherapy to RA patients with active disease. Instead, true DMARDs, either conventional or biological, should be initiated, and corticosteroids used as supplemental therapy to control symptoms as needed. Corticosteroids are associated with many side effects. Insomnia, night sweats, mood changes, and altered glucose metabolism may occur shortly after beginning corticosteroids. Prolonged corticosteroid therapy can lead to adrenal atrophy, and abrupt cessation can cause adrenal insufÞciency, hypotension, and even death (Caldwell JR, 1991; Hunter JA, 1999). Long-term use of systemic corticosteroids is associated with osteoporosis, hypertension, cataracts, acne, abnormal fat deposition, and excessive hair growth. These side effects generally occur less often with corticosteroid injections, but injection site infections, post-injection ßares, and crystal-induced synovitis can occur with parenteral delivery. A wide variety of generic corticosteroids, salts, and formulations are available in the markets under study. This section assesses two of the most commonly prescribed agents, orally administered prednisone (PÞzer’s Deltasone, generics) and intra-articular (IA) methylprednisolone acetate (PÞzer’s Depo-Medrol, generics). Mechanism of Action. Corticosteroids act as anti-inßammatory and immunosuppressive agents through multiple effects: inhibiting synthesis of proinßammatory mediators (prostaglandins, leukotrienes, and cytokines); disrupting cellular activation, migration, and proliferation; and blocking edema formation. Corticosteroids can also inhibit the expression of COX-2 and the activity of collagenase and other cartilage-damaging enzymes. Figure 7 illustrates how corticosteroids suppress prostaglandin and leukotriene synthesis by blocking the arachidonic acid pathway. Formulation. Rheumatologists employ a wide variety of corticosteroid formulations and doses to relieve RA symptoms. Oral corticosteroids are often used as bridge therapy, meaning they are initiated by primary care physicians or rheumatologists upon diagnosis, but then discontinued when the slower-acting, conventional DMARD therapy takes effect. Intravenous corticosteroids are occasionally used as bridge therapy, as well. In addition, oral and intravenous corticosteroids

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are used during periodic ßare-ups to control symptoms during times of high disease activity. For alleviating severe RA symptoms, high doses of intravenous corticosteroids can be used on a short-term basis (up to 1 g per day for three days). The intravenous treatment can be repeated after four to six weeks if necessary. Alternatively, oral corticosteroids can be initiated at a high dose (e.g., 30–40 mg prednisone daily) and then gradually tapered down in 5 mg steps to a continuous dosage of 5–10 mg per day. For milder symptoms, a low dose of oral corticosteroids can be used as short-term or long-term therapy. Because of the serious side effects associated with corticosteroids, rheumatologists prefer to discontinue these drugs as soon as possible. However, abrupt cessation of corticosteroids can cause adrenal insufÞciency, hypotension, and even death, so doses must be carefully tapered to a level where these drugs may be safely discontinued. To minimize side effects and to prevent problems with abrupt discontinuation of systemic therapy, intramuscular or IA injections are used for relief of localized, aggressive joint ßare-ups. A single intramuscular dose of a depot corticosteroid, such as methylprednisolone acetate or triamcinilone acetonide (both 40 mg/mL), is preferred by rheumatologists when rapid control is required. Furthermore, physicians and researchers have expressed concern that repeated corticosteroid injections into joints can actually exacerbate progressive cartilage damage. As a result, many rheumatologists consider this strategy a last resort and will not use it in more than one joint at a time. The ACR also recommends that intra-articular injections not be administered in the same joint more than once within three months (ACR, 2002). Prednisone. Oral prednisone (PÞzer’s Deltasone, generics) (Figure 14) is commonly prescribed in low doses (less than 10 mg daily) to treat the signs and symptoms of RA. Oral corticosteroids—including prednisone—can be used as a short-term therapy during periods of high disease activity or as a bridge therapy and can be administered for long-term use in combination with DMARDs. Corticosteroids act as anti-inßammatory and immunosuppressive agents through multiple effects: inhibiting synthesis of proinßammatory mediators (prostaglandins, leukotrienes, and cytokines); disrupting cellular activation, migration, and proliferation; and blocking edema formation. Corticosteroids can CH2OH C O

H3C

O OH

H3C

O FIGURE 14. Structure of prednisone.

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also inhibit the expression of COX-2 and block the activity of collagenase and other cartilage-damaging enzymes. Low-dose oral prednisone (7.5 mg) has been shown to provide a rapid and signiÞcant decrease in RA symptoms between two and eight weeks after treatment initiation, an effect that was maintained for three months and that was not associatedwith rebound in symptoms after treatment cessation (Kirwan JR, 2004). In a prior study, the efÞcacy, disease-modifying properties, and safety of lowdose oral prednisone were compared with those of placebo (van Everdingen AA, 2002). This two-year, randomized, double-blind study involved 81 patients with early active RA who had not been treated previously with DMARDs. Trial participants received either 10 mg daily oral prednisone or placebo, and both treatment groups were given 500 mg daily of elementary calcium. Patients were allowed to take acetaminophen or NSAIDs for pain, if necessary, and sulfasalazine (2 g daily) could be added as a rescue medication after six months if the activity of RA warranted the addition of a DMARD. At the conclusion of the two-year trial, treatment with prednisone was associated with a greater mean improvement over baseline scores for most efÞcacy measures, including early-morning stiffness, morning pain, general well-being, and 28-joint score for swelling. There were signiÞcant differences between the improvements for the two groups on two efÞcacy measures: grip strength (measured by a vigorimeter) and the 28-joint score for tenderness. A clinically relevant improvement (deÞned as 20% or greater improvement in the 28-joint scores for swelling and tenderness, and 20% or greater improvement in two of the following four variables: pain, general well-being, HAQ, and CRP level) was demonstrated by 33% and 30% of patients receiving prednisone at 12 and 24 months and by 24% and 22% of placebo recipients at these two time periods, respectively. Treatment with prednisone was associated with signiÞcant increases in mean body weight and mean serum glucose level compared with placebo. Additionally, the incidence of new vertebral fractures was higher in the prednisone group compared with the group treated with placebo. Perhaps the most interesting Þnding from the study is the ability of prednisone to reduce progression of radiologically detected joint damage over the course of two years (van Everdingen AA, 2002). At the start of the trial, radiologic evidence demonstrated that 40% of patients randomized to prednisone and 37% of patients randomized to placebo had erosive disease. After 24 months, the percentage of patients with erosive disease was 70% for the prednisone group and 78% for the placebo group. Total radiologic scores (sum of erosion and joint-space narrowing scores using the van der Heijde modiÞcation of the Sharp method for scoring radiographs) were signiÞcantly lower for the prednisone group compared with placebo at 12, 18, and 24 months. The prednisone group demonstrated a mean 8-point and 16-point increase from baseline in total radiologic score at 12 and 24 months, respectively, compared with mean 15-point and 29-point increases from baseline for the placebo group at these two time periods, respectively. However, corticosteroids are not considered true DMARDs because their disease-modifying properties are far inferior to those of conventional and biological DMARDs. For

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OR O HO

OH

O

FIGURE 15. Structure of methylprednisolone acetate (R = COCH3 ).

example, the combination of etanercept with methotrexate has achieved a 69% ACR 50 response in reducing the signs and symptoms of RA and a negative Sharp score, indicating no progression of joint damage at 52 weeks (Klareskog L, 2004). Methylprednisolone Acetate. Methylprednisolone acetate (PÞzer’s DepoMedrol, generics) (Figure 15) is the 6-methyl derivative of the corticosteroid prednisolone. Methylprednisolone acetate is indicated for a number of arthritic disorders as well as other inßammatory conditions such as asthma. Similar to other corticosteroids, methylprednisolone acetate inhibits the synthesis of proinßammatory mediators (prostaglandins, leukotrienes, and cytokines), the expression of COX-2, and the activity of collagenase and other cartilagedamaging enzymes. Because of the lack of large-scale, placebo-controlled trials comparing methylprednisolone acetate with placebo in RA patients, this section presents the Þndings of a blinded study involving 18 knees of 16 patients with RA or other arthritic conditions (e.g., psoriatic arthritis, juvenile rheumatoid arthritis) (Ostergaard M, 1996). In this study, patients underwent arthrocentesis to remove excess ßuid from their knee(s), after which 30 mg lidocaine hydrochloride and 80 mg methylprednisolone acetate were injected into these knees. Gadolinium-DTPA (diethylenetriamine penta-acetic acid) enhanced magnetic resonance imaging (MRI) was performed on each patient prior to arthrocentesis (day 0) and after treatment on days 1, 7, 30, 180, and until clinical relapse (deÞned by the presence of joint swelling and/or tenderness) to measure synovial membrane volume and joint effusion volume. On day 0, the mean synovial membrane volume and joint effusion volume of patients was 58 mL and 35 mL, respectively. There was no signiÞcant difference between the knees of patients with RA and other arthritic conditions. Compared with baseline measurements, mean synovial membrane volume decreased by 43%, 50%, and 64% on days 1, 7, and 30, while joint effusion volumes decreased 57%, 63%, and nearly 69% at the same time points. Each of these improvements was statistically signiÞcant compared with baseline measurements. On day 180, only 5 knees of the original 18 remained in clinical remission. Safety data were not reported in the study conducted by Ostergaard and colleagues. Risk of infection is associated with the use of IA corticosteroids, but this

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complication is rare if proper aseptic techniques are followed (Gifford RH, 1973). Additionally, IA corticosteroids can cause crystal-induced synovitis, which can cause a temporary increase in joint swelling and pain following administration of the drug. Concern that repeated corticosteroid injections into joints can actually exacerbate progressive cartilage damage has made this strategy a last resort in the treatment of RA symptoms. It is recommended that intra-articular corticosteroid injections not be used in more than one joint at a time and not more than every three months in the same joint (ACR, 2002). Nonpharmacological Therapies Prosorba Column. Developed by Cypress Bioscience as a novel blood-Þltering device, the Prosorba column was licensed for comarketing in the United States, Europe, and Japan to Fresenius Hemotechnology. The column received FDA approval in 1999 for patients unresponsive to or intolerant of DMARD therapy, and it was launched in the United States and Europe in 2000. This device uses approximately 200 mg of protein A (a component of the Staphylococcus bacterium that selectively binds IgG and IgG-bound antigens) covalently bound to an inert silica matrix. In a process similar to kidney dialysis, a patient’s blood is removed from a vein in one arm and passed through a machine that separates the blood cells from the plasma. The plasma is then passed through the Prosorba column, where circulating immune complexes are removed, recombined with the blood cells, and returned to the patient through the other arm. The standard course of therapy involves 12 weekly, two-hour outpatient sessions. A 99-patient, double-blind trial resulted in a 41.7% ACR 20 improvement in Prosorba-treated patients versus 15.6% with placebo (Furst D, 2000). Surgery. Surgical treatment for RA takes three main forms: carpal tunnel relief and metatarsal head resections, synovectomies (partial removal of the diseased joint lining), and arthroplasty (total joint replacement). Carpal tunnel and metatarsal head procedures are rarely performed, but they are useful when corticosteroid injections have failed or the maximum dose provides no further improvement. Synovectomies are perhaps even rarer because the long-term beneÞts are outweighed by the procedure’s cost. Arthroplasty of the knee and hip are the most common type of surgery for RA. Approximately 15–25% of all RA patients eventually undergo this procedure for one or more joints. Most of these patients have end-stage disease, and many have secondary OA. EMERGING THERAPIES The launch of highly effective biological therapies in the late 1990s—in particular, the tumor necrosis factor-alpha (TNF-α) inhibitors—has dramatically improved the therapeutic options available for the treatment of rheumatoid arthritis (RA). The availability of these effective, albeit expensive, agents has signiÞcantly raised the bar for emerging therapies in the RA market. Despite the steep

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competition in developing therapies for this market, a vast number of agents, many of them biologics, are under development for the treatment of RA. This discussion focuses on emerging therapies that have mLed comparatively late-stage development for RA (i.e., Phase II or later trials, for which clinical results are available). Table 4 summarizes emerging therapies in late-stage development for RA. The discussion of emerging therapies is organized in this section according to their mechanisms of action. Tumor Necrosis Factor-α Inhibitors Overview. Inhibitors of TNF-α represent the most recent and signiÞcant advance in the treatment of RA. The Þrst anti-TNF-α agent approved for RA, etanercept (Amgen/Wyeth/Takeda’s Enbrel), was launched in the United States in 1998. Since then, two other agents have entered the market—inßiximab (Centocor [a Johnson&Johnson subsidiary]/Schering-Plough/Tanabe Seiyaku’s Remicade) and adalimumab (Abbott/Eisai’s Humira). Because of the relatively high cost and risk of immunomodulatory side effects associated with these agents, researchers have directed their attention toward the development of improved anti-TNF-α therapies for the treatment of RA. Different approaches to targeting and blocking TNF-α include monoclonal antibodies, TNF-α receptors, TNFbinding proteins, and antisense technology. Two new TNF-α inhibitors that have progressed into Phase II clinical trials or beyond are discussed here. Currently available products are administered parenterally, and various companies have sought to develop orally active agents as a means of reducing cost and broadening the market scope for anti-TNF-α therapy. Oral delivery of TNF-α inhibition is seen as a particularly desirable goal by many rheumatologists. In practice, however, the development of orally active agents has proven difÞcult. Work on a number of apparently promising oral agents, such as AtheroGenics’ AGIX-4207, Bristol-Myers Squibb’s BMS-561392, and Isis’s ISIS-104838, has been discontinued. The most promising oral TNF-α inhibitor remaining in the pipeline is apratastat (Wyeth’s TMI-005). This agent, however, is in early Phase II development and is not discussed because of a lack of clinical trial data. Mechanism of Action. The role of TNF-α in RA has been demonstrated in transgenic mouse models in which TNF-α overexpression led to joint destruction and aggressive synovitis (Figure 6). Among RA patients, the degree of synovial TNF-α expression correlates with the degree of synovitis and bone erosion. Further investigations have shown increased expression of TNF-α receptors in synovial tissues of patients with RA but not in some other forms of arthritis. Receptors for TNF-α are found on the surface of most cells, including mononuclear cells and cells in the synovium. Cleavage of membrane-bound TNF-α receptors yields soluble TNF-α receptors that retain ligand-binding ability but cannot activate cells. Two distinct types of TNF-α receptors have been identiÞed: type I (p55) and type II (p75). TNF-α inhibitors reduce free, bioactive TNF-α by emulating the physiological role played by soluble TNF-α receptors.

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TABLE 4. Emerging Therapies in Development for Rheumatoid Arthritis Compound

Development Phase

Marketing Company

TNF-α inhibitors CDP-870 United States Europe Japan

III III —

UCB UCB —

Pegsunercept United States Europe Japan

II — —

Amgen — —

III III III

Chugai/Roche Chugai/Roche Chugai/Roche

II — —

Amgen — —

PR — —

Bristol-Myers Squibb — —

III III —

Roche/Biogen Idec/Genentech Roche/Biogen Idec/Genentech —

Conventional DMARDs Iguratimod United States Europe Japan

IIa IIa PR

Toyoma Chemical/Eisai Toyoma Chemical/Eisai Toyoma Chemical/Eisai/Taisho

Selective COX-2 inhibitors Lumiracoxib United States Europe Japan

III PR II

Novartis Novartis Novartis

Immunosuppressants Tacrolimus United States Europe Japan

III II PR

Astellas Astellas Astellas

Interleukin-based therapies Atlizumab United States Europe Japan AMG-714 United States Europe Japan Lymphocyte modulators Abatacept United States Europe Japan Rituximab United States Europe Japan

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TABLE 4. (continued) Compound MAP kinase inhibitors SCIO-469 United States Europe Japan Doramapimod United States Europe Japan

Development Phase

Marketing Company

II II —

Scios Scios —

II II —

Boehringer Ingelheim Boehringer Ingelheim —

COX-2 = Cyclooxygenase-2; DMARDs = Disease-modifying antirheumatic drugs; IL = Interleukin; MAP = Mitogen-activated protein; PR = Preregistered; TNF-α = Tumor necrosis factor-alpha.

This action modulates the amount of circulating, bioactive TNF-α by binding to the cytokine before it can activate cell-surface receptors on mononuclear cells. Because TNF-α plays an important role in the eradication of neoplastic cells, its suppression is not without hazard—particularly as a long-term therapeutic strategy. Concerns have been raised as to whether chronic immunosuppression leads to opportunistic infection, malignancies, or other complications (Alldred A, 2001; Lee JH, 2002). CDP-870. UCB is developing CDP-870, a pegylated anti-TNF-α antibody fragment, for the treatment of RA and other inßammatory disorders such as Crohn’s disease in the United States and Europe. The agent was originally under development with Celltech, which was acquired by UCB in mid 2004, and is currently in Phase III trials. Celltech had previously been developing the agent in collaboration with PÞzer under an agreement terminated in December 2003. Regulatory Þlings are expected in 2006. No development has been reported in Japan. CDP-870 binds TNF-α with high afÞnity, thereby blocking its ability to activate the inßammatory cascade. Unlike inßiximab, which is a chimeric monoclonal antibody (mAb) composed of 75% human and 25% mouse protein, CDP-870 is a humanized anti-TNF-α mAb fragment conjugated to two polyethylene glycol (PEG) subunits. The agent is being developed as an injectable formulation with a once-monthly subcutaneous (SC) dosing regimen. UCB can manufacture CDP-870 in Escherichia coli at very low cost using proprietary technology. The production cost may be as little as 10% of that for creating an antibody or receptor fusion product using mammalian cell culture. In a Phase II, double-blind, placebo-controlled study involving 203 patients, CDP-870 signiÞcantly reduced the signs and symptoms of RA (Keystone E, 2001). In the study, patients were randomized to receive CDP-870 (50, 100, 200, or 400 mg) once monthly as an SC injection or placebo for 12 weeks. Clinical response was assessed using American College of Rheumatology (ACR) criteria. The percentages of patients achieving an ACR 20, ACR 50, or ACR 70 response

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at week 12 after treatment with the most-effective dose (400 mg) of CDP-870 were 72%, 48%, and 32%, respectively, compared with 15%, 0%, and 0% for placebo. The most common adverse events reported in this trial were headache, nausea, and upper respiratory tract infection. In October 2002, Celltech presented data from a Phase II clinical trial involving approximately 600 RA patients. Of the patients receiving CDP-870 (400 mg once monthly), 75% achieved an ACR 20 response seven days after initial administration. The company reported no signiÞcant differences in adverse events between CDP-870 (400 mg or 800 mg once monthly) and placebo, and no signiÞcant occurrences of injection-site reaction or urinary tract infection. In 2004 press releases, Celltech announced preliminary results from two Phase III studies in RA, in which CDP-870 was used both in combination with methotrexate (Stada’s Rheumatrex, generics) and as monotherapy. Again, the agent met the primary end point: a signiÞcant ACR 20 response at 24 weeks. A further trial required for registration, designed to assess the impact of CDP-870 on disease progression, was scheduled to commence in the second half of 2004. Additional information on the drug’s safety or efÞcacy were not announced in the press releases, but UCB stated that full results will be released after conclusion of the entire Phase III program in RA. CDP-870 treatment was considered to be well tolerated in a Phase II study in 36 RA patients (Choi EH, 2002). The most common adverse event reported was headache, and there was a higher incidence of mild or moderate lower respiratory tract infections and urinary tract infections in the treatment group as compared with placebo. Overall, CDP-870 appears to have a similar safety proÞle to the other TNF-α inhibitors, with the most common side effects being headache, nausea, and respiratory and urinary tract infections. Pegsunercept. Amgen has been evaluating the pegylated monomeric soluble TNF receptor type I inhibitor pegsunercept (PEG-sTNF-RI) in Phase II clinical trials in the United States. Although the product remains in the corporate portfolio, no clinical development has been reported since 2003, and Amgen appears to be pursuing other opportunities in RA. Despite the lack of recent development, pegsunercept is cited here as an emerging therapy because Amgen may decide to revive the product for use in combination regimens or as a defensive posture against market encroachment by CDP-870. Pegsunercept is a second-generation TNF-binding protein (TNF-bp) that comprises a soluble TNF-receptor type I (TNF-RI) subunit joined by a pegylated (PEG) linker. Like other TNF-α inhibitors, pegsunercept binds TNF-α with high afÞnity, thereby blocking its ability to activate the inßammatory cascade. As with CDP-870, pegsunercept is manufactured in Escherichia coli at a very low cost. At the 67th annual ACR meeting in October 2003, researchers presented results of a Phase II trial in which 309 patients with active RA were randomized to receive an SC injection of pegsunercept (400, 800, or 1,100 µg/kg) twice weekly or placebo for 24 weeks (Piercarlo S, 2003). All patients received concomitant methotrexate, and the primary end point was improvement in ACR response.

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At the end of the study period, all doses of pegsunercept signiÞcantly improved ACR 20, ACR 50, and ACR 70 responses, but the most signiÞcant response was seen for pegsunercept at 800 µg/kg. The ACR 20, 50, and 70 responses for pegsunercept 800 µg/kg in combination with methotrexate were 68%, 35%, and 25%, respectively, compared with 26%, 10%, and 3% for placebo. The results indicate that pegsunercept may reduce disease activity when administered concomitantly with methotrexate. The overall incidence of adverse events was similar between treatment groups, but injection site reactions were more frequent in patients receiving pegsunercept (38%, 36%, and 50% for 400, 800, and 1,100 µg/kg) than in those given placebo (30%). Pegsunercept has been evaluated in combination with other agents in clinical trials, suggesting that its efÞcacy as a monotherapy cannot compete with that of currently available drugs. Additionally, the ACR 20, 50, and 70 response rates for pegsunercept in combination with methotrexate are lower than those demonstrated by etanercept in combination with methotrexate in the Trial of Etanercept and Methotrexate with Radiographic Patient Outcomes (TEMPO) study: 85%, 69%, and 43%, respectively (Klareskog L, 2004). Interleukin-Based Therapies Overview. Despite the relatively low efÞcacy of the interleukin (IL)-1β antagonist anakinra and the relative lack of success of this agent in the RA market, companies continue to study IL-1 and other interleukins in the hope of Þnding a novel drug for RA. Three interleukin-based therapies that have advanced to comparatively late-stage development are discussed here, including Chugai/Roche’s atlizumab (Actemra, MRA) and Amgen’s AMG-714 (HuMax IL-15), which target IL-6 and IL-15 respectively. Abbott’s ABT-874, an anti-IL-12 mAb currently in Phase II studies, is not discussed owing to a lack of clinical data in RA. Regeneron’s IL-1 inhibitor RGN 303 was also in clinical trials for RA, but development was discontinued in September 2005. Mechanism of Action. The inßammatory cascade involves many proinßammatory cytokines in addition to TNF-α —notably, IL-1, IL-6, IL-8, IL-10, IL-12, IL-15, IL-17, IL-18, and IL-23. Dysregulated expression of these cytokines has been observed in RA patients and likely contributes to progressive joint degradation. Therefore, suppressing other inßammatory pathways may be an effective way to control RA, and blocking multiple pathways in the inßammatory cascade may act synergistically to block disease progression. Atlizumab. Chugai and its parent company Roche are developing atlizumab (Actemra, MRA), a humanized anti-IL-6 receptor mAb for the treatment of RA, Crohn’s disease, multiple myeloma, and Castleman’s disease (giant lymph node hyperplasia). In February 2003, the companies announced their intention to copromote the drug in the United Kingdom, France, and Germany. Roche will codevelop and promote atlizumab worldwide, except in Japan, South Korea, and Taiwan. Chugai also retained copromotion rights in the United States, Italy, and Spain. Phase III

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trials are ongoing in Japan and a Phase III preparation program is under way in Europe and the United States. Atlizumab inhibits the proinßammatory cytokine IL-6, which has a wide range of effects, including stimulating B cells to differentiate into plasma cells to produce immunoglobulin and stimulating T lymphocytes to differentiate into cytotoxic T cells. IL-6 is implicated in the pathogenesis of RA and stimulates chondrocytes, synoviocytes, and osteoblasts to secrete prostaglandins, plasminogen activator, collagenases, and other proteases. These substances promote the breakdown of extracellular matrix and cause extensive tissue destruction. Unlike the situation with the other cytokines, increased serum levels of IL-6 appear to correlate with increased levels of acute-phase reactant proteins (such as C-reactive protein and Þbrinogen) in arthritic patients. Results presented at the 66th annual ACR meeting in October 2002 demonstrate atlizumab’s efÞcacy and tolerability (Nishimoto N, 2002). In a multicenter, double-blind, placebo-controlled Phase II study, 164 patients with active RA and an inadequate response to disease-modifying antirheumatic drugs (DMARDs) were randomized to receive a monthly infusion of atlizumab (4 or 8 mg/kg) or placebo for three months. Patients were permitted to take prednisolone (10 mg/day or less). The primary end point was the ACR 20 response at week 12, at which point both doses of atlizumab demonstrated successful outcomes. ACR 20 responses for atlizumab 4 mg/kg and 8 mg/kg were 57.42% and 78.2%, respectively, compared with 11.3% for placebo; ACR 50 responses were 25.9%, 40.0%, and 1.9% and ACR 70 responses were 20.4%, 16.4%, and 0% of patients taking atlizumab 4 mg/kg, 8 mg/kg, and placebo, respectively. Patients receiving the 8 mg/kg dose also displayed 63.1% and 63.4% reductions, respectively, in the number of tender and swollen joints, compared with 7.7% and 2.6% reductions for placebo. Open-label extension study data were presented at the ACR 2003 meeting that conÞrmed the long-term safety of atlizumab. Subsequent to the original study, 142 patients were treated with 8 mg/kg of atlizumab intravenously every four weeks. Blood cholesterol increase was frequently observed, but stabilized at 3 months (185 mg/dL at base line; 224 mg/dL at 3 months; 225 mg/dL at 12 months). No cardiovascular events were observed (Nishimoto N, 2003). AMG-714. Amgen is developing AMG-714 (HuMax IL-15) under an agreement with Genmab of Denmark for the treatment of inßammatory conditions. The lead indication is RA, in which AMG-714 is currently being evaluated in U.S. Phase II trials. In July 2003, Genmab announced that Amgen had exercised its commercial option to AMG-714 and would be responsible for all further development costs for the product. In addition to the ongoing studies in the United States, Amgen is expected to initiate clinical studies for AMG-714 in Europe. AMG-714 is a fully human monoclonal IgG1 antibody that binds to human IL-15, a pro-inßammatory cytokine present in RA synovium, and inhibits IL-15induced effector functions. Data from an interim analysis of a Phase II study were presented at the ACR Annual ScientiÞc Meeting in October 2004 (McInnes I, 2004). In this study, 110

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patients naive to therapy with biologics and with active disease despite treatment with at least one conventional DMARD were randomized to receive one of four doses of AMG-714 (40, 80, 160, or 280 mg injected subcutaneously) or placebo every two weeks for 12 weeks. Stable background methotrexate, NSAIDs, and low-dose corticosteroids were continued. Clinical and safety assessments were performed at 2-week intervals for 16 weeks and monthly over the subsequent 8 weeks. The primary efÞcacy outcome measured was ACR 20 response at 12 weeks. At this time point, those patients receiving the highest dose (280 mg) demonstrated a 62% ACR 20 response compared with 26% for placebo. The other active treatment groups demonstrated higher ACR responses compared with placebo, but the differences were not statistically signiÞcant. The total incidence of adverse events in all four AMG-714 groups (60.9%) was similar to placebo (56.5%). The incidence of infectious adverse events was also similar between the 280 mg AMG-714 group (33.3%) and placebo (34.8%). RGN-303. Regeneron’s RGN-303 (IL-1 Trap) is in Phase IIb development in the United States for the treatment of RA. In March 2003, Regeneron signed an agreement with Novartis concerning its development and commercialization. However, in February 2004, after reviewing Phase II trial results, Novartis informed Regeneron that it would no longer continue development of the agent unless the terms of the collaboration were revised. Regeneron rejected Novartis’s proposed revisions and proceeded with clinical development alone. In September 2005, Regeneron discontinued development for the RA indication. RGN-303 is a fully human IL-1 inhibitor consisting of the Fc portion of IgG1 fused to the extracellular domains of both IL-1 receptor components, thereby mimicking the cell surface receptor for IL-1. RGN-303 binds to both IL-1α and IL-1β with high afÞnity, trapping the cytokine to block its activity. Results of a 12-week, placebo-controlled Phase II study in 201 patients with moderate to severe RA who had failed at least one prior DMARD were presented at the ACR Annual ScientiÞc Meeting in October 2004 (Bingham CO, 2004). Patients were randomized to receive weekly SC injections of placebo or 25, 50, or 100 mg of RGN-303. Stable doses of background conventional DMARDs, but not other biologics, were permitted. The primary end point was improvement in ACR 20 response at 12 weeks. At the end of the study period, 46.0% of the patients receiving 100 mg RGN-303 achieved an ACR 20 response compared with 30.9% in the placebo group; these results were not statistically signiÞcant. However, signiÞcant improvements in the Disease Activity Score (DAS28) of the 100 mg RGN-303 group over placebo were apparent by one week (−0.55 versus −0.29) and continued through the end of the study (−1.121 versus −0.702). The percentage of RA patients with a moderate or good response to treatment with 100 mg RGN-303 as assessed by the DAS28 was also signiÞcantly greater than placebo (46% versus 26%). The authors of this study concluded that further evaluation of the efÞcacy of the drug at higher doses is warranted. No signiÞcant infectious complications were seen, with the most common adverse event being injection site reactions.

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Although the agent failed to meet its primary end points in this study, Regeneron considered it possible that the optimal dose level for RGN-303 had not yet been achieved (Regeneron, press release, June 10, 2003). In April 2004, the company announced its intention of initiating a Phase IIb trial with RGN-303 in the second half of 2004, following the recommendation of an independent advisory panel of medical experts. The panel recommended that the new Phase IIb trial examine higher doses of RGN-303 in a larger patient population for a longer period of time compared with previous Phase II trial protocols (Regeneron, press release, February 27, 2004). Lymphocyte Modulators Overview. A new class of biological agents has emerged that speciÞcally target T and B lymphocytes, the main cells involved in an adaptive immune response. Most currently available biological agents target the products of activated macrophages, cytokines. However, T cells help activate macrophages, and both types of lymphocytes have been implicated in the inßammation and joint destruction found in RA. The rheumatoid synovium contains activated T and B cells, and immunoglobulin produced by B cells, such as rheumatoid factor and antinuclear or anticytoplasmic autoantibodies, serve as diagnostic markers for RA that may also be involved in the disease process. This section discusses one emerging therapy that targets T cells and one that targets B cells. Excluded from this analysis are lymphocyte modulators in Phase II clinical trials for which large-scale trial data are currently lacking, such as Human Genome Sciences/Cambridge Antibody Technology’s belimumab. Mechanism of Action. Lymphocyte modulators target T and B cells, which are immune cells known to be involved in the RA disease process. Agents in this class can act in a variety of ways, but the main mechanisms of action for therapies in late-stage development are interfering with lymphocyte activation or the actual depletion of the speciÞc cell population. Either way, the lymphocytes cannot carry out their effector functions, which contribute to the inßammatory state and the production of autoantibodies. Abatacept. Bristol-Myers Squibb is developing the CD28 antagonist abatacept (CTLA4-Ig BMS-188667) for the treatment of RA, multiple sclerosis, systemic lupus erythematosus (SLE), and other autoimmune disorders. Abatacept will be the Þrst of a new class of agents termed T-cell costimulation modulators. The FDA has granted fast-track status for RA in the United States, and a rolling biologics license application (BLA) submission was initiated in November 2004.The drug received FDA approval in December 2005, and a launch is expected in the Þrst half of 2006. No development has been reported elsewhere, but BMS is likely to submit further new product applications in European markets. Abatacept is a genetically engineered fusion protein that consists of the extracellular domain of human CTLA-4 and a fragment of the Fc portion of human IgG1. Abatacept binds to CD80 and CD86 molecules on antigen-presenting cells,

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blocking their engagement with CD28 on T cells. The blockade of this costimulatory signal prevents T-cell activation and the consequent proinßammatory cytokine release and acquisition of effector functions that contribute to the RA disease process. Published clinical trial results demonstrate that abatacept improves physical function in patients with active RA (Kremer JM, 2003). In this Phase IIb study, 339 patients refractory to methotrexate were randomized to receive intravenous administrations of abatacept (2 mg/kg or 10 mg/kg) or placebo on days 1, 15 and 30, and monthly thereafter for six months. All patients received concomitant methotrexate. At the end of the study period, the most signiÞcant ACR responses were seen with abatacept 10 mg/kg. The percentages of patients in the 10 mg group who achieved ACR 20, ACR 50, and ACR 70 responses were 60%, 37%, and 17%, respectively, compared with 35%, 12%, and 2% for placebo. The incidence of adverse events was similar for abatacept and placebo. The most frequent adverse events were headache, upper respiratory tract infection, musculoskeletal pain, and nausea. Further clinical response results over two years incorporating a one-year open-label extension study were presented at the 2004 ACR scientiÞc meeting. Of those patients randomized to the abatacept treatment arm, 75 (89%) completed two years of treatment. At two years, ACR 20, 50, and 70 responses were 77.3%, 54.7%, and 29.3%, respectively. Furthermore, 15% of patients demonstrated a 90% improvement in ACR criteria at two years (Kremer JM, 2004). The company is also pursuing development of abatacept in patients refractory to TNF-α inhibitors. Data from the Abatacept Trial in Treatment of Anti-TNF Inadequate Responders (ATTAIN) were presented at the ACR 68th Annual ScientiÞc Meeting in October 2004 (Genovese MC, 2004). In this placebo-controlled study, 391 patients with active RA were treated with at least one conventional DMARD and abatacept at a Þxed dose of 10 mg/kg (258 patients) or placebo (133 patients). All patients had discontinued TNF-α therapy. After six months of treatment, signiÞcant differences were observed in the ACR response rates: 50.4% of abatacept-treated patients achieved an ACR 20 response compared with 19.5% on placebo; 20.3% of abatacept-treated patients achieved an ACR 50 response versus 3.8% on placebo; and 10.2% of abatacept-treated patients achieved an ACR 70 response versus 1.5% on placebo. The incidence of adverse events was similar between abatacept and placebo, with the most common side effects being headache and nasopharyngitis. The incidence of serious infections was the same in each group (2.3%). Rituximab. Rituximab is being developed by Roche as MabThera and codeveloped by Biogen Idec and Genentech as Rituxan as a potential treatment for RA. Both compounds have already launched their respective products worldwide for the treatment of relapsed or refractory, low-grade, CD20-positive, Bcell non-Hodgkin’s lymphoma. Rituximab is being evaluated across a range of immunological indications, including multiple sclerosis, vasculitis, SLE, and RA. It is in Phase III clinical trials for anti-TNF-α refractory RA and Phase IIb clinical trials for moderate to severe RA in the United States and Europe. No development

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has been reported in Japan for RA, but rituximab is marketed there for B-cell lymphoma by Zenyaku Kogyo. This mouse/human chimeric mAb targets CD20 on the surface of B lymphocytes and recruits the body’s natural defenses to attack and kill the marked B cells. The labeling for rituximab indicates that the drug is associated with serious adverse events, including fatal infusion reactions, tumor lysis syndrome, mucocutaneous reactions, hypersensitivity reactions, cardiac arrhythmias and angina, and renal failure. The product label includes black box warnings for the Þrst three of these adverse events. Rituximab demonstrated efÞcacy for the treatment of RA in a Phase II trial of 161 partial methotrexate responders with long-standing disease (Edwards JCW, 2004). Study participants were randomized to receive methotrexate alone (≥10 mg per week), rituximab alone (1 g administered intravenously on days 1 and 15), rituximab plus cyclophosphamide (750 mg administered intravenously on days 3 and 17), or rituximab plus methotrexate. All groups also received a 17-day course of corticosteroids (100 mg intravenous methlyprednisolone on days 1, 3, 15, and 17; 30 mg oral methlyprednisolone on days 2 and 4 through 7; and 60 mg oral methlyprednisolone on days 8 to 14). Rituximab treatment resulted in nearly complete depletion of peripheral B cells, but peripheral blood immunoglobulin concentrations remained within normal ranges throughout the study period. At week 24, 43% of patients taking rituximab plus methotrexate achieved an ACR 50 response compared with 41% of patients taking rituximab plus cyclophosphamide, 33% of patients taking rituximab alone, and 13% of patients taking methotrexate alone. All ACR 50 responses for the patients treated with a combination of rituximab and methotrexate were maintained at 48 weeks. The majority of adverse events occurred with the Þrst rituximab infusion. At 24 weeks, serious infections occurred in 2.5% of the control group and in 3.3% of the rituximab groups. Subsequent data from the study were presented at the 68th annual ACR meeting in October 2004. Results demonstrated that a single course of rituximab in combination with methotrexate produced a substantial duration of response over two years. At week 104, ACR 20 and 50 responses were achieved by 34% and 21% of the rituximab plus methotrexate group compared with 14% and 11% on methotrexate alone, while 13% of the former group achieved a major clinical response (ACR 70 maintained for ≥6 months). The authors concluded that all three rituximab treatment arms were well tolerated with no signiÞcant differences observed in rates of infection (Emery P, 2004[a]). Many rheumatologists have employed rituximab off-label on a compassionate basis for patients refractory to currently available therapies. This agent is considered a highly exciting development in the Þeld of RA because it has the potential to induce long-term remission—up to two years, in some patients. Initially, rituximab is likely to be reserved for RA patients who are refractory to other DMARDs, conventional and biologic, although it is unclear what dosing strategy would be used. In clinical trials, rituximab has been administered intravenously weekly or every two weeks, but if the agent can truly induce

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long-term remission, dosing could be much less frequent. Excitement for the potential of the drug is tempered with serious concerns over the adverse events associated with rituximab. Administration of rituximab is linked to a high rate of infusion reactions, with up to 77% incidence for the Þrst infusion. Some of these infusion reactions have proved fatal, prompting the black box warnings discussed previously. Conventional Disease-Modifying Antirheumatic Drugs Overview. The conventional DMARD class includes a wide range of agents as diverse as antimalarials, cytotoxics, and immunosuppressants. Unlike antiinßammatories, which provide symptomatic relief without altering the natural course of the disease, DMARDs typically lack a direct analgesic effect but have the potential to slow or prevent joint damage in RA patients. In 2002, the ACR published revised guidelines for the treatment of RA advocating more aggressive use of conventional DMARDs earlier in the RA treatment regimen in an effort to limit joint damage and minimize loss of joint function and disability (ACR, 2002). As a result, the use of conventional DMARDs, which were once reserved for treating severe or late-stage RA, has become widespread and is now the cornerstone of RA treatment. The development of new DMARDs for the treatment of RA has focused primarily on biological agents. Biological agents, particularly the TNF-α inhibitors, have the best efÞcacy of all drugs used for the treatment of RA, but their high cost limits their widespread use. The production costs of synthetic compounds are much lower than those of biological agents, allowing them to be priced competitively and therefore used as Þrst-line DMARD therapy. Synthetic agents are also usually available in oral formulations that offer an additional advantage over the parenteral biological agents. Emerging conventional DMARDs, however, must contend with methotrexate, which is Þrmly established as the favored DMARD because it offers the best balance of efÞcacy, cost, and safety. Mechanism of Action. Conventional DMARDs include a wide range of agents, such as antimalarials, cytotoxics, and immunosuppressants, each with a distinct mechanism of action. In general, agents in this class are anti-inßammatory and/or antiproliferative, inhibiting various molecules and cell types that play a role in the inßammatory cascade. Most have demonstrated the ability to slow the rate of progression of joint erosion and disability to varying degrees. Iguratimod. Toyama Chemical is developing a chromone derivative, iguratimod (T-614), as a potential therapy for RA. As of November 2004, iguratimod was preregistered in Japan, while a Phase IIa program had been completed in the United States and Europe. Toyama has a codevelopment and comarketing agreement with Eisai, and Toyoma licensed Japanese marketing rights to Taisho Pharmaceuticals in 2003. We anticipate iguratimod will launch in Japan in 2006. Iguratimod was discovered in a program to develop novel anti-inßammatory compounds that preferentially inhibit cyclooxygenase (COX)-2 activity. The drug

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is now recognized as a DMARD, reßecting its ability to block the synthesis of proinßammatory cytokines and immunoglobulin production in addition to inhibiting COX-2 enzyme activity and COX-2 mRNA induction (Tanaka K, 2004). Unlike most other conventional DMARDs, iguratimod has direct analgesic properties that should help the drug provide symptomatic relief in addition to its disease-modifying ability. Iguratimod has demonstrated efÞcacy at least comparable to that of sulfasalazine. In a Japanese multicenter, double-blind trial presented at the 2004 European League Against Rheumatism (EULAR) meeting, 375 patients with active RA were randomly allocated to receive either iguratimod 50 mg/day (n = 146), sulfasalazine 1,000 mg/day (n = 156), or placebo (n = 73). The efÞcacy end point was ACR 20 response at 28 weeks. The ACR 20 response of the iguratimod group was 62.5%, which was comparable to the sulfasalazine group (58.1%). The presenters also stated that iguratimod was signiÞcantly superior to placebo, although the percentage of ACR 20 respondents in the placebo group was not shown. Elevated plasma enzyme levels (e.g., aspartate aminotransferase and alanine aminotransferase) were relatively frequent in the iguratimod group but were transient and improved after discontinuation (Hara M, 2004). The failure to provide placebo data in the study results hinders determination of the true efÞcacy of this agent. If it proves to have efÞcacy comparable to that of sulfasalazine, an agent widely considered to be a comparatively ineffective DMARD, iguratimod’s use will be limited. Furthermore, concerns over the cardiovascular safety of selective COX-2 inhibitors may limit uptake of this agent, since it does preferentially inhibit COX-2 in addition to its immunosuppression. Therefore, iguratimod is likely to be used in milder forms of RA or in combination with other DMARDs in European and U.S. markets, although the agent may become an alternative to salazosulfapyridine (sulfasalazine) in Japan. Selective Cyclooxgenase-2 Inhibitors Overview. The Þrst generation of selective COX-2 inhibitors, such as rofecoxib (Merck’s Vioxx) and celecoxib (PÞzer’s Celebrex), demonstrated their ability to reduce gastrointestinal (GI) side effects in comparison with traditional nonsteroidal anti-inßammatory drugs (NSAIDs), which suppress the isoforms of COX, COX-1 and COX-2, less discriminately. These agents still cause some degree of GI upset, however, so R&D efforts have been focused on developing a second generation of inhibitors that have an even higher degree of selectivity for COX-2. Valdecoxib (PÞzer’s Bextra) and etoricoxib (Merck’s Arcoxia) were the Þrst of this second generation of selective COX-2 inhibitors to be launched; they are reviewed in “Current Therapies.” Concerns regarding the cardiovascular safety of COX-2 inhibitors have drastically lowered expectations for new selective COX-2 agents. It remains unclear whether current selective COX-2 inhibitors will remain on the market and whether agents in the pipeline will launch. Although development of earlier-stage drugs in this class may be abandoned as a consequence of their signiÞcantly lowered sales potential, development for etoricoxib in new markets and for lumiracoxib

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(Novartis’s Prexige) will likely continue. Therefore, the discussion of emerging therapies in this drug class is limited to lumiracoxib. Mechanism of Action. COX-1 (also called constitutive cyclooxygenase) is present in cells under normal physiological conditions and stimulates the synthesis of prostaglandins that help regulate renal function, blood ßow, platelet activity, and protection of the mucous membrane along the GI tract. COX-2 (also known as inducible cyclooxygenase) occurs only under pathological conditions. Its production is induced by proinßammatory cytokines, mitogens, or endotoxins, and it stimulates the production of prostaglandins that drive the inßammatory process. Selective COX-2 inhibitors predominantly inhibit COX-2 while minimizing inhibition of COX-1. As a result, COX-2 inhibitors act speciÞcally to reduce pathological inßammation caused by prostaglandins, without disrupting their beneÞcial effects. Lumiracoxib. Lumiracoxib (Novartis’ Prexige, COX-189), at a dose of 100–200 mg daily, was approved by the United Kingdom Medicines and Healthcare Products Regulatory Agency (MHRA) in September 2003 for the treatment of osteoarthritis (OA). In addition, lumiracoxib at a dose of 400 mg daily was approved for the short-term relief of moderate to severe acute pain associated with primary dysmenorrhea and dental and orthopedic surgery. With these approvals, the United Kingdom was acting as the reference member state for the European mutual recognition procedure. However, Novartis temporarily withdrew its European Union mutual recognition procedure application in November 2004 pending European Agency for the Evaluation of Medicinal Products (EMEA) review of the selective COX-2 inhibitor class in 2005. The U.K. approval of lumiracoxib came on the heels of news that the FDA had requested additional clinical data on lumiracoxib for OA and acute pain. The FDA also denied the registrational Þling for RA. By January 2003, Novartis had initiated an additional clinical trial for RA. In October 2004, the company revised its anticipated U.S. resubmission date for all indications to 2007. Similar to other agents in its class, lumiracoxib predominantly inhibits COX-2 while minimizing inhibition of COX-1. As a result, lumiracoxib acts speciÞcally to reduce pathological inßammation caused by prostaglandins without disrupting their beneÞcial effects. Data from a Phase III study demonstrated that the efÞcacy of lumiracoxib is signiÞcantly better than placebo and similar to naproxen in the reduction of RA symptoms (Geusens P, 2004). In the study, 1,124 patients with symptomatic RA were randomized to receive lumiracoxib 200 mg once daily, lumiracoxib 400 mg once daily, naproxen 500 mg twice daily, or placebo for 26 weeks. The primary end point was ACR 20 response at week 13, and secondary end points included ACR 20 response at weeks 4, 20, and 26. After 13 weeks of treatment, an ACR 20 response was achieved by 41.1% and 42.7% for lumiracoxib 200 and 400 mg, respectively, compared with 39.1% and 32.4% for naproxen and placebo. Similar response levels were observed after 26 weeks of treatment. The rate of discontinuation due to adverse events was similar in all treatment

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groups. However, incidence of GI events was 15.7% and 16.4%, respectively, for lumiracoxib 200 mg and 400 mg, compared with 19.7% and 10.9% for naproxen and placebo. A meta-analysis presented at the annual meeting of the American Society for Gastrointestinal Endoscopy in May 2003 suggests that lumiracoxib’s GI safety proÞle is superior to that of traditional NSAIDs and comparable to that of celecoxib and rofecoxib. This analysis, which included 6,295 patients and data from nine OA studies, showed that the overall incidence of GI events was 22% for lumiracoxib (200 and 400 mg daily), 22% for celecoxib (200 mg daily), 29% for rofecoxib (25 mg daily), and 47% for ibuprofen (2,400 mg daily) (Hawkey CJ, 2003). Further evidence of an improved GI safety proÞle for lumiracoxib as compared with traditional NSAIDs came from the large-scale Therapeutic Arthritis Research and Gastrointestinal Event Trial (TARGET). In this study, 18,325 OA patients were randomized to receive lumiracoxib 400 mg once daily (n = 9, 156), naproxen 500 mg twice daily (n = 4, 754), or ibuprofen 800 mg three times daily (n = 4, 415) for 52 weeks in two substudies of identical design. The primary gastrointestinal end point was the one-year incidence of upper GI ulcer complications: bleeding, perforation, or obstruction. Lumiracoxib demonstrated a signiÞcantly lower incidence of ulcers than the traditional NSAIDs in the overall population and in patients not taking aspirin, but the difference was not signiÞcant for the group taking aspirin (Schnitzer TJ, 2004). Thus, lumiracoxib demonstrated a three- to fourfold decrease in ulcer complications compared to traditional NSAIDs, although concomitant aspirin appears to negate lumiracoxib’s improved GI safety proÞle. The critical question of lumiracoxib’s cardiovascular safety was also addressed in TARGET. The primary cardiovascular end point in that study was deÞned as the incidence of major adverse cardiovascular events—nonfatal and silent myocardial infarction, stroke, or cardiovascular death. At one year, incidence of the primary end point was low with both lumiracoxib (59 events [0.65%]) and the NSAIDs (50 events [0.55%]). No signiÞcant differences were observed between lumiracoxib and the comparator NSAIDs in the overall population or in the subpopulations investigated separately (patients taking or not taking low-dose aspirin). Furthermore, no signiÞcant differences were observed in the incidence of myocardial infarctions, congestive heart failure, and other thrombotic events in the overall population groups studied (Farkouh ME, 2004). The fate of this agent is inextricably linked with that of the other members of this class, currently under review by regulatory agencies in the United States and Europe. While the results of the TARGET study provide some encouragement, increased cardiovascular risk associated with rofecoxib therapy did not become apparent until 18 months. Immunosuppressants Overview. Even though immunosuppressive agents such as azathioprine (GlaxoSmithKline’s Imuran, generics) and cyclosporine (Novartis’ Sandimmune/

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Neoral, generics) have been used for decades to treat the signs and symptoms of RA, development of novel agents in this class for RA has been sluggish because of efÞcacy and safety concerns that have arisen in clinical trials of agents under development. Nonetheless, companies are pursuing development of immunosuppressive therapies. Excluded from this analysis are immunomodulators currently in Phase II trial programs but for which large-scale trial data are currently lacking, including Androclus Therapeutics’ AT-001 and Wyeth’s temsirolimus. Mechanism of Action. Immunosuppressants function through a variety of mechanisms, but they generally target key immunologic processes and molecules. In general, these agents exert their effects by blocking the activation of T-helper and cytotoxic T lymphocytes, promoting the accumulation of anti-inßammatory molecules, and decreasing the formation of antibodies. Tacrolimus. Tacrolimus, or FK-506 (Astellas’s Prograf) (Figure 16), has been launched for atopic dermatitis and for the prophylaxis of liver and kidney allograft rejection. A supplementary new drug application (NDA) was Þled in Japan for RA in November 2002, and in April 2005 the drug was approved for the treatment of RA in patients who responded insufÞciently to conventional treatments. The drug will be marketed in Japan by Astellas, formed from the merger of Fujisawa and Yamanouchi. The agent is in Phase III trials in the United States and in Phase II in Europe. Tacrolimus is a microbial product isolated from the bacterium Streptomyces tsukubaensis. The drug binds to a T-cell-speciÞc FK-binding protein, preventing T-cell activation, proliferation, and survival. As an immunosuppressant, tacrolimus is 50–100 times more potent than cyclosporine, as measured by inhibition of lymphocyte activation in vitro. HO

H3CO

CH3 H3C HO

O

O N O O

O

H3C

HO H3C

H3C O

OCH3 OCH3 FIGURE 16. Structure of tacrolimus.

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The safety and efÞcacy proÞles of tacrolimus appear to be favorable both as monotherapy and in combination with methotrexate. A six-month, randomized, double-blind, placebo-controlled Phase III trial that enrolled 464 patients with active RA who were intolerant of or refractory to one or more DMARDs, including methotrexate, found that 18.8% and 26.8% of patients receiving 2 mg and 3 mg, respectively, of tacrolimus daily achieved an ACR 20 response compared with 10.2% of patients receiving placebo (Yocum D, 2003). Of the patients receiving 2 mg and 3 mg of tacrolimus, 11.7% and 11.8%, respectively, achieved ACR 50 responses compared with 4.5% of patients in the placebo group. A 12-month, open-label extension of this trial, which enrolled 896 patients, sought to establish the long-term safety of tacrolimus (Yocum D, 2004). This study involved patients who chose to enroll in the extension trial, together with an additional 685 patients who either had never received tacrolimus or had not received it in 11 months or more. All patients received 3 mg of tacrolimus daily; 38.4% achieved ACR 20, 18.6% achieved ACR 50, and 9.0% achieved ACR 70. The drug was found to be generally well tolerated in this study; minor side effects included diarrhea (14.6%), nausea (10.3%), tremor (9.0%), and headache (8.7%). Tacrolimus is occasionally used off-label in RA patients who are refractory to methotrexate and other marketed DMARDs, a practice that may be indicative of the limited patient population to whom this agent will be prescribed when it is approved for RA. MAP Kinase Inhibitors Overview. Biotechnology companies such as Scios (a subsidiary of Johnson&Johnson), Vertex, Celgene, and Cephalon dominate the Þeld of mitogenactivated protein (MAP) kinase-based therapeutics. These companies are developing oral small-molecule inhibitors of MAP kinase for the treatment of inßammatory disorders such as RA and Crohn’s disease as well as oncology indications. This section discusses those agents that have advanced to Phase II trials, the most advanced stage of development for members of this class. Mechanism of Action. The enzyme p38 MAP kinase is part of an intracellular signal transduction cascade involved in regulating the expression of several proinßammatory cytokines, including IL-1β and TNF-α. Patients with active RA exhibit increased activation of p38 MAP kinase (Pargellis C, 2003). Researchers believe that p38 MAP kinase plays an important role in the induction of inßammation, and that agents targeting this pathway may provide a more effective and complete blockade of inßammatory cytokines than biological cytokine inhibitors. The potential therapeutic beneÞt of targeting the MAP kinase pathway in RA is also suggested by preclinical studies with selective inhibitors of p38 MAP kinase. In one study, SmithKline Beecham’s (now GlaxoSmithKline’s) research compounds SB-203580 and SB-202190 arrested disease progression in a rat adjuvant-induced arthritis model (Liverton NJ, 1999). In a later study, these same agents blocked the expression of the matrix metalloproteinases MMP-1 and MMP-13 (Ieda Y, 2001). MMP-1 and MMP-13 play a role in cartilage destruction and inßammation in RA.

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SCIO-469. Scios is developing SCIO-469, an oral p38 MAP kinase inhibitor, for the treatment of RA and Crohn’s disease. The agent is in Phase II development in the United States and Europe. No development in Japan has been reported. SCIO-469 blocks the activity of MAP kinase, which in turn inhibits the production of proinßammatory factors such as TNF-α, IL-1β, and COX-2 mRNA. Thus, the agent can be thought of as an indirect TNF-α inhibitor with additional anti-inßammatory effects. By July 2004, Johnson&Johnson reported that two Phase IIa studies of SCIO469 for RA had been successfully completed and a Phase IIb efÞcacy and tolerability trial was under way. The main objective of the Þrst of these studies was to evaluate six escalating doses of SCIO-469 compared with placebo in 120 patients with active RA receiving concomitant methotrexate. The second study was a 24-week, placebo-controlled trial to determine the efÞcacy of SCIO-469 in RA patients not receiving DMARDs other than hydroxychloroquine. Results from the studies have not yet been reported. In April 2001, Scios reported data from a randomized, double-blind, placebocontrolled Phase Ib trial conducted in the United Kingdom that evaluated safety, tolerability, and pharmacokinetics in 30 healthy volunteers over a wide range of doses (Scios, press release, October 20, 2000). According to the company, the drug demonstrated excellent bioavailability and pharmacokinetics and was well tolerated. In October 2002, at the 66th annual ACR meeting, the company presented preclinical data on SCIO-469. In a 28-day study involving rats with preexisting RA, administration of SCIO-469 (10 mg/kg or 40 mg/kg) once daily resulted in a signiÞcant dose-related reduction in the severity of arthritis (joint swelling and erythema). By day 28, the clinical severity score (a measure of disease severity with higher scores indicating greater disease severity) was 5.10 in the control group compared with 4.25 for SCIO-469 10 mg/kg group and 2.16 for SCIO469 40 mg/kg group. In addition, the mean radiographic score for the control group was 3.30 compared with 2.62 for SCIO-469 10 mg/kg group and 0.16 for SCIO-469 40 mg/kg group (Brahn E, 2002). Doramapimod. Boehringer Ingelheim’s doramapimod (BIRB-796) is in Phase II trials in the United States and Europe for RA, psoriasis, and Crohn’s disease. No development has been reported in Japan. Doramapimod, a pyrazole phenyl urea derivative, is a reversible, competitive inhibitor of p38 MAP kinase that acts by interfering with adenosine triphosphate (ATP) binding of p38. Like SCIO-469, this agent blocks TNF-α and IL-1β synthesis. Multiple Phase I trials have demonstrated the agent’s tolerability and lack of signiÞcant adverse events. The Þrst human study conducted with this drug was a randomized, double-blind, placebo-controlled, 64-patient trial presented at the 2002 annual meeting of the American Academy of Asthma, Allergy, and Immunology. In this study, doramapimod administered as an oral solution with PEG400 exhibited safety in doses ranging from 1 mg to 600 mg (Gupta A, 2002).

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Diabetic Retinopathy

ETIOLOGY AND PATHOPHYSIOLOGY Introduction Diabetic retinopathy—described by ICD-10 code H36.0+—is the leading cause of new blindness among adults aged 20–74 (Fong DS, 2003). This condition is characterized by abnormal blood vessel growth in the retina, the light-sensitive tissue at the back of the eye that communicates light signals to the brain. Diabetic retinopathy can be classiÞed by severity into a Þrst stage of nonproliferative retinopathy and a later, progressive stage of proliferative retinopathy. Uncontrolled proliferative retinopathy can progress to the point where complications can eventually lead to severe visual acuity loss and/or blindness. Anatomy Figure 1 illustrates the anatomy of the eye. Hyperglycemia-mediated damage to the eye occurs primarily in the retina, the nerve-dense, light-sensitive tissue that lines the interior of the eye. The macula is the avascular center portion of the retina responsible for central vision. At the center of the macula is the fovea, a further specialized retinal section that is responsible for Þne, sharp vision. Retinal tissue (also detailed in Figure 1) consists of three primary cell types: neurons, glial cells, and blood vessels. Inputs from four types of retinal neurons (photoreceptors and amacrine, bipolar, and horizontal cells) are transmitted to the cells of the ganglia, which then convey electrical output to the nerve Þber layer and optic nerve. Retinal neurons are highly redundant; approximately half of neural cells can be damaged or destroyed before signiÞcant functional impairment or visual acuity loss will occur. Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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FIGURE 1. Anatomy of the eye.

M¨uller cells and astrocytes are the two key glial cell types found in the retina, and they function as the metabolic modulators for neural and vascular components of the retina (Abbott NJ, 1992). Both cell types regulate ion concentrations, neurotransmitters, and nutrients for neural cells. Astrocytes also play an important role in the development and differentiation of vascular endothelial cells, particularly during fetal development (Zhang Y, 1997). Capillaries, arterioles, and venules constitute the important components of the retinal vascular network. Arterioles are the main “valves” through which blood ßows into the retina. Smooth muscle present in arterioles allows them to regulate the resistance they pose to blood ßow. Venules are largely passive vessels that drain blood out of the retina. Although passive, venules do possess a large number of receptors for vasoactive agents. All retinal blood vessels contain two types of cells: endothelial cells and pericytes. Pericytes act like modiÞed smooth muscle cells to control capillary tone in the retina. The endothelial cells in the retina are like other vascular cells in the body except that they have tight junctions that prevent leakage. These tight junctions are the functional component of the blood-retina barrier that allows the retina to self-regulate metabolism and homeostasis and to protect retinal neurons from circulating cytotoxic agents. A number of proteins, including occludin and claudins, are responsible for maintaining the tight junctions and limiting ßuid ßow between endothelial cells (Gardner TW, 2002).

ETIOLOGY AND PATHOPHYSIOLOGY

5

FIGURE 2. Pathological features of diabetic retinopathy.

Pathophysiology In many cases, the early stages of diabetic retinopathy do not manifest any obvious symptoms (e.g., visual acuity loss). Therefore, the pathophysiology of the disease is described primarily by clinical signs of retinal abnormalities. Figure 2 illustrates the common clinical signs of diabetic retinopathy. Vascular changes are the hallmark of diabetic retinopathy, but recent research also points to cellularlevel deÞcits in neural function that may be an equally important component of the disease. Microvascular Abnormalities. With the onset of diabetic retinopathy, two important changes occur in capillary cells. The tight junctions between endothelial cells loosen and the capillaries become permeable, allowing the inÞltration of glial cells, leukocytes, and other materials. This inßux of cellular material—which adheres to vessel walls—eventually causes capillary occlusion. Second, pericytes die and leave behind “pericyte ghosts.” Without the smooth-muscle activity of the pericytes, the capillaries dilate beyond normal levels, allowing the formation of microaneurysms and other early intraretinal microvascular abnormalities (IRMAs). Capillary microaneurysms appear as small red dots distributed sporadically throughout the retina. As this condition worsens, arteriovenous shunts occur, and bleeding into the nerve Þber layer causes ßame-shaped, blot-shaped, or linear lesions.

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Neovascularization (growth of new blood vessels) reßects the eye’s attempt to restore retinal blood ßow to areas rendered hypoxic by ischemia and signiÞcant vascular occlusion. New blood vessels grow perpendicularly from the surface plane of the retina into the vitreous space. These new vessels are typically hyperpermeable (allowing the inßux of materials) and more prone to hemorrhage than the normal retinal microvasculature. Consistent with wound healing in other tissues, the vessels are eventually subjected to a process of remodeling that includes Þbrosis and replacement with collagen (Gardner TW, 2000). The Þbro-vascular tissue can hemorrhage and/or scar, causing preretinal hemorrhage, vitreous hemorrhage, and/or retinal detachment (described later in the section “Complications”). Retinal Neurodegeneration. Research suggests that defects in retinal glial cell and neuron function precede the development of microvascular abnormalities in diabetic retinopathy and may be an important step in disease pathogenesis. Indicators of glial cells’ metabolic dysfunction include decreased production of glial Þbrillary acidic protein (GFAP; a marker of astrocyte function), decreased conversion of glutamate to glutamine by M¨uller cells, and increased production of cytokines that are known to increase vascular permeability (Gardner TW, 2002). Subtle defects in vision also suggest that retinal neurons incur damage from hyperglycemia. Using electroretinogram (ERG) measurements of retinal neuron electrical activity, researchers demonstrated that the amplitude of oscillatory potentials was reduced in type 1 diabetics within the Þrst Þve years after disease onset but before the development of clinical signs of retinopathy (FrostLarsen K, 1980). Disease Severity. Diabetic retinopathy is graded in two severity stages: nonproliferative and proliferative. The gold standard for measurement of severity is the criteria developed more than a decade ago in the Early Treatment Diabetic Retinopathy Study (ETDRS). This staging system relies on clinical signs that are evident with fundus photography using seven standard stereoscopic Þelds, and the combined number and position of microvascular abnormalities dictates the severity level (ETDRS Research Group, 1991). Although widely used in clinical trials, the ETDRS staging system is fairly complex and not commonly used for severity grading in practice. In an attempt to simplify the ETDRS criteria, the International Council on Ophthalmology recently proposed new severity scales for diabetic retinopathy (Ciulla TA, 2003; International Council on Ophthalmology, 2002). The International Diabetic Retinopathy Severity Scale is essentially a simpliÞcation of the ETDRS classiÞcations but is intended for use with dilated ophthalmoscopy investigation. Table 1 summarizes the international scale. Nonproliferative retinopathy (previously called background retinopathy) is the slowly progressing, less severe stage of the disease that is characterized by nonneovascular changes in the retina. The severity of nonproliferative retinopathy is determined by the location and quantity of retinal microvascular abnormalities, including microaneurysms, intraretinal hemorrhages, venous beading, and other vascular lesions. Proliferative retinopathy is characterized by neovascularization

ETIOLOGY AND PATHOPHYSIOLOGY

7

TABLE 1. Proposed International Clinical Diabetic Retinopathy Disease Severity Scale Severity Level No apparent retinopathy Nonproliferative diabetic retinopathy Mild Moderate

Severe

Proliferative diabetic retinopathy

Dilated Ophthalmoscopy Findings

Comparable ETDRS Stage(s)

No retinal abnormalities

No retinopathy: Level 10

Microaneurysms present, no other clinical signs Microaneurysms and other signs (intraretinal hemorrhages, venous beading, and/or IRMA) present, but less than severe nonproliferative diabetic retinopathy No signs of proliferative diabetic retinopathy, with any of the following: >20 intraretinal hemorrhages in each of four quadrants, definite venous beading in ≥2 quadrants, prominent IRMA in ≥1 quadrant Neovascularization and/or vitreous or preretinal hemorrhage

Very mild nonproliferative diabetic retinopathy: Level 20 Moderate nonproliferative diabetic retinopathy: Levels 35–47

Severe and very severe nonproliferative diabetic retinopathy: Level 53

All stages (high-risk, very severe, and/or advanced proliferative diabetic retinopathy): Levels 61 and higher

ETDRS = Early Treatment Diabetic Retinopathy Study. IRMA = Intraretinal microvascular anomalies.

on the retina, disc, and/or iris and is a quickly progressing, vision-threatening stage of disease. Research has demonstrated that in the preclinical stage of diabetic retinopathy, changes in retinal function (as measured by ERG) can cause perceptive resolution deÞcits, including impaired contrast sensitivity (particularly blue-yellow contrast) and impaired night vision (Bangstad HJ, 1994; Barber AJ, 2003; Della Sala S, 1985; Sokol S, 1985). However, because it precludes the development of classic vascular abnormalities, preclinical diabetic retinopathy is not a widely accepted disease severity stage. Complications. Macular edema and retinal traction detachment are signiÞcant complications of diabetic retinopathy that can, if left untreated, lead to blindness. Macular edema is a thickening/swelling of the center portion of the retina caused by deterioration of the blood-retina barrier that allows leakage from retinal capillaries into normally nonperfused tissues. Clinically signiÞcant macular edema (CSME) is deÞned as thickening of the macula or the area within 500 µm of the macula, hard exudates within 500 µm of the macula, and/or any thickening of the retina one disk area or more in diameter that is within one disk diameter of the center of the retina (ETDRS Research Group, 1985).

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Retinal detachment generally describes the separation of the retina from the underlying retinal pigment epithelium (RPE). There are three main types of retinal detachment: rhegmatogenous, where a tear in the retain causes detachment; exudative, where ßuid accumulation in the subretina causes detachment; and tractional, where scar tissue on the retinal surface contracts and pulls the retina off the RPE. The tractional type is most often associated with diabetic retinopathy. The Þbro-vascular tissue produced in the proliferative stage of disease eventually scars, causing retinal contraction and traction detachment. If the area of detachment includes the macula, signiÞcant vision impairment can arise. Etiology The Growth Factor Hypothesis. In 1948, I.C. Michaelson published what has been termed the “growth factor hypothesis” (Aiello LP, 2000; Michaelson IC, 1948). His hypothesis was that retinal ischemia (like that found in diabetic retinopathy) promotes production of angiogenic growth factors that would be responsible for proliferative neovascularization. Researchers have since focused on a number of likely candidates, including basic Þbroblast growth factor (bFGF), growth hormone (GH), and hepatocyte growth factor (HGF). However, a large body of in vitro and in vivo evidence has narrowed the focus of this search to one highly probable etiological factor: vascular endothelial growth factor (VEGF). VEGF is a potent angiogenic agent whose production is upregulated by hypoxia in RPE cells and retinal pericytes (Adamis AP, 1993; Plouet J, 1993). A study of ocular ßuid taken from the eyes of patients with diabetic retinopathy and other neovascularizing eye disorders conÞrmed VEGF’s association with neovascular retinal disorders (Aiello LP, 1994). In the ocular ßuid of diabetics, VEGF was detectable in 83% of samples from patients with active proliferative diabetic retinopathy, 22% of samples from patients with quiescent proliferative diabetic retinopathy, and only 8% of samples from patients with nonproliferative diabetic retinopathy. Not only does VEGF promote retinal neovascularization, but it is also a highly effective inducer of vascular permeability. Research with bovine retinal endothelial cell cultures demonstrated that the administration of VEGF causes a 46–54% reduction in occludin concentrations (Antonetti DA, 1998). Occludin is the “glue” between retinal endothelial cells that helps maintain the blood-retina barrier, so this research suggests that VEGF-mediated loss of occludin may be a key process underlying the retinal vascular “leakiness” and hemorrhage that occur in diabetic retinopathy. The Role of Hyperglycemia. The metabolic abnormalities of diabetes contribute to the destruction of blood vessels in the eye, just as these abnormalities affect blood vessels in many other organs. The etiological link between hyperglycemia and diabetic retinopathy has not been conclusively established, but several metabolic mechanisms have been proposed to explain how chronic hyperglycemia leads to retinal damage (Figure 3):

ETIOLOGY AND PATHOPHYSIOLOGY

9

FIGURE 3. Pathways of hyperglycemia-induced damage.

• • •

The activation of the enzyme protein kinase C. The accumulation of advanced glycation end-products (AGEs). The overactivation of the polyol pathway.

Protein Kinase C. The protein kinase C isozymes are a family of 12 related serine/threonine enzymes found in nearly every tissue and cell type in the body (Aiello LP, 2000; Koya D, 1998). These enzymes have many responsibilities, including cell proliferation, differentiation, and apoptosis. One member of this family, protein-kinase-C beta (PKCβ), has been the target of diabetes research since researchers linked it to hyperglycemia-induced processes. The predominant hypothesis holds that AGEs and oxidants produced in nonenzymatic glycation and the polyol pathway, respectively, are converted to diacylglycerol (DAG). In turn, DAG synthesis promotes the activation of PKCβ (Xia P, 1994). Diabetic

10

DIABETIC RETINOPATHY

animal models have shown high levels of the enzyme in the retina, heart tissues, and renal glomeruli. PKCβ activation in the retina is believed to promote the overgrowth of blood vessels characterizing diabetic retinopathy, primarily through its relationship to VEGF activity. The β isozyme primarily causes cell proliferation, and both animal and human models have shown that blockade of PKCβ stops hypertrophy of the retinal vasculature. PKCβ promotes the expression of VEGF and is a critical mediator of the proliferative and permeability effects of VEGF (Aiello LP, 1997; Xia P, 1996). Furthermore, PKCβ can mediate signal transduction initiated by hormones such as vasopression and angiotensin II, possibly leading to renin-angiotensin-aldosterone system (RAAS) dysfunction. The RAAS is a critical factor in the regulation of the vasculature in the retina and in other body tissues. Advanced Glycation End Products. Advanced glycation end products (AGEs) are formed through a process called nonenzymatic glycosylation. AGEs make up a heterogeneous group of proteins, nucleic acids, and lipids that have been exposed and irreversibly bound to reducing sugars. They are believed to form when blood and tissue glucose levels increase, thereby chemically modifying various extracellular and intracellular macromolecules. One such AGE is glycosylated hemoglobin A1c (HbA1c ), the overproduction of which is a key clinical marker of diabetes-related hyperglycemia. AGEs can alter the structural and functional properties of proteins. Much of this damage is believed to originate intracellularly, particularly in vasculature endothelial cells. It is believed that AGEs damage endothelial cells by intensifying oxidative stress and possibly by upregulating gene transcription, both of which may contribute to vascular disease. The AGE-mediated overproduction of superoxide anion (O-2) creates oxidative stress, which can promote abnormal regulation of apoptosis (cell death). Hyperglycemia also affects the primary enzyme that detoxiÞes O-2, superoxide dismutase 1 (SOD1), which is deactivated by glycosylation, leaving no counterbalance to the damage caused by AGEs. AGEs accumulate in the retinal capillaries, leading to capillary basement membrane thickening, decreased elasticity, and increased leakiness. The Polyol Pathway. A large body of research has studied the role of the enzyme aldose reductase in diabetic complications. Excessive activity of aldose reductase, which reduces glucose to sorbitol via the polyol pathway, causes sorbitol to overaccumulate in cells, producing osmotic stress and creating structural and functional abnormalities in sensitive tissues such as the eye. Because aldose reductase has a low afÞnity for glucose, the polyol pathway has only minimal importance in people with normal glucose levels. In people with hyperglycemia, however, the excessive availability of glucose pushes this reaction in favor of aldose reductase activity. The polyol pathway can account for as much as 30% of glucose metabolism in diabetics with hyperglycemia. Excessive aldose reductase activity may be responsible for overactivation of the GLUT-1 transport mechanism, allowing excessive amounts of glucose to enter

ETIOLOGY AND PATHOPHYSIOLOGY

11

endothelial cells. Myoinositol competes with glucose for transport into cells. If aldose reductase enables glucose uptake to increase signiÞcantly, myoinositol will be unable to enter the cell. Abnormal sorbitol and myoinositol levels have also been implicated in increased vascular permeability in the blood/retinal barrier, as well as glomerular hyperÞltration (a kidney abnormality that is an early symptom of diabetes) and nerve cell dysfunction. Risk Factors Duration of Diabetes. The strongest predictor for both development and progression of diabetic retinopathy is the duration of diabetes. The Wisconsin Epidemiologic Study of Diabetic Retinopathy (WESDR) found that the prevalence of diabetic retinopathy (any stage) was 8% at 3 years after type 1 diabetes diagnosis, increasing to fully 80% at 15 years (Klein R, 1984[a]). The prevalence of PDR was 0% at 3 years and 25% at 15 years. Glycemic Control. Observational data from the WESDR showed a signiÞcant correlation between HbA1c at baseline at the development or progression of diabetic retinopathy among type 1 diabetics, type 2 diabetics who used insulin, and type 2 diabetics who did not use insulin. Depending upon type of diabetes and HbA1c level, the relative risk of developing diabetic retinopathy was 1.1–2.7 compared with the quartile of patients with the lowest average HbA1c . The risk of any disease progression was 1.1–4.3, and the risk of progression to proliferative diabetic retinopathy was 1.2–13.8 (Klein R, 1988; Klein R, 1994; Klein R, 2000). The 1993 publication of the landmark Diabetes Control and Complications Trial (DCCT) conclusively demonstrated that interventional strict glycemic control prevents the development and slows the progression of microvascular complications (including diabetic retinopathy) in type 1 diabetics. The United Kingdom Prospective Diabetes Study (UKPDS) and the Steno Type 2 Randomized Trial found similar results in type 2 diabetics. Table 2 highlights key points from these trials. Hypertension. Hypertension is assumed to be a risk factor for diabetic retinopathy, although it has not been proved to play a role in the progression from the nonproliferative stage to the proliferative one. Researchers believe hyperglycemia causes an increase in retinal blood pressure that causes capillary damage. The role of systemic hypertension in diabetic retinopathy is less clear; studies investigating a possible link have produced conßicting results. Observational studies have examined the role of hypertension in development and/or progression of diabetic retinopathy. The UKPDS compared retinal photographs from 1,919 newly diagnosed type 2 diabetics at diagnosis and six years later. The study found that systolic blood pressure at baseline was signiÞcantly associated with the incident of diabetic retinopathy, and patients in the highest blood pressure tertile were 2.8 times more likely to develop diabetic retinopathy than patients in the lowest tertile (Stratton IM, 2001). However, baseline blood pressure was not signiÞcantly associated with progression of existing retinopathy.

12

DIABETIC RETINOPATHY

TABLE 2. Hyperglycemia and Diabetic Retinopathy: Select Results from Clinical Trials Trial

Details of Study

Selected Results

Diabetes Control and Complications Trial (DCCT)

In patients with no initial The DCCT was conducted from retinopathy, IIT reduced the risk 1983 to 1993 and followed 1,441 of sustained disease by 76%. type 1 diabetics. Intensive insulin In patients with mild initial therapy (IIT) was employed to retinopathy, IIT reduced the risk maintain strict glycemic control, of sustained retinopathy by defined as serum HbA1c levels no higher than 7%. 54%, the risk of developing severe proliferative retinopathy by 47%, and the need for laser treatment by 56% (The DCCT Group, 1993). The U.K. The UKPDS randomized 3,867 A modest decrease in HbA1c Prospective type 2 diabetics to receive (11%) reduced the risks for Diabetes Study conventional dietary advice progression to retinopathy by (UKPDS) (fasting glucose levels at 15 21%. mmol/L) or dietary advice in This reduction in HbA1c lowered the risk of microvascular combination with either intensive complications by as much as sulphonylurea or intensive insulin 25%. treatment (fasting glucose levels The median complication-free at less than 6 mmol/L). interval was 1.3 years longer for the intensive treatment group (UKPDS Group, 1998[a]). Progression of retinopathy—either The Steno Type 2 One hundred and sixty type 2 a worsening of existing Randomized Trial diabetics with microalbuminuria retinopathy or development of (albumin excretion rate of new retinopathy—occurred in 19 30–300 mg/day) were followed intensively treated and 33 for four years. Patients were conventionally treated patients randomized to receive either (Gaede P, 1999). standard treatment according to Danish guidelines (n = 80) or intensive treatment that maintained HbA1c levels at 6.5% (n = 80). Full source citations appear in ‘‘References.’’

The WESDR also found that the risk of proliferative retinopathy after 14 years of follow-up was positively correlated with diastolic blood pressure at baseline among type 1 diabetics. However, neither systolic nor diastolic blood pressure was associated with incidence or progression of diabetic retinopathy among type 2 diabetics (Klein R, 1998, Klein R, 2002). The Þrst interventional trial to catch the attention of physicians and other experts was the EURODIAB Controlled Trial of Lisinopril in Insulin Dependent Diabetes (EUCLID). For two years, 530 type 1 diabetics in European centers were randomized to either placebo or lisinopril (10 or 20 mg, depending on severity of hypertension). At baseline, 65% of the placebo group and 59% of the lisinopril group had some evidence of retinopathy. The control of blood

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pressure with lisinopril resulted in a 50% reduction in the number of patients whose retinopathy progressed one or more stages compared with the placebo group, after controlling for glycemic control (Chaturvedi N, 1997). It should be noted, however, that the results of this trial are a matter of some debate. The lisinopril-treated group had a signiÞcantly lower average HbA1c at baseline, and some researchers have postulated that its positive effect was due to lowering of undetected baseline hypertension (Fong DS, 2003). In a UKPDS interventional study, 1,148 hypertensive type 2 diabetics were randomized to tight blood pressure control (goal of 150/85 mm Hg or lower) with an ACE inhibitor or a beta blocker or to less tight control (goal of 180/105 mm Hg or lower). Tight blood pressure control was associated with a 35% lower risk of progression of diabetic retinopathy (two or more steps on the modiÞed ETDRS scale) during the median 7.5-year follow-up, and a 47% lower risk of visual acuity loss (three or more lines on the ETDRS chart). There was no signiÞcant difference in these effects between the two antihypertensive agents (UKPDS Group, 1998[b]). A similar study—the Appropriate Blood Pressure Control in Diabetes (ABCD) trial—randomized 470 hypertensive type 2 diabetics to intensive (goal of 75 mm Hg diastolic pressure) or moderate (goal of 80–89 mm Hg diastolic pressure) blood pressure control. At the end of the Þve-year follow-up, the data showed no signiÞcant difference between the groups in the progression of diabetic retinopathy (Estacio RO, 2000). Taken together, the available data (both observational and interventional) do not paint an entirely conclusive picture of the association between hypertension and diabetic retinopathy. However, most physicians and researchers believe that the link is highly plausible and that well-designed studies in the future will provide evidence of a clear causal relationship. Diabetic Nephropathy. Epidemiological data have shown that the incidence and progression of diabetic retinopathy is highly correlated with diabetic nephropathy, as measured by micro- and/or macroalbuminuria (Cruickshanks KJ, 1993; Klein R, 1993; West KM, 1980). In fact, some researchers believe microalbuminuria alone is a predictor of increased diabetic retinopathy risk. It is unlikely that diabetic nephropathy itself inßuences the development of retinopathy, but researchers believe hyperglycemia inßicts similar damage to kidneys and the retina. One hypothesis proposes that increased VEGF in both retinal and renal tissues is to blame, given that diabetics with retinopathy display increased VEGF levels in the eye, and elevated VEGF levels are often found in the urine of diabetics with proteinuria. CURRENT THERAPIES The only proven pharmacological therapy for diabetic retinopathy is the prophylactic achievement of near-normal glucose control. As described in “Etiology and Pathophysiology,” studies have found that rigorous glycemic control can prevent the development of nonproliferative retinopathy and delay progression to proliferative disease (DCCT Group, 1993; Gaede P, 1999; UKPDS Group, 1998[a]).

14

DIABETIC RETINOPATHY

TABLE 3. Current Therapies Used for Diabetic Retinopathy Agent

Company/Brand

ACE inhibitors Lisinopril AstraZeneca’s Zestril/Acerbon, Bristol-Myers Squibb’s Carace/Coric, Merck’s Prinivil, generics Captopril Bristol-Myers Squibb’s Lopirin/Capoten, ´ Sanofi-Synthelabo’s Alopresin, generics

Daily Dose

Availability

5–10 mg qd or bid

US, F, G, I, S, UK, J

25–50 mg qd or bid

US, F, G, I, S, UK, J

ACE = Angiotension-converting enzyme; bid = Twice daily; qd = Once daily. US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan.

Most physicians and other experts have consequently tightened glycemic control in retinopathy patients, particularly since the publication of the Diabetes Control and Complications Trial (DCCT) and United Kingdom Prospective Diabetes Study (UKPDS) results. Besides antidiabetic agents, only antihypertensive drugs have been shown to have a positive effect on the development and progression of diabetic retinopathy. Nevertheless, physicians do not routinely prescribe antihypertensives speciÞcally for the treatment of diabetic retinopathy. Many diabetics already take one or more antihypertensive agents, either for cardiovascular disease or renal dysfunction, at the time of retinopathy diagnosis. Most physicians are unwilling to prescribe antihypertensives to diabetics who are normotensive and have no discernible renal dysfunction. When antihypertensives are used in these patients, they are, almost without exception, angiotensin-converting enzyme (ACE) inhibitors; they are described here and in Table 3. ACE Inhibitors Overview. Since the 1997 publication of the EURODIAB Controlled Trial of Lisinopril in Insulin-Dependent Diabetes Mellitus (EUCLID, described later in the section “Lisinopril”), many physicians and researchers contend that strong evidence proves the beneÞcial effect of ACE inhibitors on the progression of diabetic retinopathy. This section highlights the agents in the class for which there are signiÞcant Þndings from high-quality clinical trials. Other ACE inhibitors that are also prescribed for diabetic retinopathy (and diabetics in general) include ramipril (Aventis’ Altace/Triatec, AstraZeneca’s Vesdil/Unipril), enalapril (Boehringer Ingelheim’s Pres, Merck’s Vasotec, generics), fosinopril (BristolMyers Squibb’s Monopril/Fosinorm, Merck’s Fozitec), and quinapril (PÞzer’s Accupril/Accupro, SanoÞ-Synth´elabo’s Korec). Mechanism of Action. ACE inhibitors lower blood pressure by inhibiting the vasoconstrictive action of the renin-angiotensin-aldosterone system (RAAS). Additionally, ACE inhibitors are known to have some action in preventing cell

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NH2 H2C H2C

CH2 H2C CH

HOOC

CH2

CH NH

CH2

N C O

COOH

FIGURE 4. Structure of lisinopril.

proliferation, reducing platelet aggregation, and enhancing Þbrinolysis (Lonn EM, 1998). Pharmacologically, ACE inhibitors act to prevent the ACE-mediated conversion of angiotensin I into angiotensin II (AII)—a potent vasoconstrictive agent that also promotes cardiovascular tissue growth and water and sodium retention. AII is also known to stimulate contraction of the vascular smooth-muscle cells lining the vascular wall, an action ultimately leading to hypertrophy (an increase in cell size) and hyperplasia (an increase in cell number). This action manifests as a thickening of the arterial wall and a narrowing of the lumen, developments that generate an increase in the peripheral resistance of the vasculature. AII also increases production of excess reactive oxygen species, which in turn increase vasoconstriction and damage the endothelial wall (Sowers JR, 2002). Lisinopril. Lisinopril (Merck’s Prinivil, AstraZeneca’s Zestril, generics) (Figure 4) was approved in the United States in December 1987 for use as an antihypertensive agent. The agent is now available in all seven markets under study. The EUCLID study enrolled 530 type 1 diabetics who were both normotensive and either normoalbuminuric (85%) or microalbuminuric (15%) from 15 European centers. At baseline, 65% of the placebo group and 59% of the lisinopril group had some evidence of retinopathy. Patients were randomized to either placebo or lisinopril (10 mg or 20 mg). After two years, treatment with lisinopril resulted in a 50% reduction in the number of patients whose retinopathy progressed one or more stages compared with the placebo group, after controlling for glycemic control (Chaturvedi N, 1997). It should be noted, however, that the results of this trial are a matter of some debate. The lisinopril-treated group had a signiÞcantly lower average HbA1c at baseline, and some researchers have postulated that the drug’s positive effect was due to lowering of undetected baseline hypertension (Fong DS, 2003). Lisinopril’s adverse events proÞle is similar to that of other ACE inhibitors. A common side effect of agents in this class is a dry, nonproductive cough, which can occur in 5–20% of patients. Independently of their action on the RAAS, ACE inhibitors reduce breakdown of the vasodilator bradykinin, accounting for the cough frequently associated with ACE inhibitor prescription. ACE inhibitors are

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DIABETIC RETINOPATHY

CH3 HSCH2CHC O

N COOH

FIGURE 5. Structure of captopril.

contraindicated in pregnancy (the FDA has rated this class of drugs as Pregnancy Category C [risk cannot be ruled out] in the Þrst trimester and Category D [published documentation of risk exists] for the second and third trimesters). ACE inhibitors are also contraindicated for patients with serious renal stenosis or widespread vascular lesions in the kidney due to potentially decreased renal perfusion. A further concern is angioedema (swelling and accumulation of ßuid in the deep layer of the skin and the connective tissue that underlies mucous membranes); angioedema occurs in approximately 0.1% of patients taking ACE inhibitors. Captopril. Captopril (Bristol-Myers Squibb’s Capoten, generics) (Figure 5) is available in all seven markets under study. In a UKPDS study, 1,148 hypertensive type 2 diabetics were randomized to tight blood pressure control (goal of blood pressure lower than 150/85 mm Hg) with either captopril or a beta blocker (atenolol; AstraZeneca’s Tenormin, generics) or to less tight control (goal of blood pressure lower than 180/105 mm Hg). Tight blood pressure control was associated with a 35% lower risk of progression of diabetic retinopathy (an increase of two or more steps on the modiÞed Early Treatment Diabetic Retinopathy Study [ETDRS] scale) during the median 7.5-year follow-up, and a 47% lower risk of visual acuity loss (an increase of three or more lines on the ETDRS chart). There was no signiÞcant difference in these effects between the two antihypertensive agents (UKPDS Group, 1998[b]). Like lisinopril, captopril is associated with a dry, unproductive cough. It is also known to cause rashes in 2–7% of treated patients, generally within the Þrst few weeks of treatment. Captopril is contraindicated in patients with renal insufÞciency or angioedema. Nonpharmacological Approaches Although tight control of hyperglycemia and hypertension are effective in slowing the progression of diabetic retinopathy, the modest beneÞts conferred by pharmacological agents are not sufÞcient to completely prevent neovascularization. For this reason, nonpharmacological approaches such as laser photocoagulation and surgery are the only effective means to prevent vision loss. Laser Photocoagulation. Photocoagulation destroys large sections of the hypoxic retina, thereby reducing the stimulus for abnormal vascular proliferation. Generally, photocoagulation involves placing a contact lens on a patient’s

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eye after dilation and then focusing a laser beam (argon or krypton) on the retina using a slit lamp. Two techniques are commonly used to treat diabetic eye disease: panretinal photocoagulation (also called scatter photocoagulation) and focal photocoagulation. Panretinal photocoagulation treats a large section of the retina, excluding the macular and foveal areas. Using a blue-green laser, physicians apply moderately intense burns for 0.1 second. Panretinal treatment usually consists of 1,200 to 1,500 of these burns, spaced a half-burn diameter apart from one another. Most patients undergo multiple sessions of 600 to 800 burns each. Panretinal treatment is indicated for proliferative retinopathy but may also be used in patients with severe preproliferative retinopathy, depending on the physician’s assessment of the need for aggressive treatment. In the Diabetic Retinopathy Study (DRS), in which 1,742 patients were randomly assigned to either argon or xenon laser treatment, panretinal photocoagulation reduced severe vision loss by 60% after three years. The same trial also showed a 90% reduction in blindness at Þve years after photocoagulation (Diabetic Retinopathy Study Research Group, 1978). Focal photocoagulation is often used for macular edema, a common complication of diabetic retinopathy, as well as for speciÞc discrete vascular abnormalities elsewhere on the retina of diabetic retinopathy patients. This technique employs the same laser used in panretinal photocoagulation but at a lower energy level (commonly an argon green-only laser). The laser is applied either in a grid pattern or directly to leaking microaneurysms to destroy the blood vessels. In the Early Treatment Diabetic Retinopathy Study (ETDRS), focal/grid photocoagulation reduced by approximately 50% the risk of moderate visual acuity loss (deÞned as doubling of visual angle) in patients with clinically signiÞcant macular edema (ETDRS Research Group, 1985; ETDRS Research Group, 1987). Photocoagulation therapy is generally safe, but it can produce serious side effects and complications in a minority of patients. For example, some patients lose their peripheral vision—in the DRS, 5% of argon-treated eyes and 25% of xenon-treated eyes experienced a constriction of visual Þeld to less than 45◦ but greater than 30◦ (DRS Research Group, 1981[b]). Patients may also develop night vision problems. Potentially more serious is the risk that the laser might burn the wrong part of the retina, causing further deterioration and/or loss of vision. Among argon-treated eyes in the DRS, 11% had a treatment-related persistent decrease in visual acuity of one line, while 3% had a persistent decrease of two or more lines. Xenon-treated eyes had a higher rate of treatment-related side effects—19% had a persistent decrease in visual acuity of one line, and another 11% had a persistent decrease of two or more lines (DRS Research Group, 1981[b]). Vitrectomy. Vitrectomy is indicated for media opacities (primarily vitreous hemorrhage), retinal traction detachment, and, rarely, traction-induced macular edema. A more substantial surgical undertaking than photocoagulation, vitrectomy usually requires hospitalization and general anesthesia. The goal of

18

DIABETIC RETINOPATHY

vitrectomy is removal of the posterior vitreous surface, which provides the “scaffolding” upon which neovascular tissue grows in diabetic retinopathy. The procedure involves making three incisions into the pars plana of the eye, the region behind the iris and in front of the retina. Three instruments are inserted into the incisions: a small light pipe to provide illumination; an infusion port to maintain proper ßuid balance during the procedure; and the vitrector, or cutting device. Once the instruments are positioned inside the eye, subhyaloid blood is extracted and the posterior two-thirds of the vitreous gel and posterior vitreous surface are excised. Fibrovascular tissue causing retinal traction detachment may also be excised by this method. Photocoagulation may be applied thereafter in a panretinal pattern. The Diabetic Retinopathy Vitrectomy Study (DRVS) conducted two randomized trials to assess the efÞcacy of early vitrectomy in type 1 and 2 diabetics and conducted an observational study of a third group of diabetics. One interventional arm of the trial enrolled 616 diabetic patients with severe vision loss from a recent vitreous hemorrhage (5/200 or less for one month or longer). Patients were randomized to either early vitrectomy or deferral of vitrectomy for one year (conventional therapy). The results showed that 25% of the early vitrectomy group recovered good vision (10/20 or better) at two years, compared with 15% of those who received the conventional therapy (DRVS Research Group, 1985). The second interventional arm randomized 381 patients with proliferative retinopathy who still had useful vision (10/200 or better) in at least one eye to either early vitrectomy or conventional therapy. In this arm, the early vitrectomy group had a 36% recovery rate at two years, versus 12% of the conventional group (DRVS Research Group, 1988). Although the Þndings of the DRVS and other studies show that vitrectomy can produce positive results, the procedure’s complicated nature has relegated its use only to cases of severe diabetic retinopathy when photocoagulation has failed or proved technically impossible. The operation puts patients at much higher risk for complications than photocoagulation does; the complications include the buildup of silicon ßuid in the eye, recurrent vitreous hemorrhage, retinal detachment, and anterior hyaloidal Þbrovascular proliferation (vessel proliferation at the front of the eye, behind the lens capsule). EMERGING THERAPIES Of all the agents in development for the treatment of the diabetic microvascular complications, the agents outlined in this section are potentially the most exciting. No current pharmacotherapy, with the possible exception of antihypertensives, can halt the progression of retinopathy. While effective, the current retinopathy treatments (photocoagulation or surgery) are appropriate only for patients who have progressed to the more severe, proliferative stage of disease, leaving few options other than tightened glycemic control for the treatment of patients with nonproliferative disease. Most pharmacotherapies in development target the aberrant angiogenesis that characterizes proliferative diabetic retinopathy, while some

EMERGING THERAPIES

19

are also being tested for related diabetic macular edema. The most interesting therapies in development for diabetic retinopathy are outlined in Table 4. Protein Kinase Cβ Inhibitors Overview. Protein kinase C (PKC) inhibitors are potential treatments for diabetic retinopathy. Eli Lilly’s PKCβ inhibitor, ruboxistaurin, was the Þrst potential oral drug therapy for the indication to reach late-stage development. However, recent clinical trial data have cast some doubt on the potential of this class of drugs. Mechanism of Action. The protein kinase C family consists of several structurally related enzyme isoforms that are present throughout the body. Experimental evidence has shown that the beta isoform (PKCβ) is a key mediator of diabetes-induced retinal abnormalities. When activated by diacylglycerol—cellular levels of which are increased by a chronic hyperglycemic state—PKCβ promotes the synthesis of vascular endothelial growth factor (VEGF), a potent promoter of angiogenesis known to play a role in inducing proliferative diabetic retinopathy (Frank RN, 2002). Agents that inhibit the activation and/or activity of PKCβ could therefore block the effects of VEGF, including promotion of vascular angiogenesis and hyperpermeability. Ruboxistaurin. Eli Lilly’s PKCβ inhibitor ruboxistaurin (LY-333531), an orally available agent, has been discontinued. For purely historical purposes, it is of interest to note that it was highly selective for the β isozyme from the PKC family, and did reach Phase III development in the United States and Europe. In early 2004, Takeda and Eli Lilly agreed to codevelop and comarket ruboxistaurin in Japan, where it was in Phase II. Ruboxistaurin was also in Phase III development for the treatment of diabetic peripheral neuropathy. The Þrst data published from Phase III trials of ruboxistaurin in diabetic retinopathy were not as promising as physicians and researchers had expected. The Protein Kinase C Diabetic Retinopathy Study (PKC-DRS) was a Phase II/III, multicenter, double-blind, placebo-controlled, randomized trial. Two hundred Þfty-two patients with either type 1 (19%) or type 2 (81%) diabetes were randomized to one of three ruboxistaurin doses (8, 16, or 32 mg daily) or placebo for a minimum of 36 months. At baseline, patients had either moderate to severe nonproliferative diabetic retinopathy: as judged by the severity of disease in the worst affected eye, 45% had retinopathy of Early Treatment Diabetic Retinopathy Study (ETDRS) severity level 47, and 55% were at level 53. The primary outcomes measured were progression of retinopathy by three or more steps on the ETDRS scale or application of photocoagulation. At 42 months, ruboxistaurin did not have any signiÞcant effect on those primary end points: event-rate estimates were 57%, 72%, and 52% in the 8, 16, and 32 mg groups, respectively, compared with 55% in the placebo group. However, the highest-dose ruboxistaurin group did show a trend toward a positive effect on some outcome measure. The rates of moderate visual loss (loss of 15 or more letters on the Snellen chart) were

20

DIABETIC RETINOPATHY

TABLE 4. Emerging Therapies in Development for Diabetic Retinopathy Compound

Development Phase

Protein kinase Cβ inhibitors Ruboxistaurin United States Europe Japan Ocular corticosteroid implants Fluocinolone acetonide (Retisert) United States Europe Japan Dexamethasone (Posurdex) United States Europe Japan

Marketing Company

III III I

Eli Lilly Eli Lilly Eli Lilly

III — —

Bausch & Lomb/Control Delivery Systems — —

IIIa — —

Allergan/Oculex Pharmaceuticals — —

Somatostatin analogues Octreotide (Sandostatin LAR) United States Europe Japan BIM-23190 United States Europe Japan

— IIIb —

— Novartis —

— PC PC

— Ipsen Teijin

Angiotensin II receptor antagonists Candesartan (Atacand) United States Europe Japan

III III III

AstraZeneca/Takeda AstraZeneca/Takeda AstraZeneca/Takeda

VEGF antagonists Pegaptanib (Macugen)c United States Europe Japan

II II —

Eyetech Pharmaceuticals/Pfizer Eyetech Pharmaceuticals/Pfizer —

Hyaluronidase modulators Hyaluronidase (Vitrase)d United States Europe Japan

PR III PC

Ista/Allergan Ista/Allergan Otsuka

a Phase III trials are being conducted in diabetic macular edema patients. b Phase III trials are being conducted in Switzerland. c Phase II trials are being conducted in diabetic macular edema patients. Pfizer and Eyetech have publicly

discussed the potential for additional diabetic retinopathy trials, but to date no such formal development program has been announced. d Vitrase is preregistered in the United States for the treatment of vitreous hemorrhage and as a dispersion agent for other ophthalmic drugs. It is in Phase III development in Europe and preclinical development in Japan for vitreous hemorrhage, and Phase II trials for diabetic retinopathy have been conducted in Mexico. PC = Preclinical (including discovery). PR = Preregistered.

EMERGING THERAPIES

21

lower in the 32 mg group than in the placebo group at 12 months (12% versus 20%), 24 months (8% versus 29%), and 36 months (19% versus 28%), with an average risk reduction of 35% (Milton R, 2003). The second Phase III trial of ruboxistaurin—the Protein Kinase C Diabetic Macular Edema Study (PKC-DMES)—measured the drug’s effect on diabetic macular edema (DME) and found similar results. Ruboxistaurin had no signiÞcant effect on either the progression of DME or the application of photocoagulation, but data revealed a trend, with a 32 mg daily dose, toward a positive effect on the development of DME that involves or imminently threatens the center of the macula (Eli Lilly, press release, August 25, 2003). One possible explanation for the largely disappointing results from these two Phase III trials (which used partially overlapping patient populations) was the lower-than-expected event rates in the placebo groups (Davis MD, 2003). The trials were designed based upon event-rate data from the ETDRS, where the rates of both diabetic retinopathy and DME events were notably higher than those found in the PKC-DRS or PKC-DMES. Thus, the PKC-DRS and PKCDMES were perhaps not powered sufÞciently to show a treatment effect, if any existed. Eli Lilly stated already in December 2002 that these results would delay the EU and U.S. regulatory Þlings for ruboxistaurin for diabetic retinopathy, but that additional Phase III studies were to be conducted that would, the company had hoped, address the methodological issues of the PKC-DRS. These hopes did not come to fruition. Ocular Corticosteroid Implants Overview. Corticosteroids have been utilized in eyedrop formulations for a number of ophthalmic indications. However, these products are unable to penetrate the vitreous and retinal areas and therefore have little effect on retinal vasculature and inßammation. Corticosteroid implants currently in development for diabetic macular edema and retinopathy may allow site-speciÞc delivery of these compounds, which have potent anti-inßammatory and antiangiogenic qualities. Mechanism of Action. Corticosteroids act as anti-inßammatory, immunosuppressive, and antiangiogenic agents through multiple effects: inhibiting synthesis of proinßammatory mediators (prostaglandins, leukotrienes, and cytokines); disrupting cellular activation, migration, and proliferation; and blocking edema formation. Although systemic corticosteroids are effective against many inßammatory and immune-mediated disorders, their prolonged use is associated with a high risk of side effects. Insomnia, night sweats, mood changes, and altered glucose metabolism may occur shortly after beginning corticosteroid use, while long-term use of systemic corticosteroids is associated with adrenal atrophy, osteoporosis, hypertension, cataracts, acne, abnormal fat deposition, and excessive hair growth. Local corticosteroid delivery in the form of ocular drops, injections, or sustained-release implants has been postulated as a therapeutic option that would target inßammation and neovascular processes in numerous

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retinal disorders—including diabetic retinopathy—in the hopes of minimizing the risk of systemic side effects. Fluocinolone Acetonide. Fluocinolone Intraocular implant has been discontinued. Originally, Bausch&Lomb and Control Delivery Systems were developing a sustained-release intravitreal ßuocinolone implant (Retisert) using Control Delivery Systems’ Envision TD delivery technology. The agent did reach Phase III development in the United States for diabetic macular edema and diabetic retinopathy, as well as Phase III trials for uveitis and Phase II trials for age-related macular degeneration (AMD). Twelve-month data from the Þrst Phase III trial demonstrating the ßuocinolone implants’ efÞcacy were presented at the 2003 annual meeting of the Association for Research in Vision and Ophthalmology (ARVO). Eighty patients with diabetic macular edema were randomized to ßuocinolone implant (0.5 mg or 2 mg) or standard of care (observation or laser photocoagulation). The 2 mg implant was discontinued early in the trial when investigators determined it conferred no additional treatment beneÞt compared with the 0.5 mg implant. After 12 months, the ßuocinolone implant had a signiÞcant effect on retinal thickness (macular edema)—49% of patients receiving the 0.5 mg implant had complete response, compared with 25% of the control group (Bausch & Lomb, press release, May 7, 2003). This trial was not powered to demonstrate signiÞcant effects on diabetic retinopathy or overall visual acuity, but those data were presented and suggested a trend toward an effect. Fewer ßuocinolone-treated patients demonstrated a worsening of diabetic retinopathy (5%) than did patients in the control group (30%). Although the results for visual acuity changes were not statistically signiÞcant, patients receiving the ßuocinolone implant did demonstrate a trend toward a treatment effect. More ßuocinolone-treated patients had stable (70%) or improved (19.5%; 15 or more letters) visual acuity than did control patients (50% and 7.1%, respectively). Although the 12-month data from the Phase III trial looked promising—particularly for the treatment of diabetic macular edema—ßuocinolone treatment was also associated with a number of side effects. Nearly 20% of ßuocinolone-treated patients had a serious increase in intraocular pressure (≥30 mm Hg), whereas no control-group patients did. Cataract progression at six months was evident in 55% of patients in the ßuocinolone-treated group, compared with no patients in the control group. After discussions concerning the adverse event rates from this trial, the FDA had requested additional 12-month safety data from additional ßuocinolone-implanted diabetic eyes. Eventually, these effects resulted in the discontinuation of the ßuocinolone intraocular implant. Dexamethasone. Oculex has developed a biodegradable, sustained-release dexamethasone implant (Posurdex) for the treatment of macular edema. In October 2003, Allergan purchased Oculex and began to design and plan Phase III

EMERGING THERAPIES

23

trials in the United States for diabetic macular edema and nondiabetic edemas (Allergan eyes Posurdex . . ., 2003). Preliminary data from a Phase II, randomized, dose-ranging trial of intravitreal dexamethasone were presented at the 2003 annual meeting of ARVO. The study enrolled patients with persistent macular edema associated with diabetes (n = 171), uveitis or Irvine-Gass syndrome (n = 40), or central or branch retinal vein occlusion (n = 103), all with visual acuity of 20/40 or worse. Patients were randomized to intravitreal dexamethasone (350 µg or 700 µg) or standard of care/observation (Haller JA, 2003; Kuppermann BD, 2003; Williams GA, 2003). EfÞcacy data were reported for a combined study population of 306 patients three months after treatment initiation. Patients who received the 700 µg demonstrated signiÞcant improvements in visual acuity (two or more lines) compared with the control group. Both the 350 µg and 700 µg intravitreal dexamethasone groups showed signiÞcant decreases in retinal thickness and ßuorescein leakage compared with control (Oculex, press release, May 8, 2003). Like the ßuocinolone implant, intravitreal dexamethasone was associated with an increased risk of glaucoma—4% of treated patients had developed elevated intraocular pressure in the three-month assessment. Although this rate is lower than reported in the previously described trial of the ßuocinolone implant, it is premature to assume that intravitreal dexamethasone will have a more favorable adverse event proÞle. Preliminary data regarding the ßuocinolone implant also demonstrated a low occurrence of serious side effects, and not until longerterm data were available did the serious adverse event rate become substantial (approximately 40–50%). According to Allergan, the intended Phase III development program for intravitreal dexamethasone will include approximately 700 patients (diabetic and nondiabetic), with treatment periods of at least six months and follow-up periods of a year or more. Somatostatin Analogues Overview. The normal process of retinal vascular growth and apoptosis involves both proangiogenic and antiangiogenic elements. In an attempt to capitalize on the body’s naturally antiangiogenic compounds, researchers are developing analogues of somatostatin, a naturally occurring hormone with strong inhibitory effects on the endocrine system and possibly on the retinal vasculature speciÞcally. Mechanism of Action. Somatostatin is an endogenous hormone that is secreted in a number of body tissues, including the hypothalamus, intestines, and pancreas. It has a potent and multifaceted inhibitory effect on the endocrine system—endogenous somatostatin release inhibits the secretion of growth factors such as insulin, thyroid-stimulating hormone (TSH), human growth hormone (GH), and insulin-like growth factor 1 (IGF-1). Case studies and other experimental evidence have demonstrated that GH and IGF-1, in particular, are angiogenic factors that contribute to retinal neovascularization in diabetics. Some diabetics

24

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f C F W K T C T–ol FIGURE 6. Structure of octreotide.

who have undergone hypophysectomy (removal of the pituitary gland) experienced reversals of proliferative retinopathy that correlated with the degree of GH deÞciency postsurgery (Patterson JH, 1974; Poulsen JE, 1953; Sharp PS, 1987). Serum and vitreal concentrations of IGF-1 correlate positively with the presence of proliferative retinopathy in diabetics (Burgos R, 2000; Merimee TJ, 1983). Thus, by mimicking the inhibitory effects of endogenous somatostatin and blocking the secretion of GH, IGF-1, and other growth factors, somatostatin analogues may be effective against neovascularization in the diabetic retina. Octreotide. Novartis’s octreotide (Figure 6) is a somatostatin analogue marketed as Sandostatin and as Sandostatin LAR, a long-acting release formulation, worldwide for the treatment of acromegaly and diarrhea. The long-acting formulation is in Phase III trials in Switzerland for diabetic retinopathy. A 15-month pilot study of 23 patients with type 1 or type 2 diabetes and severe nonproliferative diabetic retinopathy was conducted in the United States. Octreotide-treated patients received four daily subcutaneous injections of shortacting octreotide at the maximum tolerable dose for each individual (total daily doses ranged from 200 µg to 5,000 µg). The number of patients requiring panretinal photocoagulation (PRP) during the study period was signiÞcantly reduced in the octreotide-treated group: 1 of 22 eyes required PRP compared with 9 of 24 eyes in the control group. In addition, the incidence of progression to require PRP (ETDRS score 71 or 75) was 42% in the control group compared with 9% in the octreotide group, although those results were not statistically signiÞcant (Grant MB, 2000). In a small U.K. trial, 18 patients (type 1 or type 2 diabetics) with persistent proliferative retinopathy after laser photocoagulation were randomized to either octreotide treatment (n = 9) or a control group (no placebo; n = 9). Octreotidetreated patients received three times daily subcutaneous 100 µg injections of short-acting octreotide. After three years of treatment, octreotide reduced the risk of vitreous hemorrhage and vitreoretinal surgery signiÞcantly compared with control. Average visual acuity remained stable in the octreotide-treated patients but decreased in the control group (Boehm BO, 2001). The currently marketed short-acting product is an intramuscular injection that requires physician administration, and trials using this formulation of octreotide in retinopathy have used thrice-daily injections. The ongoing Phase III diabetic retinopathy trial is utilizing the sustained-release octreotide LAR, and the dosing protocol in the study will use monthly injections. However, octreotide LAR is an intramuscular injection that requires in-ofÞce administration by a physician, so it may not offer a large convenience beneÞt over daily but self-administered injections of the short-acting octreotide product.

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BIM-23190. Ipsen’s BIM-23190 is a somatostatin analogue that is highly selective for the human somatostatin analogue receptor subtype 2 (Morgan JP, 1996). In July 2003, Teijin signed an exclusive codevelopment and comarketing agreement with Ipsen for BIM-23190 in Japan for the treatment of diabetic retinopathy. No human trial data are yet published that would allow for direct comparison of BIM-23190’s efÞcacy and safety with those of octreotide, but it appears that BIM-23190 will also be administered parenterally. Angiotension II Receptor Antagonists Overview. Since the 1997 publication of the EURODIAB Controlled Trial of Lisinopril in Insulin-Dependent Diabetes Mellitus (EUCLID, described in the “Current Therapies” section), antihypertensives (speciÞcally, ACE inhibitors) have been posited to have a strong effect on the progression of diabetic retinopathy. However, EUCLID was not designed with incidence or progression of diabetic retinopathy as primary outcomes, leaving some uncertainty about antihypertensives’ effects on the disease (Sjolie AK, 2002). Another question remains as well: Will agents that produce the same antihypertensive effect as ACE inhibitors via different mechanisms be comparable or superior to those agents in the treatment of diabetic retinopathy? To that end, angiotensin II receptor antagonists (AIIRAs) are under investigation as potential therapies for this indication. Mechanism of Action. Angiotensin II (AII) is a potent vasoconstrictive agent that also promotes cardiovascular tissue growth and water and sodium retention. AII is also known to stimulate contraction of the vascular smooth-muscle cells lining the vascular wall, an action ultimately leading to hypertrophy (an increase in cell size) and hyperplasia (an increase in cell number). This action manifests as a thickening of the arterial wall and a narrowing of the lumen, developments that generate an increase in the peripheral resistance of the vasculature. AII also increases production of excess reactive oxygen species, which in turn increase vasoconstriction and damage the endothelial wall (Sowers J, 2002). AIIRAs, like ACE inhibitors, act on the renin-angiotensin-aldosterone system (RAAS) to block the activity of AII; however, AIIRAs selectively antagonize the angiotensin II receptor subtype 1 (AT1), whereas ACE inhibitors function by blocking AII generation. Data from animal models have shown that AII can stimulate retinal neovascularization in vitro (Otani A, 1998) and that AIIRA treatment can inhibit retinal VEGF expression and angiogenesis in vivo (Moravski CJ, 2000; Nagisa Y, 2001). Candesartan. Candesartan (AstraZeneca’s Atacand/Ratacand/Amias, Takeda’s Kenzen/Blopress) (Figure 7) is marketed in all regions under study for the treatment of hypertension and is currently in Phase III trials for diabetic retinopathy. A large-scale Phase III diabetic retinopathy trial of candesartan was initiated in early 2003. The Diabetic Retinopathy Candesartan Trial (DIRECT) is a multicenter, placebo-controlled, double-blind, randomized clinical trial consisting of

26

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Me

N O N O

OH HN N

N N

FIGURE 7. Structure of candesartan.

three study arms. The three arms of the DIRECT study are a primary prevention trial in type 1 diabetics with no retinopathy, a secondary prevention trial in type 1 diabetics with nonproliferative retinopathy, and a secondary prevention trial in type 2 diabetics with nonproliferative retinopathy. Study investigators plan to enroll approximately 4,500 participants from 20 countries who will be randomized to either candesartan (up to 32 mg/day) or placebo for at least three years. The primary endpoints to be measured in DIRECT are incidence of retinopathy in those with no retinopathy at baseline (a two-step increase in ETDRS severity grade) and progression of existing retinopathy (a three-step or greater increase in ETDRS severity grade) (Chaturvedi N, 2002). According to AstraZeneca and Takeda, full results of the DIRECT study are expected by 2006. VEGF Antagonists Overview. Although many agents in development inhibit the production of vascular endothelial growth factor (VEGF), research has also focused on agents that can antagonize circulating ocular VEGF. The majority of VEGF antagonists are in active development for retinal neovascularization disorders such as AMD. However, by virtue of the common VEGF-mediated pathways that result in diabetic retinopathy and/or diabetic macular edema, these indications may be logical additional targets for those same anti-VEGF agents. Mechanism of Action. Unlike antiangiogenesis agents, which inhibit the production of VEGF through enzymatic or other processes, VEGF antagonists are a heterogeneous group of drugs in development that bind to free VEGF and render it unable to activate receptors in the retinal vasculature (or theoretically in any body tissue). Agents that can be classiÞed into this group include receptor fusion proteins, anti-VEGF aptamers, and monoclonal antibodies. Though different in composition and structure, all VEGF antagonists share the ability to mimic endogenous VEGF receptors and thus “capture” the molecule and render it inactive. Pegaptanib. PÞzer and Eyetech are jointly developing pegaptanib (EYE-001; Macugen), an intravitreal injection in Phase III trials for the treatment of

EMERGING THERAPIES

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AMD and Phase II trials for diabetic macular edema. Pegaptanib is an antiVEGF aptamer, a synthetic oligonucleotide with high afÞnity and selectivity for VEGF165. Eyetech originally licensed pegaptanib from Gilead in 2000, and in December 2002, PÞzer and Eyetech entered a codevelopment and comarketing agreement for the molecule. Under this agreement, PÞzer will fund most of the remaining development, and the companies will copromote pegaptanib in the United States. PÞzer has exclusive marketing rights outside the United States. Pegaptanib has not been tested in diabetic retinopathy patients, but Phase IIa data in AMD patients were presented in May 2002 at the TIDES meeting, a joint event that included the Sixth International Oligonucleotide Technology Conference and the Fifth International Peptide Technology Conference. Patients received three monthly intravitreal injections (0.1 mL) of pegaptanib either alone or in conjunction with photodynamic therapy (PDT). At three months, 86% of pegaptanib-treated patients demonstrated stabilization of AMD progression, compared with 51% of patients receiving PDT alone. Three-line improvements in visual acuity were evident in 26% of pegaptanib-treated patients, compared with 2% of PDT-treated patients. The combined effect of pegaptanib and PDT was even more beneÞcial: 60% of patients receiving combination therapy achieved a three-line or better improvement in visual acuity (Scypinski S, 2002). These Phase II AMD data suggest that after just three months, pegaptanib can inhibit the progression of retinal neovascularization. These results are promising for the use of this anti-VEGF aptamer to halt or possibly reverse retinal neovascularization associated with other conditions, including diabetic retinopathy. The general transferability of pegaptanib’s AMD trial results to diabetic retinopathy remains to be proven, however, because of key differences in the pathology and pathogenesis of each disease. The pathology of AMD neovascularization is distinct (the former occurs in the choroid, a normally avascular portion of the eye posterior to the retina), and the natural history of AMD progression follows a more rapid, progressively degenerative course than does the metabolic-dependent progression of diabetic retinopathy. Hyaluronidase Modulators Overview. Vitreous hemorrhage (leakage of the vitreous humor into the retina) is a complication of retinopathy that is difÞcult to treat and can occlude the imaging of the retina for screening or laser photocoagulation. The current treatment method, surgical vitrectomy, is generally effective but highly invasive. Hyaluronidase modulators, which are preparations of a naturally occurring enzyme, offer promise as treatment for vitreous hemorrhage secondary to proliferative diabetic retinopathy and may also be effective for preventing progression of nonproliferative disease. Mechanism of Action. Hyaluronidase is a naturally occurring enzyme that digests proteoglycans. This group of molecules, which are largely present in connective tissues, includes hyaluronan, hyaluronic acid, and chondroitin sulfate. When introduced into a medium containing proteoglycans, hyaluronidase digests

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the molecules and decreases the viscosity of the medium. Research has shown that intraocular injections of hyaluronidase can induce posterior vitreous detachment (PVD; a loosening or separation of the vitreous humor from the retina), which allows for the clearing of vitreous hemorrhage. Researchers postulate that liquefaction or detachment of the vitreous humor will remove the “scaffolding” upon which retinal neovascular tissue forms and therefore may prevent or delay progression to proliferative retinopathy. Hyaluronidase. Ista Pharmaceuticals is developing an intravitreal injection of highly puriÞed ovine hyaluronidase (Vitrase) for the treatment of vitreous hemorrhage and diabetic retinopathy and as a spreading agent to facilitate the dispersion and absorption of other ophthalmic agents. Ista partnered with Allergan in March 2000 to comarket hyaluronidase in the United States and all international markets except Mexico and Japan. In late 2001, Ista gave exclusive development and marketing rights for hyaluronidase in Japan to Otsuka. Hyaluronidase is currently in preregistration in the United States and Japan and in Phase III development in Europe for the treatment of vitreous hemorrhage. The FDA gave hyaluronidase fast-track designation in 1998 for this indication. Ista submitted hylauronidase for FDA approval in December 2002 for the treatment of vitreous hemorrhage but received an approvable letter in April 2003 citing insufÞcient statistical evidence to support its approval. In August 2003, Ista Þled a second NDA for hylauronidase for approval as a dispersion enhancer for other ophthalmic agents. The FDA has accepted this second NDA. In addition to these development programs, Phase II trials for diabetic retinopathy have been conducted in Mexico. Initial results from a randomized, placebo-controlled, Phase IIa trial of hyaluronidase showed it was effective at inducing PVD. The study enrolled approximately 60 diabetics with nonproliferative retinopathy from Mexico City. Patients were randomized to receive one of four treatments: an injection of hyaluronidase, a placebo (saline) injection, a single treatment with sulfurhexaßuoride gas (SF6; a surgical adjunct for retinal detachment treatment), or a single, combination treatment with SF6 and a hyaluronidase injection. Interim results showed that 16 weeks following treatment, 60% of patients who received hyaluronidase injections demonstrated complete PVD, as measured via ultrasound. Complete PVD rates were 53% in the SF6-treated group, 50% in the SF6/hyaluronidase combination group, and 6% in the placebo group (Ista, press release, October 19, 2000). The full one-year data measured the effect of hyaluronidase on ETDRS diabetic retinopathy levels and found that the agent prevented worsening of diabetic retinopathy. At 12 months, 67% of hyaluronidase-treated patients had stable ETDRS scores, compared with 40%, 43%, and 38% of the SF6-treated group, the SF6/hyaluronidase combination group, and the placebo group, respectively. Fewer hyaluronidase-treated patients (13%) had a worsening of ETDRS severity, compared with 20% of the SF6-treated group, 21% of the SF6/hyaluronidase combination group, and 38% of the placebo group (Ista, press release, January 8, 2002).

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Garancini P, et al. Prevalence of retinopathy in diabetic subjects from out-patient clinics in Lombardy (Italy) and associated risk factors. Diabetes Research and Clinical Practice. 1989;6:129–138. Gardner TW, Aiello LP. Pathogenesis of diabetic retinopathy. In: Flynn HW, Smiddy WE, eds. Diabetes and Ocular Disease: Past, Present, and Future Therapies. San Francisco: The Foundation of the American Academy of Ophthalmology; 2000. Gardner TW, et al. Diabetic retinopathy: more than meets the eye. Survey of Ophthalmology. 2002;47(suppl 2):S253–S262. Grant MB, et al. The efÞcacy of octreotide in the therapy of severe nonproliferative and early proliferative diabetic retinopathy. Diabetes Care. 2000;23(4):504–509. Grusser M, et al. [Preventive care for early detection of diabetes mellitus complications: a model project in Wolfsburg.] Zeitschrift f¨ur Arztliche Fortbildung und Qualit¨atssicherung. 2000;94:411–416. Haller JA, et al. Treatment of persistent macular edema associated with central and branch retinal vein occlusion with extended delivery of intravitreal dexamethasone. Presented at the 2003 annual meeting of the Association for Research in Vision and Ophthalmology. May 4–9, 2003; Fort Lauderdale, FL. Harris MI, et al. Is the risk of diabetic retinopathy greater in non-Hispanic blacks and Mexican Americans than in non-Hispanic whites with type 2 diabetes? A U.S. population study. Diabetes Care. 1998;21:1230–1235. [a] Harris MI, et al. Prevalence of diabetes, impaired fasting glucose, and impaired glucose tolerance in U.S. adults: The Third National Health and Nutrition Examination Survey, 1988–1994. Diabetes Care. 1998;21:518–524. [b] Henricsson M, et al. Mortality in diabetic patients participating in an ophthalmological control and screening programme. Diabetic Medicine. 1997;14(7):576–583. Hesse L, et al. Population-based study of diabetic retinopathy in Wolfsburg. Opthalmologie. 2001;98:1065–1068. Hoogeveen EK, et al. Hyperhomocysteinemia is associated with the presence of retinopathy in type 2 diabetes mellitus: the Hoorn study. Archives of Internal Medicine. 2000;160:2984–2990. International Council on Ophthalmology. International clinical diabetic retinopathy disease severity scale. October 2002. www.icoph.org/standards/pdrdetail.html. Accessed September 25, 2003. Joner G, et al. A nationwide cross-sectional study of retinopathy and microalbuminuria in young Norwegian type 1 (insulin-dependent) diabetic patients. Diabetologia. 1992;34:1049–1054. Kato S; Retinopathy in older patients with diabetes mellitus. Diabetes Research and Clinical Practice. 2002; 58:187–192. Keeffe JE, et al. Impact of vision impairment on functioning. Australian and New Zealand Journal of Ophthalmology. 1998;26(suppl 1):S16–S18. Khaleeli AA, et al. Diabetic retinopathy: outcome at Þve-year follow-up of 203 people with diabetes. Practical Diabetes International . 1999:16(3):68–70. Klein R, et al. The Wisconsin Epidemiologic Study of Diabetic Retinopathy, part II: prevalence and risk of diabetic retinopathy when age at diagnosis is less than 30 years. Archives of Ophthalmology. 1984;102:520–526. [a]

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Klein R, et al. The Wisconsin Epidemiologic Study of Diabetic Retinopathy, part III: prevalence and risk of diabetic retinopathy when age at diagnosis is 30 or more years. Archives of Ophthalmology. 1984;102:527–532. [b] Klein R, et al. An alternative method of grading diabetic retinopathy. Ophthalmology. 1986;93:1183–1187. Klein R, et al. Glycosylated hemoglobin predicts the incidence and progression of diabetic retinopathy. Journal of the American Medical Association. 1988;260:2864–2871. Klein R, et al. The incidence of macular edema. The Wisconsin Epidemiologic Study of Diabetic Retinopathy XI. Ophthalmology. 1989;96:1501–1510. Klein R, et al. Epidemiology of diabetic retinopathy. Diabetes Care. 1992;15(12): 1875–1891. Klein R, et al. Is gross proteinuria a risk factor for the incidence of proliferative diabetic retinopathy? Ophthalmology. 1993;100:862–867. Klein R, et al. Relationship of hyperglycemia to the long-term incidence and progression of diabetic retinopathy. Annals of Internal Medicine. 1994;154:2169–2178. Klein R, et al. The Wisconsin Epidemiologic Study of Diabetic Retinopathy, part XVII: the 14-year incidence and progression of diabetic retinopathy and associated risk factors in type 1 diabetes. Ophthalmology. 1998;105:1801–1815. Klein R, et al. The NEI-VFQ-25 in people with long-term type 1 diabetes mellitus. Archives of Ophthalmology. 2001;119:733–740. Klein R, et al. The association of atherosclerosis, vascular risk factors, and retinopathy in adults with diabetes: the Atherosclerosis Risk in Communities Study. Ophthalmology. 2002;109:1225–1234. Klein R, et al. Has the frequency of proliferative diabetic retinopathy declined in the U.S.? Diabetes Care. 2003;26:2691–2692. Klein R, Klein BEK. Epidemiology of eye disease in diabetes. In: Flynn HW, Smiddy WE, eds. Diabetes and Ocular Disease: Past, Present, and Future Therapies. San Francisco: The Foundation of the American Academy of Ophthalmology; 2000. Kostraba JN, et al. The Epidemiology of Diabetes Complications Study, part IV: correlates of diabetic background and proliferative retinopathy. American Journal of Epidemiology. 1991;133:381–391. Koya D, King GL. Protein kinase C activation and the development of diabetic complications. Diabetes. 1998;47:859–866. Kozak LJ, et al. National Hospital Discharge Survey: 2000 Annual Summary with detailed diagnosis and procedure data. National Center for Health Statistics. Vital Health Statistics. 13(153). 2002. Kuppermann BD, et al. An intravitreous dexamethasone bioerodible drug delivery system for the treatment of persistent macular edema. Presented at the 2003 annual meeting of the Association for Research in Vision and Ophthalmology. May 4–9, 2003; Fort Lauderdale, FL. Kuzuya T, et al. Prevalence of chronic complications in Japanese diabetic patients. Diabetes Research and Clinical Practice. 1994;24(suppl):S159–S164. Laakso M, et al. Age of onset and type of diabetes. Diabetes Care. 1985;8:114–117. Lee PP, et al. Longitudinal prevalence of major eye diseases. Archives of Ophthalmology. 2003;121:1303–1310.

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Dyslipidemia

ETIOLOGY AND PATHOPHYSIOLOGY Introduction The term dyslipidemia refers to any abnormality in circulating lipid levels. This abnormality can include an elevation in any one of the lipid subfractions of lowdensity lipoprotein (LDL), very-low-density lipoprotein (VLDL), or triglycerides (TG). The term additionally encompasses any reduction in the circulating levels of high-density lipoprotein (HDL). The manifestation of dyslipidemia is generally deÞned according to the cholesterol levels set forth by the U.S. Third Report of the National Cholesterol Education Program (NCEP) Expert Panel on Detection, Evaluation, and Treatment of High Blood Cholesterol in Adults (Adult Treatment Panel III [ATP III]), published in 2001. According to ATP III guidelines, a total serum cholesterol (TSC) level of 200–239 mg/dL is considered “borderline high,” and a TSC level greater than or equal to 240 mg/dL is considered “high.” Such lipid abnormalities are known risk factors for atherogenesis, coronary heart disease (CHD), and cardiovascular disease (CVD), and can result from accelerated synthesis or from retarded degradation of lipoproteins. (Note that CHD is a subcategory of CVD, which encompasses all diseases that affect the heart and blood vessels, including ischemic stroke and peripheral arterial disease.) Lipid Pathways and Mechanisms Overview. To fully understand the development of dyslipidemia, familiarity with lipid pathways and mechanisms is essential. Cholesterol and TGs are the Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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DYSLIPIDEMIA

FIGURE 1. Cholesterol biosynthesis.

two key lipids transported by blood plasma. Cholesterol, which is used by adrenal glands and sex organs to manufacture steroid hormones, is present in all cell membranes. The majority of cholesterol synthesis in adults occurs in the liver, gut, and central nervous system. Cholesterol derives from acetyl-coenzyme A (CoA); the pivotal, nonreversible step in its synthesis is the formation of mevalonate from 3-hydroxy-3-methylglutaryl CoA (HMG-CoA). Figure 1 depicts the cholesterol biosynthesis sequence. The enzyme responsible for this step, HMG-CoA reductase, can be inhibited by a variety of physiological factors, the most inßuential being the intracellular level of cholesterol. The enzyme is also the target for the most popular class of cholesterol-lowering drugs, the HMG-CoA reductase inhibitors, also known as statins. The major breakdown products of cholesterol are bile acids, which are highly effective detergents that are involved in the solubilization of dietary lipids. Bile acid action increases the surface area of lipids, promoting hydrolysis by lipases and facilitating absorption in the intestine. Because of their role in lipid solubilization, bile acids are the target site for bile acid sequestrants, another class of antidyslipidemic therapeutics (see “Current Therapies”). Plasma TGs are derived from dietary fat or synthesized in the body from other energy sources, such as carbohydrates. Stored in adipose (fat) cells until needed, TGs consist of glycerol combined with three fatty acids.

ETIOLOGY AND PATHOPHYSIOLOGY

39

FIGURE 2. Lipoprotein structure.

Types of Lipoprotein and Lipoprotein Components. In plasma, proteinlipid particles called lipoproteins carry cholesterol, TGs, phospholipids (lipids that have been phosphorylated), and apolipoproteins (proteins that combine with lipids to form lipid-protein complexes). More than half of the coronary heart disease (CHD) in the United States is attributable to abnormalities in the levels and metabolism of plasma lipids and lipoproteins (Ginsberg HN, 2001). Figure 2 illustrates the structure of lipoprotein. The lipid content of lipoproteins, mainly cholesteryl esters and phospholipids, increases with age in all healthy arteries. At birth, cholesterol levels average 1.5 mmol/L (60 mg/dL); by the Þrst year, they rise to 4.5 mmol/L (175 mg/dL). A second rise begins after age 20 and continues to age 50–60. Six classes of lipoproteins have been identiÞed: chylomicrons, chylomicron remnants, VLDL, intermediate-density lipoprotein (IDL), LDL, and HDL. These various types differ from one another in terms of size, density, and the amount of cholesterol, TG, phospholipid, and apolipoprotein they contain. Figure 3 illustrates the composition of the four lipoproteins most often associated with dyslipidemia (chylomicron, VLDL, LDL, and HDL). Each lipoprotein, discussed in the following sections, performs a speciÞc function in terms of the type of lipid it transports and where it is transported to. TGs, also discussed in more detail, are

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DYSLIPIDEMIA

FIGURE 3. Composition of lipoproteins.

another lipoprotein component that plays a signiÞcant role in lipid metabolism and dyslipidemia. Chylomicrons. Chylomicrons transport exogenous TGs (TGs absorbed from the gut). They typically appear only transiently in the blood after a fatty meal, but defects in lipid metabolism, usually genetic in origin, may lead to chylomicrons and chylomicron remnants being present in plasma in the fasting state. Chylomicron remnants—a by-product of chylomicron lipolysis in the liver—transport exogenous cholesterol. They are considered to have only some atherogenic potential because they do not appear in the absence of other underlying lipid abnormalities, such as high serum TG levels. Very-Low-Density Lipoprotein Cholesterol. VLDLs are small, cholesteroland TG-rich lipoproteins synthesized by the liver that account for 10–15% of TSC. VLDLs are responsible for the transport of endogenous TGs (newly synthesized TGs). A large body of evidence suggests that VLDL and VLDL remnants play an important role in atherogenesis (Sacks FM, 2000; Thompson GR, 1998). Serum levels of VLDL are usually combined with LDL to give a measure of nonHDL cholesterol and are therefore treated alongside LDL abnormalities. ATP III identiÞes elevated VLDL levels as the surrogate for elevated atherogenic remnants in people with TGs ≥200 mg/dL.

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Low-Density Lipoprotein Cholesterol. LDLs transport cholesterol to tissues and typically make up 60–70% of TSC. A strong and graded positive association exists between LDL cholesterol and risk of CVD events over a wide range of cholesterol concentrations, as identiÞed in the ATP III guidelines. The association applies to patients without CVD as well as to patients with established CVD, and other factors modify the risk considerably. Even with moderately elevated LDL levels (130–159 mg/dL), factors such as low HDL and nonlipid risk factors can increase the potential for CVD events. (See the “Secondary Dyslipidemia” section for more on nonlipid risk factors.) The results of epidemiological studies and of trials with angiographic or clinical end points conÞrm the importance of LDL as a cause of atherosclerosis. Reduction of LDL must therefore be a primary concern in both primary and secondary prevention of CVD. High-Density Lipoprotein Cholesterol. Researchers have found an inverse association between plasma HDL and the risk of CVD (Assman G, 1992). This phenomenon has been documented for both men and women and for both asymptomatic people and patients with conÞrmed CVD. Low HDL is considered an independent risk factor for CVD; the 2004 update of the ATP III guidelines places increased emphasis on treating low HDL levels (<40 mg/dL) (Grundy SM, 2004). Low HDL is also an increasingly popular clinical study target for prevention of CVD (e.g., Whitney EJ, 2005). However, the relationship between HDL and CVD is not entirely understood. Concentrations of HDL tend to be low when TG concentrations are high, and HDL may, to a large extent, be a reciprocal measure of atherogenic lipoproteins such as VLDL. HDL actively transports cholesterol out of foam cells in an atherosclerotic plaque and delivers it to the liver, where HDL particles are selectively taken up. Studies suggest that the antioxidant and anti-inßammatory properties of HDL may protect against atherosclerosis. Low HDL levels in the absence of other lipid abnormalities are rare but may occur in patients with type 2 diabetes or metabolic syndrome (see the “Secondary Dyslipidemia” section). Low plasma HDL concentrations may also identify people with atherogenic risk factors; HDL is relatively low in smokers, obese people, and sedentary persons. Results from a meta-analysis of 17 major lipid interventional trials in more than 44,000 patients, presented at the 2004 American Heart Association (AHA) meeting, reveal that HDL levels are a more accurate predictor of cardiovascular event reduction than LDL, TSC, or TGs (Alsheikh-Ali AA, 2004). Large-scale prospective trials are needed to clarify the future role of HDL-targeted therapy. Triglycerides. Research suggests that TGs may play a more important role in CVD than previously recognized. These fats are a component of atherosclerotic plaques and may contribute to atherosclerosis by promoting coagulation and interfering with Þbrinolysis. Serum TG levels can vary from 50 mg/dL to more than 500 mg/dL but are typically less than 100 mg/dL. The NCEP ATP III considers levels below 150 mg/dL to be desirable. Meta-analyses and epidemiological studies have more clearly identiÞed elevated levels of TGs as an independent

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risk factor for CVD (Assmann G, 1998; Austin MA, 1998). ATP III considers elevated TGs to be a marker for other lipid and nonlipid abnormalities and a target for lipid-lowering therapy. Lipoprotein Generation and Transport. The pathophysiological consequences of lipid abnormalities can ultimately be traced to malfunctions in the lipoprotein generation and transport system. Lipid transport is divided into exogenous and endogenous pathways, which are regulated by speciÞc apolipoproteins, lipoprotein receptors, lipolytic enzymes, and transfer proteins. Each of these components plays a vital role in maintaining the balance of cholesterol and TGs in tissues and plasma. Figure 4 illustrates exogenous and endogenous lipid transport from the intestine to the plasma after being absorbed from dietary fat. Note that it is important to distinguish between the TG- and cholesterol-rich remnant lipoproteins (exogenous lipids), which are elevated in combined dyslipidemia (the condition in which both TGs and cholesterol are elevated), and the cholesterol-rich LDL lipids (endogenous lipids), which are elevated in isolated hypercholesterolemia. Exogenous Pathway. Approximately two-thirds of the cholesterol absorbed from the intestine is derived from bile; the other third is derived from the diet. Solubilized cholesterol, in the form of micelles, is absorbed at the brush border of enterocytes lining the intestine and taken into the cytoplasmic compartment. Internalized cholesterol is transferred to the endoplasmic reticulum, where it is esteriÞed by acyl-CoA:cholesterol acyltransferase (ACAT) to form cholesteryl ester. Free cholesterol, cholesteryl esters, and TGs are packaged into chylomicrons, which are secreted and transported to the liver. When chylomicrons reach adipose tissue, they adhere to speciÞc receptors on the capillary wall. TGs are then hydrolyzed by lipoprotein lipase, a critical step in the breakdown of plasma TG. Following lipolysis (the decomposition of fat), some of the chylomicron surface is shed, forming HDL. Alternatively, chylomicrons exchange TGs for cholesterol from mature HDL, a process mediated by the enzyme cholesteryl ester transfer protein (CETP). The remaining exogenous lipids (mainly cholesterol) are carried in chylomicron remnants to the liver, where they bind to chylomicron remnant receptors via the binding ligand apolipoprotein E (ApoE). The surface-bound process remnants are taken into hepatocytes by receptor-mediated endocytosis and degraded by lysosomes. Some of the cholesterol reaching the liver is converted to bile acid; the remainder is excreted, unmetabolized, into the bile. Endogenous Pathway. The liver converts carbohydrates from the diet into fatty acids, which are esteriÞed with glycerol to form TGs. TGs are then transported to tissue capillaries in VLDLs. Once they reach the capillaries, TGs are hydrolyzed by the enzyme lipoprotein lipase, and the TG remnants are then transported by IDLs, which have one of two fates: they are either catabolized by the liver following binding to LDL receptors on the surface of hepatocytes, or they remain in the plasma. Plasma IDLs undergo further transformation, removing TGs and forming cholesterol-rich LDL.

ETIOLOGY AND PATHOPHYSIOLOGY

43

FIGURE 4. Plasma triglyceride and cholesterol transport.

One function of LDL is to supply cholesterol to a variety of extrahepatic cells (e.g., lymphocytes, renal cells), which also have LDL receptors on their surfaces. The cholesteryl esters of LDL are hydrolyzed by a lysosomal cholesteryl esterase (acid lipase), and the cholesterol released is used for membrane synthesis and as a precursor to steroid hormone synthesis. LDL that is not hydrolyzed by the extrahepatic cells is degraded via a scavenger cell system, which releases unesteriÞed cholesterol into the plasma. UnesteriÞed cholesterol Þrst binds to

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DYSLIPIDEMIA

TABLE 1. The Frederickson/WHO Classification of Dyslipidemia Phenotype Classification Type I Type IIA Type IIB Type III Type IV Type V

Lipids Increased

Lipoproteins Increased

Triglycerides Cholesterol Cholesterol with moderate increase in triglycerides Cholesterol with marked increase in triglycerides Triglycerides Marked increase in triglycerides and cholesterol

Chylomicrons LDL LDL and VLDL IDL and chylomicrons VLDL Chylomicrons and VLDL

IDL = Intermediate-density lipoprotein. LDL = Low-density lipoprotein. VLDL = Very-low-density lipoprotein.

HDL, then undergoes an esteriÞcation process catalyzed by the plasma enzyme lecithin:cholesterol acyltransferase (LCAT). The cholesteryl esters that form on the surface of HDL are then transferred to VLDL or IDL through the activity of CETP, and eventually back to LDL, thereby completing a cycle in which LDL delivers cholesterol to extrahepatic cells and cholesterol is returned to LDL via HDL. Etiology The various conditions that fall under the umbrella term dyslipidemia are classiÞed by lipoprotein phenotype according to the Frederickson/World Health Organization (WHO) classiÞcation system for hyperlipoproteinemias. Table 1 outlines this classiÞcation system. Patients with primary defects in the synthesis or degradation of lipoprotein particles suffer from primary dyslipidemia. Dyslipidemia that is a manifestation of secondary clinical factors that alter lipoprotein status is known as secondary dyslipidemia. Primary Dyslipidemia. Primary dyslipidemia encompasses phenotypes that may result from genetic diseases whose primary effects are on lipoprotein metabolism. Hypercholesterolemia is one of the most frequently observed primary dyslipidemias, particularly familial-combined hypercholesterolemia (FCH) and polygenic hypercholesterolemia. FCH is not present in childhood; elevations in plasma cholesterol and/or TG levels appear at puberty and continue throughout life. These lipid elevations tend to be mild and to vary with time. Most of the factors that result in polygenic hypercholesterolemia are unknown. Subtle genetic differences likely affect many of the processes that govern cholesterol metabolism. For example, there may be genetic polymorphisms in the proteins that govern the rates of intestinal cholesterol absorption, bile acid synthesis, cholesterol synthesis, and LDL synthesis or catabolism. Both FCH and polygenic hypercholesterolemia exhibit altered LDL levels and manifest most commonly as Type IIa under the Frederickson classiÞcation. Table 2 describes the characteristics of the primary dyslipidemia disorders.

ETIOLOGY AND PATHOPHYSIOLOGY

45

Secondary Dyslipidemia. The most common forms of dyslipidemia seen in clinical practice are secondary to other disorders (such as chronic, uncontrolled diabetes mellitus; metabolic syndrome; hypothyroidism; and nephrosis), to alcohol consumption and cigarette smoking, or to drugs such as oral contraceptives and corticosteroids. In secondary dyslipidemia, alterations in lipid metabolism seen in primary dyslipidemia can be mimicked or exacerbated. Clinically, secondary dyslipidemia is particularly important because the disorders associated with it may be causes/indicators of considerable morbidity. In addition, disordered lipoprotein metabolism may accelerate the progress of a primary disease, as has been hypothesized in connection with renal and liver disease. Table 3 summarizes the clinical disorders, physiological changes, and pharmacological treatments that are associated with secondary dyslipidemia, some of which are discussed in the following subsections. In general, the mechanistic cause underlying these common forms of secondary dyslipidemia is insulin resistance/deÞciency. Insulin resistance can result from an abnormality in the insulin, an abnormality in the insulin receptor, or an abnormality in insulin signal transduction. Type 2 Diabetes. Two signiÞcant abnormalities in lipoprotein metabolism are seen in type 2 diabetes patients: higher concentrations of VLDL than in nondiabetic patients and lower HDL concentrations. However, although altered composition of lipoproteins appears to be a feature of patients with type 2 diabetes, their TSC remains relatively unaltered; usually, only slight elevations in lipid levels are observed. The mechanism by which VLDL is raised and HDL is lowered in this patient group is thought to be insulin deÞciency or insulin resistance. In insulin-resistant patients, insulin accomplishes relatively little activation of lipoprotein lipase; consequently, clearance of VLDL particles is impeded and HDL levels are low. Metabolic Syndrome. The metabolic syndrome, also known as syndrome X or the insulin resistance syndrome, is receiving increasing attention as a cause of CHD and type 2 diabetes (Park YW, 2003). The mechanisms underlying the development of metabolic syndrome are poorly understood. This complex disorder is deÞned by the presence of obesity, insulin resistance, high fasting glucose concentration, presence of prothrombotic factors, high TGs, and low HDL. Alcohol Consumption. Ethanol elevates plasma TG levels primarily by inhibiting fatty acid oxidation and enhancing fatty acid synthesis. Fatty acids are esteriÞed to TGs; excess TGs lead to increased secretion of VLDL. Patients who suffer from severe alcoholic dyslipidemia appear to have a defect in the catabolism of VLDL particles; this defect can in turn result in increased plasma concentration of chylomicrons. Cigarette Smoking. Cigarette smoking has been associated with elevated levels of serum TG and depressed levels of HDL. Data from the Prospective Cardiovascular M¨unster Heart Study (PROCAM), which enrolled more than 20,000 men and 10,000 women, demonstrated that smokers had, on average, 7% lower

46

TABLE 2. Characteristics of Genetic Conditions Leading to Dyslipidemia Genetic Disorder

Phenotype Classification

Familial hypercholesterolemia (FH)

Inheritance

Pathogenesis

IIA, rarely IIB

Mutations in LDL-receptor gene.

Inefficient LDL uptake resulting in elevated plasma LDL levels.

Polygenic hypercholesterolemia

IIA

Impaired clearance of LDL.

Familial combined hypercholesterolemia (FCH) Remnant hyperlipoproteinemia (also known as type III hyperlipoproteinemia or dysbetalipoproteinemia) Familial hypertriglyceridemia Familial apoprotein CII deficiency

IIA, IIB, or IV, rarely V III

Susceptible genotype exacerbated by high dietary saturated fat and cholesterol intake. Gene mutation inherited as autosomal dominant trait.

Familial lipoprotein lipase deficiency (also known as familial chylomicronemia)

IV, rarely V I or V

I

LDL = Low-density lipoprotein. VLDL = Very-low-density lipoprotein.

Usually inherited; also observed in dysproteinemias and hypothyroidism.

Gene mutation of apo E inherited as autosomal dominant trait. Inherited deficiency of lipase-activating protein apo C-II. Inherited deficiency of lipoprotein lipase activity.

Excessive hepatic production of apoprotein B can result in excess LDL, VLDL, or both. Accumulation of chylomicron and VLDL remnants, leading to high plasma cholesterol and triglycerides.

Variable elevations of VLDL; higher observed incidence of atherosclerosis. Ineffective removal of chylomicrons and VLDL triglycerides due to inactive lipoprotein lipase (see below). Ineffective removal of chylomicrons; manifests as recurrent attacks of pancreatitis.

Estimated Prevalence 1 in 500 (heterozygous form); 1 in 1 million (homozygous) 1 in 25

1 in 200

1 in 10,000

1 in 300 <1 in 1 million

1 in 1 million

ETIOLOGY AND PATHOPHYSIOLOGY

47

HDL and 15% higher TG levels than nonsmokers (Cullen P, 1998). Smoking is also strongly associated with elevations in both C-reactive protein (CRP) and Þbrinogen, which are markers of systemic inßammation and increased thrombotic activity, respectively (Fr¨ohlich M, 2003). These two factors have been linked to the pathogenesis of atherogenesis and CVD. (See the “Cardiovascular Disease Markers in Dyslipidemia and Atherosclerosis” section for more on CRP.) It is unclear whether these lipid abnormalities are a result of physiological or dietary changes associated with smoking. Nevertheless, experts believe that smoking is the strongest independent predictive factor for CVD and recommend immediate cessation for dyslipidemic patients (Third Report, 2001). Oral Contraceptives. Estrogens tend to raise serum TG levels because they increase hepatic VLDL production. In most women, the catabolism of VLDL also increases when taking oral contraceptives, so the overall increase in plasma TG level is modest. However, in women who have an underlying genetic disorder (such as familial hypertriglyceridemia or FCH), the plasma VLDL-TG level can increase markedly, and hyperchylomicronemia can develop when estrogencontaining medications are taken. Pathophysiology Dyslipidemia: A Risk Factor for CVD. High serum cholesterol, particularly cholesterol associated with LDL and VLDL, is one of the principal risk factors for atherogenesis. Characterized by an accumulation of lipid within artery walls, atherogenesis is a key factor in the development of CVD. When lipoproteins cross the endothelial cell barrier and enter the vessel wall, they become progressively oxidized. In this state, they are taken up by macrophages, which subsequently transform into foam cells. Oxidized LDL (ox-LDL) may also activate endothelial cells. Both foam cells and endothelial cells secrete growth factors that stimulate the migration and proliferation of vascular smooth-muscle cells and monocytes. This inßammatory process results in the formation of atherosclerotic lesions. Secreted growth factors also increase the binding of LDL to endothelial cells, where apolipoprotein components promote thrombosis by interfering with the conversion of plasminogen to plasmin. Thus, a cycle of inßammation may be started, ultimately leading to arterial infarction. Cardiovascular Disease Markers in Dyslipidemia and Atherosclerosis. Despite substantial evidence that links LDL to CVD risk, a proportion of the CVD population shows no discernible lipoprotein abnormalities. Consequently, other factors have been identiÞed as potential markers of disease. These factors can be classiÞed as lipoprotein particles, such as lipoprotein (a); inßammatory markers, such as CRP; and nonlipid factors such as homocysteine. Questions remain about whether these factors are consequences or causes of CHD. Lipoprotein Markers. The use of lipoprotein(a) (Lp[a]) particles and apolipoproteins as prognostic indicators for CVD has been explored and is receiving continuing attention. In particular, a substantial body of evidence points to the clinical

48

DYSLIPIDEMIA

TABLE 3. Diseases, Physiological Changes, and Pharmacological Treatments Associated with Secondary Dyslipidemia Disease, Physiological Change, or Pharmacological Treatment

Lipoprotein Pattern

Endocrine and metabolic Type 2 diabetes IV (rarely V)

Glycogen storage disease

IV (rarely V)

Lipodystrophies

IV

Cushing’s syndrome

IIA or IIB

Hypothyroidism Anorexia nervosa

IIA (rarely III) IIA

Metabolic syndrome

Elevated TGs, low HDL

Drug-induced Alcohol consumption

IV (rarely V)

Oral contraceptives

IV (rarely V)

Cigarette smoking

Glucocorticoids

Low HDL, increase in atherogenic particles IIA or IIB

Renal Uremia

IV

Nephrotic syndrome

IIA or IIB

Hepatic Primary biliary cirrhosis and extrahepatic biliary obstruction

Elevated cholesterol, elevated phospholipids, elevated lipoprotein X

Proposed Mechanism for Dyslipidemia Increased secretion of VLDL and lower HDL due to insulin resistance; decreased catabolism of VLDL and chylomicrons due to reduced lipoprotein lipase activity. Increased secretion of VLDL and lower HDL due to insulin resistance; decreased catabolism of VLDL and chylomicrons due to reduced lipoprotein lipase activity. Increased secretion of VLDL due to insulin resistance. Increased secretion of VLDL with conversion to LDL due to insulin resistance. Decreased catabolism of VLDL and IDL. Reduced biliary excretion of cholesterol and bile acids. Multiple mechanisms.

Increased secretion of VLDL in persons genetically predisposed to hypertriglyceridemia. Increased secretion of VLDL in persons genetically predisposed to hypertriglyceridemia. Multiple mechanisms, possibly including interaction with apolipoprotein E allele 4 (apo epsilon[4]) gene. Increased secretion of VLDL with conversion to LDL. Decreased catabolism of VLDL due to reduced lipoprotein lipase activity. Increased secretion of VLDL; direct secretion of LDL from liver; decreased catabolism of VLDL and LDL. Diversion of biliary cholesterol and phospholipids into bloodstream.

ETIOLOGY AND PATHOPHYSIOLOGY

49

TABLE 3. (continued) Disease, Physiological Change, or Pharmacological Treatment

Lipoprotein Pattern

Acute hepatitis

IV

Hepatoma

IIA

Immunological Systemic lupus erythematosus Monoclonal gammopathies (myeloma, macroglobulinemia, lymphoma) Stress-induced Emotional stress, acute myocardial infarction, extensive burns, acute gram-negative sepsis

I III or IV

IV

Proposed Mechanism for Dyslipidemia Decreased hepatic secretion of lecithin cholesterol acyltransferase (LCAT). Lack of feedback inhibition of hepatic cholesterol synthesis by dietary cholesterol. Presence of IgG or IgM that binds heparin, thereby decreasing activity of lipoprotein. Presence of IgG or IgM that forms immune complex with chylomicron remnants and/or VLDL, thereby decreasing their catabolism.

Increased secretion and decreased catabolism of VLDL.

HDL = High-density lipoprotein. TGs = Triglycerides. Ig = Immunoglobin. LDL = Low-density lipoprotein. VLDL = Very-low-density lipoprotein.

utility of apolipoprotein B (ApoB). As the principle protein component of LDL, IDL, VLDL, and chylomicrons, ApoB is the major apolipoprotein of all atherogenic lipoproteins. Each lipoprotein particle contains a single molecule of ApoB. Therefore, ApoB level is a quantitative indicator of the number of atherogenic lipoprotein particles in plasma and a good indicator of atherosclerotic risk. In addition, elevated levels of ApoB have been found to exhibit a stronger association with other risk factors for CVD than LDL (Williams K, 2003). Several companies have marketed assays to measure ApoB levels, including Abbott, Exocell, and Japan Immunoresearch Labs. Inflammatory Markers. Because inßammatory response has a pivotal role in the development of atherosclerosis, inßammation markers are potentially useful clinical indicators of CVD. Several inßammation markers have been identiÞed, including CRP, interleukin-6 (IL-6), plasminogen activator, intracellular adhesion molecules, tumor necrosis factor alpha (TNF-α), soluble CD40 ligand, and lipoprotein phospholipase A2. CRP has been associated with acute myocardial events for several decades. Of particular interest are asymptomatic patients who have both elevated CRP

50

DYSLIPIDEMIA

levels and high cardiovascular risk despite falling below the recommended LDL threshold values for treatment (Ridker PM, 2002[a]). Observations that obesity and the metabolic syndrome are commonly accompanied by increases in CRP also suggest a close link between metabolic derangement and inßammation. Studies have provided evidence that statins, the standard pharmacological treatment for dyslipidemia, also lower CRP levels (Benjamin J, 2003; Nissen SE, 2005). Some researchers theorize that CRP not only signals inßammation but also actively mediates inßammatory processes involving ox-LDL, which is also an index of inßammation (Chang MK, 2002). Ox-LDL can also trigger liver production of CRP via the messenger cytokine IL-6 (Jialal I, 2001). Native LDL is thought to undergo oxidation when entering the artery wall from the circulation, although the exact nature of this process is not well understood. Prospective clinical studies will be important to elucidate the relationship between lipoprotein levels and CRP. Currently, ATP III does not recommend routine measurement of inßammatory markers for the purpose of modifying LDL goals in primary prevention. Yet, despite numerous studies, the predictive clinical value of CRP as an independent risk marker, compared with traditional risk factors, such as lipoprotein levels, remains uncertain. Findings from one large study indicate that CRP is only a relatively moderate predictor of CHD, particularly when adjustments are made to account for smoking status, other established coronary risk factors, and socioeconomic status (Danesh J, 2004). Research also points to a weak correlation between CRP and established lipid markers such as TSC, HDL, and LDL (Ridker PM, 2002[a]). In 2003, a statement from the Centers for Disease Control and Prevention (CDC) and the American Heart Association (AHA) recommended that CRP measurement should be used as an adjunct to global risk assessment and lipid screening in detecting patients without CVD who might be at higher risk than predicted by established risk factors alone (Pearson TA, 2003). The proposal is in part made possible by the growing number of commercially available highsensitivity CRP (hs-CRP) assays that can be used in an outpatient setting; more than 20 such assays are available in the United States alone (manufactured by Dade Behring [Illinois] and others). In addition, many insurance plans in the United States reimburse the cost of the hs-CRP assay. The CDC is in the process of standardizing testing (Roberts WL, 2004). Note that levels of CRP will increase in response to various inßammatory stimuli as well as to vascular injury or infection. This fact implies that CRP may have limited speciÞcity as a tool for cardiovascular risk assessment in patients with concomitant inßammatory states, including arthritis and chronic pulmonary disease. Homocysteine. Elevations of serum homocysteine are positively correlated with risk for CVD (Bostom AG, 1999; Giles WH, 2000; Whincup PH, 1999). However, the mechanism of the link between homocysteine and CVD is not well understood, and an elevation of homocysteine is less common than elevation of the major risk factors. Several clinical trials are under way to test whether

CURRENT THERAPIES

51

homocysteine-lowering therapy also reduces CVD risk. ATP III does not recommend routine measurement of homocysteine as part of risk assessment to modify LDL goals for primary prevention. CURRENT THERAPIES Dyslipidemia includes any elevation of serum low-density lipoprotein (LDL) or triglyceride (TG) levels, reduction in high-density lipoprotein (HDL) levels, or combination of these factors. Treatments for dyslipidemia may target any one of these lipid disorders. The primary goal of therapy, however, has been and remains the lowering of LDL because elevations in LDL are strongly associated with progression of coronary heart disease (CHD). Because LDL is currently the primary drug target in dyslipidemia treatment, the marketplace for this indication is dominated by the statins (HMG-CoA reductase inhibitors). Dyslipidemia treatments differ in their mechanisms of action and in their ability to modulate the different lipid disorders. Pharmacological agents also vary considerably in their side-effect proÞles, drug interactions, and dosing. In some countries, particularly Italy, over-the-counter (OTC) supplements such as omega-3 fatty acids have been a popular patient choice for lowering TG levels. These agents are now available by prescription for the treatment of hypertriglyceridemia and as adjuvant therapy after myocardial infarction in Germany, Italy, the United Kingdom, and most recently, the United States. The major pharmacological agents currently used in the treatment of dyslipidemia are summarized in Table 4. Statins (HMG-CoA Reductase Inhibitors) Overview. Statins are key components of lipid-lowering treatment regimens; members of this class are some of the most valuable agents in the entire pharmaceutical marketplace. Statins lower serum LDL cholesterol concentrations by 20–65%, depending on the particular statin and the dosage used. Major studies that have assessed the beneÞts of statins in primary prevention of CHD include the West of Scotland Coronary Prevention Study (WOSCOPS), the Air Force Texas Coronary Atherosclerosis Prevention Study (AFCAPS/TexCAPs), and the Heart Protection Study (HPS) (McDougall C, 2003). Some physicians prescribe statins for primary prevention only in men aged 45–64 who are at very high risk of acute myocardial infarction (AMI) or stroke. Others believe that the WOSCOPS and AF/TexCAPS results should encourage much more widespread use of statins in primary prevention, especially in elderly patients and patients with only moderately elevated lipids. Much of the recent debate about statins centers on the optimal LDL threshold for high-risk patients. Several ongoing trials seek to determine whether lowering LDL levels below 100 mg/dL further reduces cardiovascular risk in patients with CHD or CHD-risk equivalents (the “lower is better” hypothesis). In addition to these trials, the 2004 updated National Cholesterol Education Program (NCEP) Expert Panel on Detection, Evaluation, and Treatment of High Blood Cholesterol in Adults (Adult Treatment Panel III [ATP III]) guidelines are likely to inßuence prescribing behavior by promoting statin use. Additionally, as generic versions

52

DYSLIPIDEMIA

TABLE 4. Current Therapies Used for Dyslipidemia Agent

Company/Brand

Daily Dose

Availability

10–80 mg qd

US, F, G, I, S, UK, J US, F, G, I, S, UK, J

Statins (HMG-CoA reductase inhibitors) Atorvastatin Simvastatin

Rosuvastatin Pravastatin

Fluvastatin

Lovastatin

Pitavastatin

Pfizer’s Lipitor/Tahor/Sortis/ Torvast/Cardyl Merck’s Zocor/Sinvacor, Boehringer Ingelheim’s Denan, generics AstraZeneca’s Crestor Bristol-Myers Squibb’s Pravachol/Pravasin/Selectin/ Lipemol/Lipostat/Elisor, Sankyo’s Mevalotin/Sanaprav, generics Novartis’ Lescol/Locol, Solvay’s Digaril Merck’s Mevacor/Mevinacor, Andrx’s Altoprev (extended-release), generics Sankyo/Nikken/Kowa Kogyo’s Livalo

10–80 mg qd

5–40 mg qd 10–40 mg qd

US, F, I, UK, J US, F, G, I, S, UK, J

20–80 mg qd, up to 40 mg tid 10–80 mg qd

US, F, G, I, S, UK, J US, G, S

1–4 mg qd

J

Statin combination therapies Simvastatin/ ezetimibe Lovastatin/ nicotinic acid (ER)

Merck and Schering-Plough’s Vytorin/Inegy Kos’s Advicor

Atorvastatin/ amlodipine

Pfizer’s Caduet

US, G, UK Lowest dose is 500 mg niacin/20 mg lovastatin qd; maximum dose is 2,000 mg/40 mg qd. Maximum dose is 10 mg amlodipine/80 mg atorvastatin qd.

US

67 mg tid, 200 or 267 mg qd

US, F, G, I, S, UK, J

200 mg up to tid

F, G, I, S, UK, J

450–750 mg bid

US, F, G, I, S, UK

US

Fibrates (fibric acid derivatives) Fenofibrate

Bezafibrate Gemfibrozil

Fournier’s Lipantil/Lipidil/Fulcro/Secalip, Abbott’s Tricor, generics Roche’s Bezalip/Cedur/Befizal, generics Pfizer’s Lopid/Lipur, generics

CURRENT THERAPIES

53

TABLE 4. (continued) Agent

Company/Brand

Daily Dose

Availability

Cholesterol absorption inhibitors Ezetimibe

Merck and ScheringPlough’s Zetia/Ezeterol

10 mg qd

US, F, G, S, UK

Kos/Merck KGaA’s Niaspan, Upsher-Smith’s Slo-Niacin

0.5–2.0 g qd

US, F, G, UK, J

Genzyme/Sankyo’s Welchol Bristol-Myers Squibb’s Questran, generics

1.88 g bid

US

Up to 36 g qd, in up to four divided doses

US, F, G, I, S, UK

Nicotinic acid derivatives Extendedrelease nicotinic acid Bile acid sequestrants Colesevelam Cholestyramine

bid = Twice daily; ER = Extended release; qd = Per day; tid = Three times daily. US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan.

of agents such as pravastatin (Bristol-Myers Squibb’s Pravachol) and especially the universally popular simvastatin (Merck’s Zocor) become available in more countries, the reduced cost of therapy may encourage more widespread uptake of statins. Although statins are generally well tolerated and cause few side effects, their safety proÞle has drawn increased scrutiny in recent years following the withdrawal of cerivastatin (Bayer’s Lipobay) in August 2001 and more recent concerns about rosuvastatin (AstraZeneca’s Crestor; see the “Rosuvastatin” section). Bayer withdrew cerivastatin after higher doses were linked to cases of lifethreatening rhabdomyolysis (the destruction of skeletal muscle) that led to 31 deaths in the United States; 12 deaths occurred in patients who were receiving concomitant therapy with the Þbrate gemÞbrozil (PÞzer’s Lopid, generics). Bayer’s action resulted in a review of statins’ safety and publication of new guidelines for their use by the key regulatory bodies in the seven major pharmaceutical markets (United States, France, Germany, Italy, Spain, United Kingdom, and Japan) (Pasternak RC, 2002). Because of rare but potentially serious hepatotoxicity, liver function monitoring is now recommended before and during treatment with statins. The principal reason for potentially harmful interactions of statins with other drugs is competitive elimination via the P450 isoenzyme 3A4. Drugs that use this metabolic pathway include cyclosporin, erythromycin, azole antifungals, and various drugs for hypertension and diabetes. Statins that are not metabolized via P450 3A4 are ßuvastatin (Novartis’ Lescol), which primarily uses the P450 2C9 pathway, and pravastatin, which is at least partially metabolized via other

54

DYSLIPIDEMIA

routes. Therefore, ßuvastatin and pravastatin are often preferred in treating renally compromised patients, especially when immunosuppression is employed. Mechanism of Action. Statins are structurally similar to the cholesterol precursor HMG-CoA; they act as competitive inhibitors of HMG-CoA reductase, the rate-limiting enzyme in cholesterol biosynthesis. Statins act to inhibit the conversion of hydroxymethyglutaryl to mevalonic acid. In response to reduced hepatic cholesterol biosynthesis, activation of the transcription factor sterol regulatory element binding protein (SREBP) upregulates LDL receptor gene expression, which leads to enhanced clearing of serum lipids by the liver (Liao JK, 2003). Statins are commonly believed to confer additional beneÞcial effects on cardiovascular health, independently of their cholesterol-lowering activities. Termed pleiotropic effects, these actions stem from statins’ ability to modify endothelial function, possibly by promoting the production of nitric oxide and inhibiting the production of inßammatory molecules in the endothelium (Wassmann S, 2004). This class effect of statins may reduce inßammation, stabilize atherosclerotic plaques, inhibit platelet aggregation, and improve blood ßow, all of which act to prevent coronary events. Atorvastatin. Atorvastatin (PÞzer’s Lipitor) (Figure 5) was introduced to the United States and to European markets in 1997. By 2001, the compound had overtaken Merck’s simvastatin in sales, and in 2004, it was the global salesleading pharmaceutical product. In addition to its use in primary dyslipidemia, atorvastatin is indicated for patients with heterozygous and homozygous familial hypercholesterolemia (FH) and combined dyslipidemia. Like other agents in the statin class, atorvastatin acts to competitively inhibit HMG-CoA reductase, the rate-limiting enzyme in cholesterol biosynthesis. A therapeutic response is seen within two weeks of treatment; the maximum response is usually achieved at four weeks of therapy, and this response is maintained throughout chronic administration. Pleiotropic actions of atorvastatin have been observed in several recent studies, including improvements in endothelial function in both diabetic and nondiabetic patients (Dalla NE, 2003; John S, 2005; Sakabe K, 2004; Wassmann S, 2004).

FIGURE 5. Structure of atorvastatin.

CURRENT THERAPIES

55

Many clinical studies have assessed the effects of lipid-lowering with atorvastatin to prevent cardiovascular disease in dyslipidemia patients. Published data from the Anglo-Scandinavian Cardiac Outcomes Trial (ASCOT) support the use of atorvastatin, particularly in patients with an elevated cardiovascular risk proÞle (Sever PS, 2003). The lipid-lowering arm of this study compared the effect of 10 mg atorvastatin with placebo on CHD end points in 9,000 hypertensive patients with total cholesterol greater than 250 mg/dL. Atorvastatin-treated patients showed a 36% reduction in nonfatal MI and fatal CHD, with a 29% reduction in all coronary events compared with the placebo arm (p = 0.0005). These highly signiÞcant reductions prompted early termination of the lipidlowering arm of this study in October 2002; open-label atorvastatin was then offered to all patients who continued in the antihypertensive arms of the study. No signiÞcant differences in adverse events or liver enzyme abnormalities were observed when treatment and placebo groups were compared over the median follow-up period of 3.3 years. Several large-scale clinical trials have assessed the actions of atorvastatin and compared the agent with other commonly prescribed agents in the statin drug class. The Comparative Dose EfÞcacy Study of Atorvastatin Versus Simvastatin, Pravastatin, Lovastatin, and Fluvastatin in Patients with Hypercholesterolemia Study (CURVES) was the Þrst study to compare the safety and efÞcacy of Þve statins (Jones P, 1998). Atorvastatin produced greater reductions in LDL cholesterol than milligram-equivalent doses of pravastatin, simvastatin, ßuvastatin, or lovastatin (Merck’s Mevacor, generics). Also, compared with higher doses of the other statins, the usual starting dose of atorvastatin 10 mg/day produced similar or greater LDL cholesterol reductions. The Aggressive Lipid-Lowering Initiation Ablates New Cardiac Events (ALLIANCE) study is one example of several recently published trials that have tried to establish the value of aggressively lowering LDL below recommended target levels in patients with CHD. This study included 2,442 patients with a history of CHD and compared cardiovascular outcomes of patients who received atorvastatin (up to 80 mg) and were titrated to LDL levels of <80 mg/dL with the outcomes of patients who received usual care: lipid-lowering treatment that includes lifestyle modiÞcations such as diet and exercise and drug treatment if necessary (Koren MJ, 2004). Patients began the study on 10 mg of atorvastatin; the dose was titrated upward until an LDL cholesterol level of <80 mg/dL was reached (the maximum dose of atorvastatin was 80 mg). The investigators reported that patients taking atorvastatin experienced fewer nonfatal heart attacks than patients receiving usual care: 4.3% versus 7.7%. The atorvastatin patient arm also experienced a 17% reduction in overall negative cardiovascular outcomes, including cardiac death, myocardial infarction (MI), and stroke. Fewer patients in the atorvastatin arm were hospitalized. Data from the Treating to New Targets (TNT) study, presented at the American College of Cardiology (ACC) conference in March 2005, lend further credence to the “lower is better” hypothesis of LDL lowering. In this trial, 10,001 patients with established CHD were treated with open-label 10 mg atorvastatin daily

56

DYSLIPIDEMIA

to a mean LDL level of 98 mg/dL. Patients were then randomized to receive either 10 mg or 80 mg atorvastatin daily and followed for a median 4.9 years. The primary end point, occurrence of a major cardiovascular event—deÞned as death from CHD; nonfatal, nonprocedure-related MI; resuscitation from cardiac arrest; or fatal or nonfatal stroke—was lower in patients treated with 80 mg atorvastatin daily than in patients treated with 10 mg daily: 8.7% versus 10.9%, a 22% relative risk reduction (p < 0.001) (LaRosa JC, 2005). However, TNT did not demonstrate a difference in overall mortality between the two treatment groups; an increase in noncardiovascular deaths was seen in the 80 mg atorvastatin group. The Reversal of Atherosclerosis with Aggressive Lipid Lowering (REVERSAL) study is particularly interesting in terms of its implications for treatment of dyslipidemia (Nissen SE, 2004). This randomized, double-blind study, involving 654 patients in 34 centers across the United States, investigated the effects of aggressive lipid-lowering with atorvastatin (80 mg) or pravastatin (40 mg) on the progression of atherosclerosis over an 18-month period. Baseline LDL cholesterol (mean 150.2 mg/dL) was reduced to 110 mg/dL in the pravastatin-treated group and to 79 mg/dL in the atorvastatin-treated group. C-reactive protein (CRP), a marker of cardiovascular risk, was reduced by 5.2% in the pravastatin group and by 36.4% in the atorvastatin-treated group. The primary end point of this study was the progression of coronary atherosclerosis. Progression did occur in the pravastatin-treated group (2.7%; 95% conÞdence interval [CI] 0.2 to 4.7%; p < 0.01); in the atorvastatin-treated group, no progression was observed (0.4%; CI −2.4% to 1.5%; p = 0.98). PÞzer will seek to provide more evidence for the “lower is better” hypothesis from the ongoing Incremental Decrease in Endpoints through Aggressive Lipid lowering (IDEAL) study, initiated in 1999. The 8,888 CHD patients enrolled in IDEAL include a large number of elderly patients; all patients will be randomized to 80 mg atorvastatin or 20/40 mg simvastatin daily and followed for Þve and one-half years. The primary end point in the trial is the time to initial occurrence of a major coronary event (Pedersen TR, 2004). Simvastatin. Simvastatin (Merck’s Zocor, generics) (Figure 6) was launched in Europe in 1988 and in the United States and Japan in 1992. In August 1999, the FDA approved the use of simvastatin for raising low HDL cholesterol levels, the Þrst such approval for a statin. Of the major markets under study, only the United States currently affords patent protection to simvastatin. Prior to the launch of atorvastatin (PÞzer’s Lipitor), simvastatin was the most potent of all the statins on the market. Simvastatin competitively inhibits HMG-CoA reductase, the rate-limiting enzyme in cholesterol biosynthesis. Pleiotropic effects, such as improvements in endothelial function, have been demonstrated in some studies of simvastatin (Pereira EC, 2003; Rezaie-Majid A, 2003). Strong clinical data support the beneÞts of simvastatin in the treatment of dyslipidemia. Owing to results of the landmark Scandinavian Simvastatin Survival Study (4S) in 1994, simvastatin was the Þrst lipid-lowering drug to receive

CURRENT THERAPIES

57

FIGURE 6. Structure of simvastatin.

approval from the FDA for use in preventing AMI in patients with coronary artery disease and dyslipidemia. This randomized study with a follow-up of 5.4 years was designed to investigate the actions of simvastatin treatment versus placebo on mortality and morbidity in patients with CHD. The study enrolled 4,444 patients who had angina pectoris or had suffered a previous MI and had total serum cholesterol (TSC) of 5.5–8.0 mmol/L (about 210–310 mg/dL). These patients were placed on a lipid-lowering diet and received either 20 mg once daily of simvastatin or placebo; if serum cholesterol did not reach the target of 3.0–5.2 mmol/L (about 115–200 mg/dL) with simvastatin, doses were adjusted to either 10 or 40 mg daily. Over the course of the study, treatment with simvastatin reduced TSC (−25%) and LDL cholesterol (−35%), and increased HDL (+8%) with few adverse side effects. Simvastatin treatment also signiÞcantly reduced mortality: 256 deaths (12%) occurred in the placebo group; 182 deaths (8%) occurred in the simvastatin group. Other beneÞts included a 37% reduction in the risk of undergoing myocardial revascularization procedures (Kjekshus J, 1994). A recent subgroup analysis of 3,933 nondiabetic patients in the 4S study revealed that simvastatin demonstrated equivalent relative risk reduction in patients with or without metabolic syndrome over the Þve year follow-up period (Pyorala K, 2004). Data from the Heart Protection Study (HPS), published in 2002, provide further support for simvastatin as an important treatment for lipid abnormalities. In this study, 20,536 high-risk patients were randomized to receive simvastatin (40 mg daily) or placebo, with a mean follow-up of 5.5 years. Simvastatin reduced the coronary mortality rate by 18% (p = 0.0005): total mortality was reduced from 14.7% in the placebo arm to 12.9% in the simvastatin arm (p = 0.0003). SigniÞcant reductions were also seen in stroke, major cardiovascular events, and the need for coronary and noncoronary revascularization (all p < 0.0001). BeneÞt was clearly seen in women, the elderly, diabetics, and patients with prior noncardiac atheromatous disease; these results will likely extend the use of simvastatin within these populations (Heart Protection Study, 2002). Moreover, the HPS demonstrated that even in patients with LDL values below 100 mg/dL, there was a similar, incremental reduction in major vascular events.

58

DYSLIPIDEMIA

The Japan Lipid Intervention Trial (J-LIT) was the Þrst large-scale study to examine the relationship between coronary risk factors and incidence of CHD in Japanese patients treated with simvastatin therapy. The J-LIT study enrolled 47,294 patients without CHD and 5,127 patients with CHD, all with TSC levels >220 mg/dl; all patients received open-label simvastatin at doses of 5–10 mg/daily for six years under standard clinical conditions. In the primary prevention cohort, simvastatin treatment reduced TSC, LDL, and TG by 18.4%, 26.8%, and 16.1%, respectively, and increased HDL by 4.5%. In the secondary prevention cohort, simvastatin treatment reduced TSC, LDL, and TG by 19.8%, 28.6%, and 15.9%, respectively, and increased HDL by 4.7% (Mabuchi H, 2002; Matsuzaki M, 2002; Matsuzawa Y, 2003). During the study period, major coronary events were observed in 336 patients in the primary prevention cohort (n = 47, 294) and in 102 patients in the secondary prevention cohort (n = 5, 127); these data were used to develop Japanese risk assessment charts for guidelines published by the Japanese Society of Atherosclerosis (JAS). More recently, simvastatin studies have been conducted in patients who demonstrate combined LDL and non-LDL lipid abnormalities. The Comparative HDL EfÞcacy and Safety Study (CHESS) study produced evidence that simvastatin is superior to atorvastatin in raising HDL levels. The head-to-head trial compared maximum doses of simvastatin (80 mg daily) and atorvastatin (80 mg daily) in 917 hypercholesterolemic patients, including some who fulÞlled NCEP ATP III metabolic syndrome criteria. Simvastatin increased HDL cholesterol values signiÞcantly more than atorvastatin for the mean of 6 and 12 weeks (8.9% versus 3.6% and 4.9% versus −0.9%, respectively) and the mean of 18 and 24 weeks (8.3% versus 4.2% and 3.7% versus −1.4%, respectively). However, reductions in LDL and TG levels were greater in the atorvastatin-treated group (Ballantyne CM, 2003). The beneÞt of aggressive LDL lowering with high-dose simvastatin therapy is being evaluated in the ongoing Study of the Effectiveness of Additional Reduction in Cholesterol and Homocysteine with Simvastatin and Folic Acid/Vitamin B12 (SEARCH). In this trial, 12,000 patients are randomized to receive 80 mg or 20 mg simvastatin, with or without folate and vitamin B12, with a primary end point of death from CHD or nonfatal MI. SEARCH will also assess the effects of lowering homocysteine with folic acid (MacMahon M, 2000). Data from the trial are expected in 2006. Rosuvastatin. Rosuvastatin (Crestor) (Figure 7) was developed by AstraZeneca under license from Shionogi Pharmaceuticals. In February 2003, AstraZeneca launched rosuvastatin (10–40 mg) in Canada (at a 15% price reduction compared with atorvastatin [PÞzer’s Lipitor]), followed in March by launches in the Netherlands (the reference state for Europe) and in the United Kingdom (under the mutual recognition [MR] procedure), where the drug is priced the same as atorvastatin. The drug is currently launched in 25 countries, most recently in Japan in 2005. Despite the mutual recognition procedure in Europe, launches in Germany and Spain are currently on hold while these nations await further

CURRENT THERAPIES

59

FIGURE 7. Structure of rosuvastatin.

safety data. In any case, AstraZeneca may reconsider marketing rosuvastatin in Germany owing to prohibitive reference price restrictions in this country. Rosuvastatin competitively inhibits HMG-CoA reductase, the rate-limiting enzyme in cholesterol biosynthesis. As mentioned earlier, some concerns have arisen about the use of this drug. Rosuvastatin has an adverse-event proÞle similar to that of atorvastatin, pravastatin, and simvastatin, with no signiÞcant increases in creatinine kinase levels reported (Olsson AG, 2002). However, myopathy was reported in Phase III trials in some patients receiving the 80 mg dose; subsequently, AstraZeneca voluntarily withdrew this dose of rosuvastatin. In 2005, the FDA approved revisions to rosuvastatin’s product label, submitted by AstraZeneca following a review of postmarketing study data. The changes include more emphasis on the safe use of the 40 mg dose to reduce the risk of myopathy and explicit recommendations that certain classes of patients start therapy with the 5 mg dose: patients taking cyclosporin, Asian patients, and patients with severe renal insufÞciency, who have predisposing factors for myopathy. Concerns have also been raised about the agent’s interaction with warfarin, following the report of an adverse event in the continuing JustiÞcation for the Use of Statins in Primary Prevention: An Intervention Trial Evaluating Rosuvastatin (JUPITER) study: an elderly woman receiving long-term warfarin therapy developed increased bruising and lightheadedness after four weeks of rosuvastatin therapy. This patient’s international normalized ratio (INR) rose from 2.0 to 8.0, resulting in hospital admission (Barry M, 2004). AstraZeneca has conducted a series of studies investigating potential interactions between vitamin K antagonists (the class of agents to which warfarin belongs) and rosuvastatin, and the company has emphasized that patients receiving warfarin and rosuvastatin (or any other statin) in combination should be carefully monitored. Long-term efÞcacy and safety data are required to reassure physicians and further support the use of this superstatin. Rosuvastatin demonstrates unique binding to the HMG-CoA reductase enzyme; it is relatively hydrophylic, promoting easy uptake by and activity in hepatic cells (key cells associated with statin actions). The agent also displays minimal metabolism via cytochrome P450 enzymes and no metabolism via the 3A4 pathway, thus reducing the potential for interactions with other agents using

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this pathway (Schuster H, 2003). Pleiotropic effects, including an improvement in vascular endothelial function, have been observed in treatment with this agent. A wealth of clinical data is available on the clinical efÞcacy of rosuvastatin in treatment of a wide range of dyslipidemias. In two dose-ranging studies, 206 patients with hypercholesterolemia were treated over six weeks with 1, 2.5, 5, 10, 20, 40, or 80 mg/day of rosuvastatin. Mean reductions in LDL of 34% (1 mg dose) and 65% (80 mg dose) were recorded by the end of the study; in both instances, 90% of the reduction was observed by the second week. Increases in HDL (from 9% to 14%) and decreases in TGs (from 10% to 35%) were observed, although these effects did not appear to be dose-dependent. Adverse events were similar across treatment and placebo groups, with no elevations in alanine amino transferase or creatinine kinase levels in either group (Olsson AG, 2001[b]). Rosuvastatin’s main advantage over other statins is its potency. In a long-term study that compared rosuvastatin with other statins, 477 patients were started on rosuvastatin (5 mg or 10 mg), pravastatin (20 mg), or simvastatin (20 mg). After 12 weeks, if NCEP ATP II treatment goals were not achieved, the statin dose was titrated to a maximum of 80 mg for rosuvastatin, 40 mg for pravastatin, or 80 mg for simvastatin. At 12 weeks, statistically signiÞcant decreases in LDL levels were observed in the 5 mg and 10 mg rosuvastatin-treated patients compared with patients who had received 20 mg pravastatin or simvastatin (39%, 47%, 27%, and 35%, respectively; p < 0.05). After 52 weeks, a greater number of rosuvastatin-treated patients remained at their starting dose. After dose titration, 88% of the rosuvastatin 5 mg and 10 mg groups achieved their ATP II LDL cholesterol goals, compared with 60% of the pravastatin group and 72.5% of the simvastatin group. Additionally, a greater increase in HDL levels and a greater reduction in TG levels were observed in patients treated with rosuvastatin (Brown WV, 2002). AstraZeneca is supporting rosuvastatin with a large-scale postmarketing study program. This large-scale clinical trial program, known as GALAXY, comprises 14 ongoing clinical studies with more than 40,000 patients recruited worldwide. The program includes a range of patient populations that are at risk of coronary events and aims to test rosuvastatin’s effects on inßammatory markers, on atherosclerosis, and on reducing cardiovascular morbidity and mortality. A six-week trial under the GALAXY program umbrella, the Statin Therapies for Elevated Lipid Levels Compared Across Doses to Rosuvastatin (STELLAR) study, compared 10, 20, 40, and 80 mg rosuvastatin with equivalent doses of atorvastatin, simvastatin, or pravastatin in 2,431 dyslipidemic patients. Results showed that dose for dose, rosuvastatin lowered LDL and raised HDL to a signiÞcantly greater degree than any of the other statins (Jones PH, 2003). Recently published data from other studies in the GALAXY program support the use of rosuvastatin in high-risk patients versus other statins. In the Measuring Effective Reductions in Cholesterol Using Rosuvastatin therapy (MERCURY I) study, 86% of patients on 10 mg rosuvastatin reached the LDL goal (116 mg/dL), compared with 80% of patients on 10 mg atorvastatin, 72% on 20 mg simvastatin, and 66% on 40 mg pravastatin; these results were achieved after eight weeks of

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therapy in 3,140 patients with dyslipidemia and comorbid CHD, atherosclerosis, or type 2 diabetes (Schuster H, 2003). Data from the follow-up MERCURY II study, presented at the 75th European Atherosclerosis Society (EAS) Congress in April 2005, demonstrated that more patients achieved the LDL target after switching to 10 mg rosuvastatin daily from 10 mg atorvastatin or 20 mg simvastatin, compared with patients who stayed on the initial statin therapy throughout the treatment period. Rosuvastatin has also demonstrated beneÞt over other statins in addressing low HDL levels in patients with metabolic syndrome. The Comparative Study with Rosuvastatin in Subjects with Metabolic Syndrome (COMETS) trial compared 10 mg and 20 mg rosuvastatin with equivalent doses of atorvastatin in 397 patients. Rosuvastatin reduced LDL by 42–49%, reduced TGs by 19–24%, and raised HDL by 9.3–10.5%, compared with ranges of 36–43%, 20–24%, and 4.8–5.7%, respectively, for atorvastatin. A subgroup analysis of 811 patients in the STELLAR trial also demonstrated that rosuvastatin had the most favorable effect on atherogenic dyslipidemia associated with the presence of metabolic syndrome (Deedwania PC, 2005). Pravastatin. Sankyo Þrst launched pravastatin (Figure 8) in 1989 in Japan as Mevalotin; licensee Bristol-Myers Squibb subsequently launched the drug in Western markets during the early 1990s as Pravachol. The product has since become the leading ethical pharmaceutical in Japan; together, Japan and the United States account for more than 80% of product sales. Generic pravastatin has been launched in France, Germany, Spain, and the United Kingdom. Pravastatin competitively inhibits HMG-CoA reductase, and pleiotropic effects have been demonstrated in pravastatin treatment (Mulder HJ, 2003). Unlike other statins, pravastatin is not extensively metabolized by P450 isoenzymes and therefore does not compete with agents such as cyclosporin, thereby enhancing its value in renal transplant patients. Also, according to some experts, pravastatin is not as toxic to muscles as some other statins because it is less readily absorbed. Therefore, for patients who have myositis (inßammation of muscle tissue) but

FIGURE 8. Structure of pravastatin.

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do not have a high creatine phosphokinase (CPK) level (indicative of muscle damage), pravastatin may be the drug of choice. The remarkable wealth of trial data supporting pravastatin has ensured its continued use in clinical practice. The Þrst major primary prevention trial for a statin—the West of Scotland Coronary Prevention Study (WOSCOPS)—examined pravastatin (Shepherd J, 1995). The results of this study are the basis of this drug’s indication for primary prevention of heart disease. This randomized, double-blind, placebo-controlled trial that included 6,595 hypercholesterolemic men assessed the actions of 40 mg pravastatin daily. Treatment with pravastatin lowered TSC by 20% and LDL by 26%; no change was observed with placebo. There were 248 coronary events in the placebo group compared with 174 events in the pravastatin group, a relative reduction of 31%. Similar reductions were observed with respect to nonfatal MIs and death from CHD and all cardiovascular causes. A 22% reduction in risk of death from any cause also occurred with pravastatin treatment. Further analysis of data from the WOSCOPS study suggests that pravastatin therapy results in a 30% reduction in the risk of diabetes development (Freeman DJ, 2001). Two major secondary prevention studies have examined pravastatin. The 4,159patient Cholesterol And Recurrent Events (CARE) study demonstrated that pravastatin (40 mg daily) therapy leads to a 20% decrease in total cholesterol and a 28% decrease in LDL cholesterol (Sacks FM, 1996). An association between the inßammatory markers CRP and serum amyloid A (SAA) and coronary events was signiÞcant in the placebo group but not in patients treated with pravastatin, raising the possibility that the efÞcacy of pravastatin may partly result from antiinßammatory properties as well as lipid-lowering properties. The Long Term Intervention with Pravastatin in Ischemic Disease (LIPID) study, which included 9,014 patients with a history of CHD, demonstrated after a mean follow-up of 6.1 years that the incidence of all cardiovascular outcomes was consistently lower in patients receiving pravastatin (40 mg daily) (LIPID Study Group, 1998). When the 13,173 patients from the CARE and LIPID studies were assessed together, a 22% reduction in total stroke incidence and a 25% reduction in nonfatal stroke was recorded (Byington RP, 2001[b]). A recent substudy analysis of 2,073 patients in the LIPID study with HDL <40 mg/dL showed that the relative reduction of CHD risk or all-cause mortality risk is signiÞcantly reduced with pravastatin therapy (Colquhoun D, 2004). Pravastatin fared less well with respect to clinical outcomes in the lipidlowering arm of the Antihypertensive and Lipid-Lowering Treatment to Prevent Heart Attack trial (ALLHAT-LLT), which examined a speciÞc subgroup of patients (older, moderately hypercholesterolemic, hypertensive, with at least one cardiovascular risk factor). Participants in this study were aged 55 years or older and had LDL cholesterol levels of 120–189 mg/dL (or 100–129 mg/dL if known to have CHD) and TG levels lower than 350 mg/dL. Patients were randomized to receive pravastatin (40 mg daily) or usual care. The primary outcome was an improvement in all-cause mortality. The mean follow-up for this trial was 4.8 years. Results at year 4 show a 17% reduction in cholesterol levels with

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pravastatin treatment and an 8% reduction with usual care. LDL was reduced by 28% in the pravastatin arm and 11% in the usual care arm. However, no signiÞcant difference in all-cause mortality or in CHD event rate was observed between the two groups. The investigators emphasize that the difference between the results obtained in this trial and other statin trials may be explained by the smaller than expected differential between the two treatment groups. At four years, patients in the usual-care arm of the study had reductions of 8% and 11% in TSC and LDL cholesterol, respectively, in contrast to previous statin trials where little or no difference was seen in the placebo group (ALLHAT Investigators, 2002). Clinical studies have been assessing the lipid-lowering beneÞts and safety of higher doses of pravastatin (80 mg and 160 mg), which would facilitate up-titration of doses in patients who are not achieving lipid goals. Two placebocontrolled investigations were carried out for a total of six weeks; 291 patients received 80 mg pravastatin, and 286 patients received 160 mg pravastatin. In both studies, the dose-related increases in lipid efÞcacy were achieved without compromising the safety and tolerability of the test agent. The safety of pravastatin at 160 mg supports the manufacturer’s decision to pursue development of an 80 mg version of the product (Rosenson RS, 2003). In an attempt to maintain market share for the brand, Bristol-Myers Squibb launched its combination blister pack (two pills packed together) pravastatin/aspirin (Pravigard PAC) in August 2003 in the United States. The combination is approved for secondary prevention only and is available at the same price as branded pravastatin. Fluvastatin. In 1994, ßuvastatin (Novartis’ Lescol) (Figure 9) was the fourth statin to be launched in the United States, at approximately half the cost of simvastatin. By 1997, however, the product had gained only approximately 7% of the lipid-lowering market in the United States. This lack of popularity resulted from the perception that ßuvastatin was less effective than the other statins. By 1996, an 80 mg/day dose of ßuvastatin was recommended; at launch, the recommended dose was 20 mg/day. At the higher dose, ßuvastatin lost its price advantage.

FIGURE 9. Structure of fluvastatin.

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Fluvastatin is a competitive inhibitor of HMG-CoA reductase. Like all other statins, this agent has pleiotropic effects that have been highlighted in studies (Shovman O, 2004). In addition to the constraints already described, ßuvastatin’s success was limited by a relative scarcity of supportive trial information: no primary or secondary prevention data were available in 1994, the product’s Þrst year on the market. Since then, data have been made available. The Lipoprotein and Coronary Atherosclerosis Study (LCAS) enrolled 429 people with angiographic evidence of coronary artery disease (CAD) and LDL levels of 115–190 mg/dL to receive ßuvastatin (20 mg twice daily) or placebo. After 2.5 years, the mean LDL cholesterol in patients treated with ßuvastatin declined by approximately 23%, with signiÞcantly less atherosclerotic lesion progression in the ßuvastatin arm (Herd JA, 1997). Subsequently, the FDA approved ßuvastatin for patients with CAD and moderately elevated LDL cholesterol levels. In March 1999, ßuvastatin received approval for reducing serum TG levels and apolipoprotein B (ApoB) levels. In late 2000, Novartis launched an extended-release (ER), once-daily, 80 mg formulation of ßuvastatin in several markets as Lescol XL. The Lescol Intervention Prevention Study (LIPS) compared early postoperative treatment with Lescol (80 mg/day) with placebo treatment in 1,667 patients who had average cholesterol levels and were undergoing a Þrst percutaneous coronary intervention. The primary end point of major adverse cardiac events (MACEs) was 21.4% in ßuvastatin patients and 26.7% in placebo controls. MACE rates were signiÞcantly lower in patients with diabetes (p = 0.04) or multivessel CAD (p = 0.01) who were receiving ßuvastatin (Serruys PW, 2002). Both ßuvastatin and warfarin are common substrates for cytochrome P450 2C9. Warfarin levels may increase if the two agents are given together; therefore, patients using both drugs should be carefully monitored. Lovastatin. Lovastatin (Merck’s Mevacor, generics) (Figure 10), launched in 1987, was the Þrst statin to be approved in the United States. The drug reached the European market in 1989, but is available only in Germany and Spain. In the United States, which accounted for more than 80% of sales of this drug, lovastatin became the Þrst available generic statin in December 2001. Following patent expiry in the United States, Andrx has reformulated lovastatin into an extendedrelease (ER), once-daily product (Altocor), which received FDA approval in June 2002. The FDA has twice rejected Merck’s applications to bring branded lovastatin to OTC status, in July 2000 and January 2005. Lovastatin is a competitive inhibitor of HMG-CoA reductase. The Expanded Clinical Evaluation of Lovastatin (EXCEL) study established the efÞcacy and safety of lovastatin in combination with a lipid-lowering diet in 8,000 patients (Bradford RH, 1991). With the launch of other statins, lovastatin lost ground in the United States. However, Merck continued to support lovastatin with clinical trials, in particular the AF/TexCAPS (Downs JR, 1998). The efÞcacy of ER lovastatin has been compared with that of the immediaterelease (IR) formulation in a randomized, double-blind, cross-over study that measured change in LDL as the primary end point. Patients (n = 179) received

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FIGURE 10. Structure of lovastatin.

FIGURE 11. Structure of pitavastatin.

20 mg IR or ER lovastatin for 12 weeks, placebo for 6 weeks, and then the alternate therapy (ER or IR) for 12 weeks. Statistically signiÞcant reductions in TSC and LDL were observed in patients treated with ER lovastatin; the efÞcacy proÞles of the two formulations suggested that 20 mg ER lovastatin was approximately equivalent to 30 mg IR lovastatin (Lukacsko P, 2004). The safety proÞles of both drugs were similar. Pitavastatin. Pitavastatin (Sankyo/Nikken/Kowa Kogyo’s Livalo) (Figure 11) is the most recently approved statin. The agent was codeveloped by Nissan Chemical, Kowa Kogyo, Nikken, Sankyo, and Novartis; it is reportedly more potent and longer-acting than pravastatin or simvastatin. Sankyo shares Japanese and U.S. co-marketing rights with Kowa; Kowa and Novartis are splitting European marketing rights. Kowa has also signed a licensing agreement with Recordati to market pitavastatin in Italy. Pitavastatin was launched in Japan in September 2003 for treatment of dyslipidemia; Phase III and Phase IIb clinical investigations are still under way in Europe and the United States, respectively. Pitavastatin is a competitive inhibitor of HMG-CoA reductase, the rate-limiting enzyme in the cholesterol biosynthesis pathway. Although the enzymes P450–29C and P450–28C are involved in pitavastatin’s metabolism, the agent has a low afÞnity for them, thus reducing the potential for drug interactions (Kajinami K, 2003). Pitavastatin can be safely administered with Þbrates, including gemÞbrozil, because these drugs do not affect the agent’s metabolism. Pleiotropic effects

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of pitavastatin, including improved endothelial function and anti-inßammatory actions, have been demonstrated in vitro and in vivo (Kajinami K, 2003). In a 12-week, randomized, multi-center, double-blind study, 240 patients with primary hypercholesterolemia were randomized to receive 2 mg/day pitavastatin or 10 mg/day pravastatin (Sankyo’s Mevalotin). LDL reductions were signiÞcantly greater after pitavastatin treatment than after pravastatin treatment (37.6% decrease from baseline levels versus 18.4% decrease). TG reductions were also signiÞcantly greater in the pitavastatin group. Increases in HDL were equivalent in the two treatment arms, and the safety proÞle was similar for both agents (Saito Y, 2002). A long-term study investigated the actions of pitavastatin in 308 Japanese dyslipidemia patients. Patients were given a range of doses of pitavastatin (1–4 mg/day) and followed for 44 weeks. At the end of the follow-up period, treatment with pitavastatin had decreased mean LDL levels from 202 to 121 mg/dL. TG levels also showed signiÞcant reductions (25–31%). HDL cholesterol increased from 56 to 63 mg/dL. The safety proÞle of pitavastatin in this study emulated that of other currently marketed agents (Kajinami K, 2003). Pitavastatin should both support and extend Sankyo’s antidyslipidemic franchise in Japan, which currently centers on pravastatin. Statin Combination Therapies Overview. The use of polypharmacy regimens in cardiovascular indications is often associated with poor patient compliance, particularly in many primary prevention patients, who do not experience symptoms of their disease. The increasing use of multiple agents (in addition to other therapies patients may be taking) in an attempt to establish patients at lipid goals outlined by the NCEP ATP III guidelines may also result in compliance issues. One approach to tackling such problems is the single-pill combination therapy. In recent years, there has been a surge in the development of such pills by pharmaceutical companies. The latest to reach the market is Merck and Schering-Plough’s LDL-lowering, single-pill combination of simvastatin and the cholesterol absorption inhibitor ezetimibe (Merck/Schering-Plough’s Zetia/Ezetrol), launched in the United States in July 2004. Kos Pharmaceuticals, in partnership with DuPont, was the Þrst to enter the marketplace with Advicor, a single-pill combination of lovastatin and nicotinic acid (also known as niacin). In June 2004, PÞzer announced the U.S. launch of Caduet, its single-pill combination of the blockbuster statin atorvastatin (Lipitor) and the antihypertensive amlodipine (Norvasc). Mechanism of Action. Single-pill combination therapies incorporate the mechanisms of action of each individual agent included in the single-pill formulation. Simvastatin/Ezetimibe. Merck and Schering-Plough have jointly developed a combination of simvastatin and ezetimibe (Figure 12) in a once-daily, Þxedcombination tablet under the brand name Vytorin. Merck will aim to compensate

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FIGURE 12. Structure of ezetimibe.

for the erosion of branded simvastatin sales with the new combination product; simvastatin’s market exclusivity expires in 2006 in the United States, but simvastatin/ezetimibe will be protected from generics competition until approximately 2015. In April 2004, Merck and Schering-Plough announced the Þrst European approval for the combined pill in Germany, the reference state for the European Union (EU); the combination’s brand name in this territory is Inegy. Simvastatin/ezetimibe completed the European mutual recognition procedure (MRP) in October 2004, thereby signaling the start of further approvals in Europe. In June 2005, Inegy was launched in the United Kingdom. (Ezetimibe is discussed as a monotherapy later.) Operating through two different mechanisms—hydroxymethyglutaryl (HMG)CoA reductase inhibition and inhibition of cholesterol absorption—the combination therapy promises a synergistic, LDL-lowering effect and the prospect of improved patient compliance. Merck claims that the combination will provide LDL lowering comparable to that of three titration steps of simvastatin and will lower LDL levels to a greater extent than simvastatin or other statins alone. In 668 patients with baseline LDL between 145 mg/dL and 250 mg/dL and TGs greater than 350 mg/dL, treatment with ezetimibe (10 mg/day) in combination with simvastatin (10, 20, 40, or 80 mg/day) for one month produced additional reductions in LDL (13.7%) and TGs (7.5%), and raised HDL by an additional 2.4%, compared with pooled simvastatin doses. Co-administration of ezetimibe and simvastatin is well tolerated, with a safety proÞle similar to that of simvastatin alone (Davidson MH, 2002). The superior lipid-modifying efÞcacy of the combination therapy in patients with primary hypercholesterolemia was also demonstrated in a study presented at the 2004 ACC conference (Capece R, 2004). Another study that included 1,528 hypercholesterolemic patients with LDL of 145–250 mg/dL and TGs of ≤350 mg/dL, with a follow-up of 12 weeks, investigated the action of the ezetimibe/simvastatin combination on levels of remnant lipoproteins, an independent predictor of future coronary events. The ezetimibe/simvastatin single tablet, given at a range of doses (10/10 mg to 10/80 mg), was shown to be signiÞcantly more effective than the corresponding dose of

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simvastatin alone in reducing potentially atherogenic, remnant-like lipoprotein particles (Bays HE, 2004). Data from another study compare the efÞcacy of the simvastatin/ezetimibe combination with that of atorvastatin in patients with hypercholesterolemia. Following a four-week diet/placebo regimen, patients were randomized to receive the combination or atorvastatin for four six-week periods. Baseline LDL and HDL cholesterol levels were similar in all treatment groups. Results demonstrate that at the end of the Þrst six-week period, reductions in LDL and the mean increase in HDL were signiÞcantly greater (p < 0.01) in the 10/10 mg ezetimibe/simvastatin group (n = 252) and the 10/20 mg ezetimibe/simvastatin group (n = 253) than in the 10 mg atorvastatin group (n = 250). In period 4 of the study, maximum doses of ezetimibe/simvastatin (10/80 mg) and atorvastatin (80 mg) were compared; again, the combination proved superior to atorvastatin with respect to reduction in LDL (−59.4% versus −52.5% change from baseline) and increase in HDL (+12.3% versus +6.5% change from baseline) (Ballantyne CM, 2004). A recent six-week study compared the LDL-lowering effect of ezetimibe/simvastatin versus atorvastatin across a dose range in 1,902 patients with high LDL levels. At the most commonly used starting doses of these agents, LDL was reduced by 51% in the 10/20 mg ezetimibe/simvastatin group and by 36% in the 10 mg atorvastatin group. In addition, 82% of high-risk patients (CHD or CHD-risk equivalent) on 10/20 mg ezetimibe/simvastatin achieved an LDL level below 100 mg/dL, versus 47% of patients on 10 mg atorvastatin (Ballantyne CM, 2005). Lovastatin/Nicotinic Acid (ER). Kos Pharmaceuticals has developed Advicor (formerly known as Nicostatin), a single-tablet formulation of ER nicotinic acid (also called niacin; Kos/Merck KGaA’s Niaspan) (Figure 13) and the statin lovastatin (Figure 10). This product is available only in the United States. Kos and Merck KGaA are awaiting marketing clearance in Europe. Lovastatin is a competitive inhibitor of HMG-CoA reductase, the rate-limiting enzyme in cholesterol biosynthesis. The mechanism of action of nicotinic acid has not been fully established. In a Phase III study, 814 patients with dyslipidemia were treated with escalating doses of up to 2,000 mg/day ER nicotinic acid and 40 mg/day lovastatin for four months; subsequently, they were treated at the top dose of nicotinic acid and 40 mg lovastatin for up to one year. At four months, LDL and TG levels

FIGURE 13. Structure of niacin.

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were lowered by 47% and 41%, respectively, and HDL cholesterol was increased by 30%. The decreases in LDL and TG were consistent throughout the 52-week study period; the increase in HDL rose to 41% by the study’s conclusion. Lp(a) and CRP also decreased. Treatment was well tolerated; the most common adverse event was ßushing (vasodilation of blood vessels that can cause a burning, tingling, and itching sensation), prompting the withdrawal of 10% of the patients (Kashyap ML, 2002). The effectiveness of the lovastatin/nicotinic acid combination was compared with that of atorvastatin and simvastatin in the Advicor Versus Other CholesterolModulation Agents Trial Evaluation (ADVOCATE) (Bays HE, 2002). In this study, 315 dyslipidemic patients were treated over 16 weeks with escalating doses of ER nicotinic acid combined with lovastatin (up to a maximum dose of 1,000 mg/40 mg), simvastatin (10 mg −40 mg), or atorvastatin (10–40 mg). By week 8, the lovastatin/nicotinic acid combination reduced LDL to the same degree as 10 mg atorvastatin (38%) and to a greater degree than 10 mg simvastatin (28%). Furthermore, HDL was raised by a signiÞcantly greater degree (20% versus 3% and 7%, respectively), and TG was lowered by a signiÞcantly greater amount (28% versus 15% and 10%, respectively). A retrospective analysis that assessed the use of lovastatin/ER nicotinic acid for treatment of patients with metabolic syndrome examined data from 757 dyslipidemic patients who participated in a 52-week study of the combination therapy. Patients were initially dosed with a 500 mg nicotinic acid/10 mg lovastatin tablet; at week 26 of the study, the dose was escalated to 40 mg lovastatin/2,000 mg nicotinic acid. Results showed that the combination therapy is equally efÞcacious and tolerable in patients with or without metabolic syndrome (Simmons PD, 2004). The presence of nicotinic acid in this combination therapy potentially increases the risk of side effects and contraindications. The safety of lovastatin/ER nicotinic acid was recently tested in an open-label study involving 4,499 patients, in which the primary end points were study compliance, increases in liver transaminases greater than three times the upper limit of normal, and clinical myopathy (RubenÞre M, 2004). A dose regimen of one tablet of 20/500 mg lovastatin/nicotinic acid for four weeks, followed by two tablets for eight weeks, resulted in ßushing as the most common side effect (in 18% of patients), raised liver enzymes (in <0.3% patients), and an overall compliance rate of 77%. Uptake of this agent in the United States has been slow, perhaps because of the unpopularity of the statin component (lovastatin). Kos is currently developing a simvastatin/nicotinic acid combination (see “Emerging Therapies”). Atorvastatin/Amlodipine. PÞzer’s combination therapy of atorvastatin∗ (Fig 1) and amlodipine∗ was launched in the United States as Caduet in June 2004 for treatment of hypertension and dyslipidemia. The company is also seeking approval in Europe; France is acting as the reference state in this process. The combination is indicated for use in patients for whom treatment with both agents is appropriate—that is, dyslipidemic patients with comorbid hypertension.

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Atorvastatin is an inhibitor of HMG-CoA reductase. Amlodipine, the antihypertensive component of the combination pill, is a long-acting calcium-channel blocker (CCB). CCBs induce vasodilation by inhibiting the entry of calcium ions into smooth-muscle cells, which causes relaxation of these cells and ultimately lowers blood pressure. Data from the Anglo-Scandinavian Cardiac Outcomes Trial (ASCOT) provide support for the use of statins in combination with antihypertensive medication in patients who have multiple cardiovascular risk factors (Sever PS, 2003). The lipid-lowering arm of this study compared the effect of 10 mg atorvastatin with that of placebo on CHD end points in 9,000 hypertensive patients with total cholesterol greater than 250 mg/dL who were taking an antihypertensive treatment (amlodipine [5/10 mg] ± perindopril [Servier’s Conversyl] [4/8 mg]) (Sever PS, 2001). Atorvastatin-treated patients showed a 36% reduction in nonfatal MI and fatal CHD and a 29% reduction in all coronary events, compared with the placebo arm (p = 0.0005). PÞzer reports that FDA approval of the atorvastatin/amlodipine combination pill was based on a study of 1,600 patients with comorbid hypertension and dyslipidemia. Patients received once-daily treatment with one of four regimens: one of eight dose combinations of amlodipine/atorvastatin (5/10, 10/10, 5/20, 10/20, 5/40, 10/40, 5/80, or 10/80 mg); amlodipine alone (5 or 10 mg); atorvastatin alone (10, 20, 40, or 80 mg); or placebo. The results showed that all combination groups demonstrated statistically signiÞcant dose-related reductions in systolic blood pressure (SBP), diastolic blood pressure (DBP), and LDL, compared with placebo. PÞzer is assessing the atorvastatin/amlodipine combination therapy in approximately 500 African-Americans in the ongoing Clinical Utility of Caduet in Simultaneously Achieving Blood Pressure and Lipid Endpoints in a SpeciÞc Patient Population (CAPABLE) trial. Patient groups will receive one of eight available doses of open-label atorvastatin/amlodipine over the course of 20 weeks. Atorvastatin/amlodipine is generally well tolerated. However, some adverse side effects have been reported, including headache, abdominal pain, nausea, insomnia, dizziness, rash, and chest pain. Fibrates (Fibric Acid Derivatives) Overview. Compared with trials of the statins, few clinical trials have assessed the beneÞts of Þbrates (Þbric acid derivatives) in the treatment of dyslipidemia. Fibrates have traditionally been used to lower TG levels and are currently the agent of choice in patients with severe hypertriglyceridemia. However, recent investigations of these agents have demonstrated reductions in LDL of 10–35%, depending on the Þbrate used—and more signiÞcantly, increases in HDL levels. Also, the publication of several recent clinical trials has spurred some interest in Þbrates (UK HDL-C Consensus Group, 2004). Generally, Þbrates are well tolerated. However, they can cause a myositis-like syndrome (inßammation of muscle tissue), especially in patients with impaired renal function. Fibrates are contraindicated in patients with severe renal and

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FIGURE 14. Structure of fenofibrate.

hepatic impairment. Fibrates may also reduce libido or even cause impotence; these effects often result in noncompliance with therapy. Other common side effects of Þbrates include nausea, loss of appetite, and gastric pain. Fibrates are not a homogeneous class of drugs; they differ in their pharmacokinetic proÞles and potency. Consequently, use of individual Þbrates in dyslipidemia treatment varies considerably, with most doctors now favoring the more potent later-generation Þbrates, such as fenoÞbrate (Fournier’s Lipantil/Abbott’s Tricor, generics) and bezaÞbrate (Roche’s Bezalip, generics). This section discusses in detail the three most commonly used members of this drug class: fenoÞbrate, bezaÞbrate, and gemÞbrozil (PÞzer’s Lopid, generics). Other Þbrates include ciproÞbrate (SanoÞ-Aventis’s Modalim and Lipanor). Mechanism of Action. Fibrates exert their effects via activation of the peroxisome proliferator activated receptor alpha (PPARα). Activation of this receptor leads to alterations in expression of genes involved in lipid metabolism, causing an increase in HDL cholesterol and a reduction in TGs. Newer members of the Þbrate class, including fenoÞbrate, have also been shown to reduce LDL cholesterol and the proportion of harmful, small, dense LDL particles (UK HDL-C Consensus Group, 2004). Fenofibrate. FenoÞbrate (Fournier’s Lipantil/Abbott’s Tricor, generics) (Figure 14) was approved for treatment of dyslipidemia in Europe in 1975 and in the United States in 1998. In 2004, Abbott received FDA approval for a new formulation of Tricor that enables patients to take the drug without food, thereby simplifying the treatment regimen in an effort to improve patient compliance. FenoÞbrate activates the PPARα receptor, altering genes involved in lipid metabolism. It has been shown to lower TG levels signiÞcantly and to achieve reductions in LDL levels of up to 35%; it has also been shown to increase HDL levels more than statins. Also, studies have reported that fenoÞbrate treatment produces inßammatory effects and improves endothelial function (Kon Koh K, 2004; Mareshi S, 2003). Several randomized trials have assessed the efÞcacy of fenoÞbrate in patients with dyslipidemia. One study investigated patients with type IIa or IIb hypercholesterolemia who were recruited from 11 medical centers. Two hundred and twenty-seven patients were randomized to receive either placebo or 100 mg fenoÞbrate three times daily for 24 weeks. A subpopulation of 192 patients was studied for an additional 24 weeks, during which all patients received

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fenoÞbrate open-label. In patients with type IIa hypercholesterolemia, signiÞcant reductions were observed in TSC (−18%), LDL (−20%), very-low-density lipoprotein (VLDL) (−38%), and TGs (−38%), compared with baseline. Mean plasma HDL increased by 11%. Similar effects were seen in the 24 patients with type IIb hypercholesterolemia, with the exception of effects on LDL, which was low to start with in these patients. No signiÞcant effects were seen in the placebo group. The trend toward change was observed throughout the open-label study period; further reductions in TSC and LDL were observed in the patients who had received fenoÞbrate during the initial 24 weeks of the study (Brown WV, 1986). The Diabetes Atherosclerosis Intervention Study (DAIS), an angiographic study, sought to determine whether fenoÞbrate therapy affects coronary atherosclerosis in patients with type 2 diabetes. The study found that fenoÞbrate therapy signiÞcantly increased HDL levels (p < 0.001 versus placebo) and that the increase in percentage diameter stenosis was signiÞcantly smaller in the fenoÞbrate group than in the placebo group (DAIS Investigators, 2001). Early clinical trials revealed problems with the combination of statins and Þbrates. With some combinations, despite reductions in lipid levels, there appeared to be an increased potential for development of severe myopathies (UK HDL-C Consensus Group, 2004). However, other investigations have proved more promising. One randomized, placebo-controlled, cross-over study examined the actions of a combination of low-dose simvastatin and fenoÞbrate in the treatment of combined dyslipidemia. The trial had three randomized phases, each lasting three months: double placebo, 10 mg/day simvastatin plus placebo, and 10 mg/day simvastatin plus 200 mg/day fenoÞbrate. The combination of simvastatin and fenoÞbrate was not associated with an increase in adverse events or serious side effects such as myopathy; furthermore, a statistically signiÞcant increase in HDL (23%) was seen with the combination treatment, compared with the statin-only treatment (Vega GL, 2003). The recent Simvastatin Plus FenoÞbrate for Combined Hyperlipidemia (SAFARI) study has provided similar positive results for a regimen of combined simvastatin and fenoÞbrate. Patients were randomized to receive 20 mg simvastatin monotherapy daily (n = 207) or combined 20 mg simvastatin and 160 mg fenoÞbrate (n = 411) for 12 weeks. No liver function abnormalities or myopathy were reported, and HDL levels increased by 18.6% in the combination therapy group versus 9.7% in the simvastatin monotherapy group (Grundy SM, 2005). A pooled analysis of clinical trials that assessed various Þbrates, including fenoÞbrate in combination with ßuvastatin, also demonstrated safety and efÞcacy in patients with mixed dyslipidemia or primary hypercholesterolemia (Farnier M, 2003). Further large-scale trials of this nature are certainly warranted, particularly in patients with combined dyslipidemia, who stand to beneÞt most from this treatment approach. Bezafibrate. BezaÞbrate (Roche’s Bezalip, generics) (Figure 15) is another commonly used Þbrate. Like other Þbrates, bezaÞbrate activates the PPARα receptor, altering genes involved in lipid metabolism. Clinical trial data assessing

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FIGURE 15. Structure of bezafibrate.

the efÞcacy of this agent in dyslipidemic patients are limited compared with data available for the statins; however, this section does discuss several key studies. The double-blind BezaÞbrate Infarction Prevention (BIP) study investigated the effects of 400 mg of bezaÞbrate per day versus placebo in 3,090 patients with CHD. Results showed a 9% reduction in coronary events, which was not signiÞcant; however, a 39% reduction in events was observed in patients with TGs < 2.2 mmol/l (<85 mg/dL). Also, HDL levels were signiÞcantly increased (18%) with bezaÞbrate treatment (BIP Investigators, 2000; UK HDL-C Consensus Group, 2004). Studies have also assessed the efÞcacy and safety of bezaÞbrate in combination with a statin in patient groups that may beneÞt from such a treatment approach. One open-label, 21-month investigation examined the actions of bezaÞbrate in combination with simvastatin in 148 patients who had type 2 diabetes with diabetic dyslipidemia (controlled by diet and oral hypoglycemic agents). Forty-eight patients received slow-release bezaÞbrate (400 mg/day), and 100 patients received simvastatin (20 mg/day) for a total of six months; after the initial treatment period, all patients were switched to a combination of simvastatin (20 mg/day) and bezaÞbrate (400 mg/day). Treatment with the combination therapy reduced TSC by 23%, TGs by 42%, and LDL by 25%, and increased HDL by 10%. A 19% reduction in Þbrinogen and lipoprotein(a) (Lp[a]) levels was also observed. The rate of cardiovascular events was reduced from 9.5% during the Þrst six months of the investigation to less than 2% during the combination treatment period. Two patients developed myopathy on the combined regimen, and one patient treated with the statin alone developed myopathy. Overall, the combination appeared efÞcacious and safe (Gavish D, 2000). Further studies are required owing to a renewed interest in statin/Þbrate combinations. Gemfibrozil. GemÞbrozil (PÞzer’s Lopid, generics) (Figure 16) is a well-established member of the Þbrate class that has been approved in every major market under study except Japan; the drug was Þrst approved by the FDA for the treatment of dyslipidemia in 1982. Like other Þbrates, gemÞbrozil activates the PPARα receptor, altering genes involved in lipid metabolism. However, gemÞbrozil differs from other agents in this class in one respect: usually, only modest decreases in LDL are observed with gemÞbrozil treatment. The most important primary prevention trial that examined Þbrates is the Helsinki Heart Study, which investigated gemÞbrozil. This randomized,double-blind,

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FIGURE 16. Structure of gemfibrozil.

placebo-controlled investigation involved 4,081 male patients aged 40–55; the follow-up period was Þve years. This study showed that patients treated with gemÞbrozil experience a 34% reduction in CHD events but do not experience a reduction in total mortality (Fick MH, 1987). In patients whose baseline HDL level was less than 1.08 mmol/L (40 mg/dL), a 44% reduction in CHD events was observed (UK HDL-C Consensus Group, 2004). The Veterans Administration-HDL-Cholesterol Intervention Trial (VA-HIT), a randomized, placebo-controlled study, enrolled 2,531 male patients with a combination of risk factors (low HDL, raised TGs, and average to low LDL) who were followed for a mean of 5.1 years. In this study, gemÞbrozil (1,200 mg/day) signiÞcantly reduced the relative risk of major coronary and cardiovascular events by 22% and 24%, respectively. Therapy raised HDL cholesterol levels (6%) and lowered TG levels (31%) without lowering LDL cholesterol concentrations (Rubins HB, 2000). As previously stated, several studies have reported that combination therapy with statins and Þbrates (including the combination of gemÞbrozil and lovastatin) is effective in lowering TGs and LDL cholesterol (Illingworth DR, 1989). However, combination therapy regimens involving gemÞbrozil have historically proved to have a potential for side effects, including myopathy (UK HDL-C Consensus Group, 2004). Second-generation Þbrates such as fenoÞbrate are therefore more likely to be used in combinations than gemÞbrozil, unless further trials that demonstrate the safety of gemÞbrozil become available. Cholesterol Absorption Inhibitors Overview. Dietary and biliary cholesterol is absorbed in the form of bile acid micelles into enterocytes in the small intestine. Recent evidence suggests that this process is not passive; rather, it is mediated via speciÞc cholesterol transporters. This uptake process therefore presents a potential target for cholesterollowering agents. The use of a cholesterol absorption inhibitor in combination with a statin may result in additive or synergistic cholesterol-lowering effects because statins inhibit the endogenous hepatic production of cholesterol. Ezetimibe (Merck/Schering-Plough’s Zetia/Ezeterol) was the Þrst selective cholesterol absorption inhibitor to reach the market, in 2002. Mechanism of Action. The mechanisms involved in cholesterol uptake from the gut lumen are not well deÞned. However, in a publication in the journal Science, researchers demonstrate that the protein Niemann-Pick C1 Like-1

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(NCP1L1) plays a key role in the process. Expression of NCP1L1 is abundant in the small intestine and is found in the brush border membrane of enterocytes. NCP1L1 knockout mice absorb less cholesterol from the gut, which is unaffected when bile acids are supplemented into the diet. The cholesterol absorption inhibitor ezetimibe had no effect on serum cholesterol levels in these animals, which suggests that this protein is crucial to the drug’s mechanism of action (Altmann SW, 2004). Ezetimibe. Developed as a joint venture between Merck and Schering-Plough, ezetimibe (Zetia/Ezeterol) (Figure 12) was launched in the United States and Germany in October 2002 and is now available in France, Spain, and the United Kingdom through the mutual recognition procedure. Bayer will comarket ezetimibe in Japan, where the agent is undergoing regulatory review. The agent’s mechanism of action is not fully understood; however, identiÞcation of the involvement of NCP1L1 in cholesterol absorption processes has provided some new insight. Many clinical trials have been carried out to assess the potential of this agent in the treatment of dyslipidemia. In a dose-ranging study, 243 patients with baseline LDL of 130–250 mg/dL and TGs of less than 300 mg/dL were randomized to receive either placebo or ezetimibe (0.25, 1, 5, or 10 mg), administered once daily. A second arm treated 189 patients with either placebo or ezetimibe (5 or 10 mg). When data were pooled, after 12 weeks of treatment, reductions in LDL of 15.7% and 18.5% were observed in the 5 mg and 10 mg groups respectively, compared with the placebo group. HDL was raised by 2.9% and 3.5% in the 5 mg and 10 mg dose groups, compared with the placebo group. Statistically insigniÞcant decreases in TG levels were also observed (Bays HE, 2001). The efÞcacy of ezetimibe in lowering cholesterol levels has been assessed in combination with statin therapy in several randomized, placebo-controlled studies. The six-week, Ezetimibe Add-on to Statin for Effectiveness (EASE) randomized, multicenter clinical trial included 3,030 patients with LDL levels ranging from 123–167 mg/dL who were currently taking marketed statins (atorvastatin [40%], simvastatin [29%], pravastatin [22%], or other [10%] [n = 2, 020]), or placebo (n = 1, 010) (Gagne C, 2002[a]; Pearson TA, 2005). The study population included both men and women and was multiracial (Caucasians, AfricanAmericans, and Hispanics); the mean age was 62 years. Thirty-eight percent of patients were diabetic, and 60% of patients fulÞlled the NCEP ATP III metabolic syndrome criteria. Results showed that addition of 10 mg of ezetimibe to a stable dose of any of the statins reduced LDL by an additional 26%, compared with a 3% reduction seen with placebo (p < 0.001). SigniÞcant reductions in LDL were observed irrespective of age, gender, race, CHD risk category, statin brand, statin dose, or the presence of diabetes or metabolic syndrome. One study assessed ezetimibe in conjunction with atorvastatin in 621 patients with LDL ≥130 mg/dL; all the patients in the study had one of the following: CHD, at least two cardiovascular risk factors, or heterozygous familial hypercholesterolemia. Patients received open-label 10 mg atorvastatin and were then randomized to receive either 10 mg ezetimibe or an additional 10 mg atorvastatin.

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In patients who did not achieve the target LDL goal of ≤100 mg/dL, atorvastatin was titrated to a maximum of 40 mg/day in the combined treatment group and to a maximum of 80 mg/day in the monotherapy group. After 14 weeks, 22% of the combined therapy group reached the LDL goal versus 7% of patients receiving monotherapy (Stein E, 2004). A recent study has tested the strategy of combining ezetimibe with simvastatin therapy. Patients with documented CHD and an LDL range of 2.6–4.2 mmol/L (100 mg/dL −162 mg/dL) (n = 372) were randomized to receive, for six weeks, either 10 mg ezetimibe (n = 181) or placebo (n = 191) coadministered with 10 mg or 20 mg simvastatin, which had been previously administered in a fourweek, open-label run-in period (Farnier M, 2005). After six weeks, the percentage of patients who achieved the primary end point—LDL ≤100 mg/dL—was signiÞcantly higher in the ezetimibe group than in the placebo group (74.3% versus 16.7%). The effect of ezetimibe was consistent across both doses of simvastatin. The average LDL reduction from baseline in the ezetimibe plus simvastatin cohort was 25.2%, compared with 0.9% in the placebo plus simvastatin group. There were no signiÞcant differences in side effects between the two groups. In clinical trials, ezetimibe has a side-effect proÞle similar to that of placebo. Gastrointestinal disorders were reported in approximately 1% of patients (Gagne C, 2002). A slight increase in serum transaminases has been reported in patients receiving concurrent ezetimibe and statin therapy. Myopathy and rhabdomyolysis have not been associated with ezetimibe treatment. The safety and effectiveness of ezetimibe in combination with Þbrates has not been established. Nicotinic Acid Derivatives Overview. Despite being a low-cost and effective agent with a superior ability to increase HDL levels and modulate Lp(a), nicotinic acid (also known as niacin) is infrequently prescribed outside the United States. Nicotinic acid’s clinical use is limited owing to its side-effect proÞle, which can create compliance issues. Vasodilatation (which can lead to skin ßushes resulting from increased production of prostaglandins), headaches, dizziness, and palpitations are particularly worrying to patients. In the United States, available products include several immediate-release, nonprescription, generic formulations, and Upsher-Smith’s controlled-release OTC preparation, Slo-Niacin, which must be administered twice daily. In 1997, Kos Pharmaceutical’s Niaspan, a reformulated, once-daily, ER form of nicotinic acid to be taken at bedtime, was approved for prescription use only in the United States for treating mixed lipid disorders. Niaspan is now available in several European markets. Mechanism of Action. Nicotinic acid lowers both cholesterol and TG concentrations by inhibiting their synthesis, but the precise mechanism of action is unknown. With use of nicotinic acid, reductions in LDL concentrations tend to be paralleled by increases in HDL concentrations. A 1 g dose of nicotinic acid produces an average 10–20% reduction in LDL, a 30–70% reduction in TGs, and a 20–35% increase in HDL. Treatment with nicotinic acid also decreases levels of ApoB-100 and of Lp(a), an independent coronary risk factor.

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Extended-Release Nicotinic Acid. As mentioned previously, several brands of nicotinic acid (or niacin) (Figure 13) are currently marketed, including two ER products: Upsher-Smith’s Slo-Niacin and Kos’s Niaspan, a once-daily formulation that is now available in the United States, France, Germany, and the United Kingdom. Niaspan is associated with a lower incidence of ßushing and liver function abnormalities than other brands and is therefore discussed in this section. (In 2002, as discussed earlier, a Þxed combination of ER nicotinic acid plus lovastatin [Advicor] was launched in the United States.) The mechanism of action of all forms of nicotinic acid, including the controlled-release agents, has not been fully established. Clinical studies of ER nicotinic acid have been positive. A 25-week clinical study of 131 patients with primary hypercholesterolemia demonstrated that Niaspan (500–3,000 mg/day) reduced LDL cholesterol by up to 21%, TGs by up to 44%, and Lp(a) by up to 26%. HDL was increased by up to 30% (Goldberg A, 2000). Kos reports an 80% decrease in the incidence of ßushing with this controlled-release formulation, compared with other forms of nicotinic acid. Combinations of low doses of nicotinic acid and statins promise the same beneÞcial effects on HDL and TG levels that nicotinic acid alone provides, but with fewer side effects. Data from the Arterial Biology for the Investigation of the Treatment Effects of Reducing Cholesterol (ARBITER 2) study were presented at the 2004 ScientiÞc Sessions of the American Heart Association. The oneyear study tracked the progression of atherosclerosis, measured by the change in carotid intima-media thickness (CIMT), in 167 patients with established CHD and low HDL (<45 mg/dL) who received 1,000 mg Niaspan in addition to background statin therapy. Patients receiving Niaspan plus statin therapy showed a lower rate of disease progression than the group receiving statin therapy alone (Taylor AJ, 2004). Nicotinic acid given in conjunction with simvastatin in the three-year HDLAtherosclerosis Treatment Study (HATS) resulted in an average increase in HDL of 26% in patients with conÞrmed CHD, normal LDL (≤145 mg/dL), and low HDL (≤35 mg/dL) (Brown BG, 2001). There was no statistical difference in the incidence of ßushing observed in HATS between the simvastatin plus nicotinic acid group and the control groups (30% versus 23%, p = 0.28). In the treatment group, glycemic control in diabetic patients declined initially but returned to pretreatment levels after eight months and remained stable for the rest of the study. Although nicotinic acid is contraindicated in diabetes patients because of its potential to increase blood sugar levels, its beneÞcial effects on HDL cholesterol and TG levels in patients with diabetes and peripheral arterial disease have been demonstrated. The Arterial Disease Multiple Intervention Trial (ADMIT) studied 468 patients, including 125 diabetics, with peripheral arterial disease. In patients with diabetes, nicotinic acid increased HDL by 29% and reduced TGs by 23%. Although glucose levels increased in both cohorts, there were no signiÞcant differences in nicotinic acid discontinuation or hypoglycemic therapy between diabetic patients who received nicotinic acid and diabetic patients who received placebo (Elam MB, 2000).

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Bile Acid Sequestrants Overview. Before the introduction of statins, bile acid sequestrants, also known as resins, were frequently used as Þrst-line pharmacological therapy for patients with hypercholesterolemia. However, since the introduction of the statins, the use of bile acid sequestrants has declined considerably because of their unfavorable side-effect proÞle (common side effects include bloating, abdominal discomfort, nausea, and constipation), the need for multiple daily dosing, and their lack of potency. Often reformulated from dry powders, they are awkward to administer and are frequently unpalatable; these factors create compliance problems. Bile acid sequestrants are now generally reserved for use in patients younger than age 20 and in women of childbearing age, because they act locally in the gut; therefore, they do not cause side effects such as reproductive toxicity. These agents may also be used in combination with a statin in patients with extremely high cholesterol levels (1,000–2,000 mg/dL). Bile acid sequestrants commonly used in the United States and Europe are cholestyramine (Bristol-Myers Squibb’s Questran, generics) and colestipol (PÞzer’s Colestid). Colestimide (also known as colestilan; Mitsubishi Pharma’s Cholebine) is commonly prescribed in Japan. Cholestyramine, colestipol, and colestimide are equally effective in reducing cholesterol levels. In September 2000, Genzyme and its marketing partner Sankyo launched a new bile acid sequestrant, the nonabsorbed hydrogel resin colesevelam (Welchol, formerly Cholestagel), in the U.S. market, thereby fueling some renewed interest in this drug class. In March 2004, colesevelam received European regulatory approval; Genzyme is seeking to outlicense product rights in Europe. Mechanism of Action. Bile acid sequestrants act to reduce cholesterol by interrupting enterohepatic circulation of bile acids. This action promotes excretion of both cholesterol and bile acids as well as increased synthesis of bile acids in hepatocytes. Because use of these agents results in removal of bile acids from the system, the liver increases the requirement for the existing cholesterol pool to form more bile acids. To compensate for the fall in cholesterol levels, the liver increases production of LDL receptors; consequently, hepatocytes take up more cholesterol. The increased number of LDL receptors, in turn, leads to increased clearance of LDL from the plasma. Therefore, despite increased cholesterol uptake, overall cholesterol and LDL levels decrease as a result of faster clearance of LDL. Bile acid sequestrants’ maximum lowering effect on LDL levels occurs within two weeks of therapy initiation, when levels have typically dropped by 20%. However, these drugs may increase TGs, thereby making them unsuitable for use in patients with hypertriglyceridemia. Colesevelam. Colesevelam (Genzyme/Sankyo’s Welchol) (Figure 17) is the latest bile acid sequestrant to be approved. Since its initial launch in the United States in September 2000, colesevelam has garnered a signiÞcant share of the bile acid sequestrant market at the expense of older bile acid sequestrants such as cholestyramine and colestipol.

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FIGURE 17. Structure of colesevelam.

Like the other members of the bile acid sequestrant class, colesevelam interrupts the enterohepatic circulation of bile acids. Several clinical trials have been conducted to assess the efÞcacy and safety of this agent as a monotherapy and in combination with a statin, focusing on patients with type IIa primary hypercholesterolemia. In a double-blind, placebo-controlled trial involving 494 patients with primary hypercholesterolemia, colesevelam lowered mean LDL cholesterol levels by 9–18% in a dose-dependent manner. The reduction in LDL cholesterol levels was greatest after 2 weeks and was sustained throughout 24 weeks of treatment. The agent was shown to be well tolerated, without incidence of adverse events (Insull W, 2001[b]). In a randomized, double-blind, placebo-controlled trial involving 94 primary hypercholesterolemic patients, colesevelam (3.8 mg/day) was investigated in combination with atorvastatin (10 mg/day); a 48% reduction in LDL was seen, as opposed to 38% for the statin alone at the 10 mg/day dose. The combination of lipid-lowering agents was found to be well tolerated (Hunninghake D, 2001). As previously mentioned, clinical studies of older bile acid sequestrants reported mild increases in TG levels. Genzyme reports that in clinical trials of colesevelam, although modest increases were seen compared with baseline, these changes were not dose-related and were not signiÞcantly different from

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FIGURE 18. Structure of cholestyramine.

changes observed with placebo treatment. These observations, together with the drug’s positive safety record in clinical studies both as a monotherapy and in combination with a statin, may make colesevelam more attractive to physicians. Cholestyramine. Cholestyramine (Bristol-Myers Squibb’s Questran, generics) (Figure 18) is a well-established bile acid sequestrant. Like other bile acid sequestrants, cholestyramine interrupts the enterohepatic circulation of bile acids. The Lipid Research Clinic Coronary Primary Prevention Trial (LRC-CPPT) assessed cholestyramine’s (24 g/day) lipid-lowering efÞcacy in reducing the risk of CHD in 3,806 asymptomatic, middle-aged men with hypercholesterolemia. Patients treated with cholestyramine had 8.5% and 12.6% greater reductions in TSC and LDL than patients in the placebo group. The cholestyramine group also had a 19% reduction in risk of the primary endpoints: conÞrmed CHD, death, or nonfatal MI (p < 0.05) (Rifkind BM, 1984). Cholestyramine has also been assessed in combination with a statin. One study of 2,012 patients assessed the effectiveness of pravastatin and cholestyramine combined in the primary care setting. The study showed that, although not statistically signiÞcant, there was a trend toward a better cholesterol-lowering effect when patients received both agents (Eriksson M, 1998). Cholestyramine is associated with a collection of adverse side effects, including constipation, abdominal pain, bloating, weight loss, and ßatulence, which can often reduce patient compliance. Also, several drug interactions have been observed in cholestyramine treatment, such as reduced absorption of vitamin K antagonists, cardiac glycosides, and thiazide diuretics. These agents should therefore be administered one hour before or four to six hours after cholestyramine. Nonpharmacological Approaches Lifestyle Modifications. Lifestyle changes—including diet modiÞcation, weight reduction, exercise, and management of associated risk factors such as smoking—are the Þrst steps in managing dyslipidemia, both in primary and secondary prevention. The therapeutic lifestyle changes (TLCs) recommended in the NCEP ATP III guidelines suggest a diet low in cholesterol (less than 200 mg/day), low in total fats (25–35% of entire diet), and low in saturated fats (less than 7% of the entire diet). Japanese guidelines also recommend a diet low in cholesterol (less

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than 300 mg/day). If initial TLC changes are unsuccessful, ATP III guidelines additionally recommend the introduction of plant stanols/sterols (which have been shown to lower LDL cholesterol by 6–15%, with little or no change in HDL or TG) and soluble Þbre (10–25 g/day). A meta-analysis of the effects of reduced fat intake on cardiovascular risk reduction found a 9% reduction in mortality and a 16% reduction in cardiovascular events (Hooper L, 2001). Likewise, the Lyon Diet Heart Study reported a 50–70% reduction in recurrent coronary events in patients who adopted a Mediterranean-style diet. These results suggest that signiÞcant dietary modiÞcations may lead to beneÞts equivalent to those achieved by pharmacotherapies (Kris-Etherton P, 2001; Robertson RM, 2001). More-recent studies show that the usefulness of dietary modiÞcation is limited. Although diet modiÞcation reduces lipids about as much as pharmacotherapy, dietary modiÞcations lack certain antiatherogenic beneÞts associated with pharmacological treatment: patients on certain pharmacotherapies experience less inßammation, less calciÞcation of atherosclerotic plaques, and less need for revascularization than patients depending on dietary modiÞcation alone. The latter group also shows a continual elevation of inßammatory markers over Þve years (Fonarow GC, 2003). Lipid Apheresis and Plasma Exchange. Approximately 1–3% of patients refractory to pharmacological treatment (usually patients who have severe, rare, genetic abnormalities) may undergo lipid apheresis or plasma exchange. Both procedures involve the “puriÞcation” of the plasma. Lipid apheresis entails removal of the ApoB-100 (which contains VLDL, intermediate density lipoprotein [IDL], and LDL) from the plasma. Plasma exchange involves removal of plasma and replacing it with plasma or albumin containing normal cholesterol levels. Such procedures are conducted only in patients with phenomenally high lipid deposits (i.e., plasma cholesterol of 500–600 mg/dL) and then only rarely because the procedures are extremely expensive. EMERGING THERAPIES The introduction of statins in the late 1980s revolutionized therapy for dyslipidemia, particularly for patients with elevated low-density lipoprotein (LDL) levels. The focus of R&D today is on novel treatments for non-LDL lipid abnormalities, including raised triglycerides (TGs) and lowered high-density lipoprotein (HDL). This focus will drive the use of combination therapies, in which the effects of statins are supplemented by agents that affect one or more changes in lipid levels—be it the reduction of LDL or TGs or the increase of HDL—via other mechanisms of action. In addition to the combination therapies, drug classes in development include the cholesterol ester transfer protein (CETP) inhibitors, reverse lipid transport pathway (RLTP) activators, and peroxisome proliferator activated receptor (PPAR) agonists. Other agents in clinical development include acyl-CoA cholesterol acyltransferase (ACAT) inhibitors, microsomal triglyceride transport protein inhibitors, squalene

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synthase inhibitors, lipoprotein lipase (LPL) activators, lipoprotein (a) antagonists, and bile acid modulators. Antioxidant and vascular protectant approaches to lipid modulation have spurred some interest; however, the lead compound in this class, AGI-1067, is no longer in development for dyslipidemia, although it continues in Phase III trials for atherosclerosis. The cannabinoid receptor antagonist, rimonabant (SanoÞ-Aventis), is another agent that has demonstrated lipid-modulating properties in the clinical setting, but is not in development for dyslipidemia and as such is not discussed in detail here. Data from large-scale trials of rimonabant for the treatment of obesity and smoking cessation have demonstrated that rimonabant improves HDL by 10–25%, and also reduces TGs. Research into nuclear hormone receptors has identiÞed several potential drug targets that affect non-LDL dyslipidemia, hepatic lipogenesis, and insulin sensitivity. Nuclear hormone receptors in development include farnesoid X receptors, retinoid X receptors, and liver X receptors. Agonists of PPARs, which belong to this family of receptors, have reached late-stage clinical development. Farnesoid X receptors and liver X receptors are nuclear receptors for bile acids; activation of these receptors affects bile acid, lipid, and carbohydrate metabolism (DuranSandoval D, 2005; Zhao A, 2004). In addition, liver X receptors raise HDL by regulating reverse cholesterol transport (Miao B, 2004). Retinoids, derivatives of vitamin A, induce hypertriglyceridemia through reduced clearance of very-lowdensity lipoprotein (VLDL) by a lipoprotein lipase (LPL)-dependent pathway. The retinoid X receptors may be important in mediating the effects of retinoids on TG clearance (Haugen BR, 2004). Currently, agonists for farnesoid X receptors and retinoid X receptors are in development by biotechnology companies. Allergen and X-Ceptor Therapeutics are developing farnesoid receptor agonists, and MaxoCore Pharmaceuticals has a retinoid receptor agonist program. GlaxoSmithKline, Tularik, and CV Therapeutics are researching the potential for liver X receptor modulators in dyslipidemia treatment. Other research is targeting apolipoprotein subfractions, including modulation of Apo-A1, Apo-B100, and Apo-E. GlaxoSmithKline and Avanir Pharmaceuticals are actively pursuing preclinical programs for Apo-A1 modulators, and Phaedrus Pharmaceuticals has two Apo-E modulators—PPI-01 and PPI-02—in preclinical development. Modulators for Apo-B100 have reached clinical development: Isis Pharmaceuticals is developing the Apo-B100 modulator ISIS-301012 in Phase I studies. These agents are not discussed further in this this section because they are still in early development. Table 5 lists emerging therapies in development for the treatment of dyslipidemia. Statin Combination Therapies Overview. The development of Vytorin—Merck and Schering Plough’s simvastatin/ezetimibe Þxed-dose, single-pill combination product—has proved to be

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TABLE 5. Emerging Therapies in Development for Dyslipidemia Compound Statin combination therapies Atorvastatin/torcetrapib United States Europe Japan Simvastatin/nicotinic acid (ER) United States Europe Japan

Development Phase

Marketing Company

III — —

Pfizer — —

III — —

Kos — —

Cholesteryl ester transfer protein inhibitors JTT-705 United States II Europe II Japan I CETi-1 United States S Europe — Japan —

Avant Immunotherapeutics — —

Reverse lipid transport pathway activators ETC-1001 United States I Europe — Japan —

Pfizer — —

Acyl-CoA cholesterol acyltransferase inhibitors Eflucimibe United States II Europe — Japan —

Pierre Fabre/Eli Lilly — —

Pactimibe United States Europe Japan

II II PC

Peroxisome proliferator activated receptor agonists Tesaglitazar United States III Europe — Japan — GW-501516 United States Europe Japan

— II —

Microsomal triglyceride transfer protein inhibitors Implitapide United States — Europe II Japan —

Roche Roche Japan Tobacco

Sankyo/Kyoto Sankyo/Kyoto Sankyo/Kyoto

AstraZeneca — — — GlaxoSmithKline/Ligand Pharmaceuticals —

— Bayer —

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TABLE 5. (continued) Compound

Development Phase

Marketing Company

Squalene synthase inhibitors TAK-475 United States Europe Japan

I II PC

Takeda Takeda Takeda

Lipoprotein lipase activators Ibrolipim United States Europe Japan

II — —

Otsuka/TAP Pharmaceuticals — —

Lipoprotein (a) antagonists Gemcabene United States Europe Japan

II — —

Pfizer — —

Bile acid reabsorption inhibitors S-8921 United States Europe Japan

— II S

— Shionogi Shionogi

PC = Preclinical (including discovery); S = Suspended.

a successful franchise protection strategy. Accordingly, other such combination therapies will likely prove popular, adding market life to current agents through enhanced efÞcacy and extended therapeutic range. These single-pill combinations may include atorvastatin with the CETP inhibitor torcetrapib and simvastatin with the extended-release (ER) nicotinic acid derivative Niaspan (Kos/Merck KGaA). A combination of pravastatin with the antihypertensive angiotensin-II receptor antagonist, olmesartan, is in preclinical development. Mechanism of Action. Single-pill combination therapies incorporate the mechanisms of action of each individual agent into a single-pill formulation. Atorvastatin/Torcetrapib. In the United States, PÞzer is developing a Þxeddose combination of atorvastatin and the CETP inhibitor, torcetrapib∗ , for dyslipidemia. PÞzer has completed enrollment for the Phase III ILLUMINATE study, a double-blind, randomized trial of the combination product compared with atorvastatin alone; this trial will measure the occurrence of major cardiovascular events in 13,000 CHD patients or risk-equivalents over a Þve-year period. In addition to standard lipid parameters, the atorvastatin/torcetrapib development program involves use of intravascular ultrasound and carotid ultrasound technology for efÞcacy studies. Such a therapy will have a dual mechanism of action: HMG-CoA reductase inhibition and cholesterol ester transfer protein (CETP) inhibition (see the section “CETP Inhibitors”). Its effect will be to reduce LDL and increase HDL.

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Company-reported data indicate that the combination—if used across the entire statin dose range—can raise HDL by more than 50% while lowering LDL as much as 80%. In trials in which atherosclerotic patients were given 90 mg/day of torcetrapib in combination with 20 mg/day of atorvastatin, the addition of torcetrapib generated an incremental 15% reduction in LDL over the reduction achieved with the statin monotherapy; the HDL-raising efÞcacy of torcetrapib was not adversely affected in combination with atorvastatin. A recent, single-blind, placebo-controlled clinical study investigated the actions of torcetrapib both as a monotherapy and in combination with atorvastatin (20 mg) (Brousseau ME, 2004). Nineteen patients were enrolled; nine received atorvastatin and ten did not. All patients received placebo for the Þrst four weeks of the study, followed by 120 mg of torcetrapib daily for an additional four weeks. Six patients in the non-atorvastatin arm received 120 mg of torcetrapib twice daily for an additional four weeks. Results showed a 61% increase in plasma HDL in the atorvastatin group following treatment with 120 mg of torcetrapib daily; a 46% increase was seen in patients not on statin therapy. Treatment with 120 mg torcetrapib twice daily during the additional study period increased HDL by 106%. A further reduction in LDL levels of 17% was observed in the atorvastatin/torcetrapib treatment arm (p = 0.02). Coronary heart disease (CHD) is generally associated with low levels of large HDL particles and higher levels of small, dense LDL particles; in this study, the number of large HDL particles increased signiÞcantly with torcetrapib treatment. Torcetrapib was well tolerated. Further analysis of the effects of torcetrapib on ApoA-1 in patients in this study show that twice-daily administration of 120 mg of torcetrapib increases the amount of ApoA-1 in alpha-migrating HDL by 382%, compared with placebo (Brousseau ME, 2005). (ApoA-1 is the major protein component of HDL. Studies show that levels of this agent are inversely proportional to CHD incidence [Luc G, 2002].) The effects of torcetrapib monotherapy have been highlighted in several clinical studies. A recently published Phase I study investigated the HDL-raising properties of torcetrapib in healthy young adults (Clark RW, 2004). Five groups of eight persons were randomized to receive 10, 30, 60, or 120 mg daily for 14 days or 120 mg twice daily for 14 days. CETP inhibition with this agent dose-dependently increased HDL by 16–91%. In the group treated with 120 mg torcetrapib twice daily, Apo A-I and E were elevated 27% and 66%, respectively, and Apo-B100 was reduced 26%. Researchers note that the effects of CETP inhibition are similar to those observed in CETP-deÞcient subjects. PÞzer is involved in several imaging trials at the Cleveland Clinic in the United States; the company is looking at changes in plaque morphology associated with torcetrapib treatment. The FDA requires these studies to support the use of combination therapies. The studies use intravascular ultrasound screening (IVUS) and intima to media thickness (IMT) imaging. Whether these studies will prove sufÞcient for FDA approval is unknown. Because studies using IVUS are susceptible to interoperator variation, large, multicenter clinical trials are not possible, thus limiting the patient pool.

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Questions also remain about the side-effect proÞle of torcetrapib. Occasional, unexplained increases in blood pressure have been noted in patients taking torcetrapib in Phase II studies (Davidson M, 2005). Simvastatin/Nicotinic Acid (ER). Kos is developing a combination of its extended-release (ER) nicotinic acid (Figure 13) tablet, Niaspan, and simvastatin. The company is conducting two pivotal U.S. Phase III trials: the open-label Evaluation of the Safety and EfÞcacy of a Combination of Niacin ER and Simvastatin in Patients with Dyslipidemia (OCEANS) and the Safety & EfÞcacy of a Combination Niacin ER/Simvastatin in Patients with Dyslipidemia (SEACOAST) dose-ranging study. Kos has also initiated a study in metabolic syndrome patients: the Atherothrombosis Intervention in Metabolic Syndrome with Low HDL-C/High Triglyceride and Impact on Global Health Outcomes (AIM-HIGH) trial will follow 3,300 patients over six years to evaluate the beneÞts of treating low HDL and high TG levels in patients who have attained guideline-deÞned LDL targets. Kos expects to submit a new drug application (NDA) in 2006 and to launch the agent in 2007, in line with simvastatin’s patent expiry date. Data from several small studies support the beneÞt of raising HDL in conjunction with statin therapy. The one-year Arterial Biology for the Investigation of the Treatment Effects of Reducing Cholesterol (ARBITER 2) study tracked the progression of atherosclerosis, measured by the change in carotid intimamedia thickness (CIMT), in 167 patients with established CHD and low HDL (<45 mg/dL) who received 1,000 mg Niaspan in addition to background statin therapy. Patients receiving Niaspan plus statin therapy showed a lower rate of disease progression than the group receiving statin therapy alone (Taylor AJ, 2004). ER nicotinic acid (Upsher Smith’s Slo-Niacin), given in conjunction with simvastatin in the three-year HDL-Atherosclerosis Treatment Study (HATS), resulted in an average 26% increase in HDL in patients with conÞrmed CHD, normal LDL (≤145 mg/dL) and low HDL (≤35 mg/dL) (Brown BG, 2001). Flushing (vasodilation of blood vessels that can cause a burning, tingling, and itching sensation) is a commonly observed adverse effect with both single-agent nicotinic acid and concomitant simvastatin and nicotinic acid. For example, in ARBITER-2, the placebo-adjusted rate of ßushing in patients was 56.5%, and the prescribing information for Niaspan reports that up to 88% of patients suffered from ßushing episodes in clinical studies. Combining Niaspan and simvastatin will protect the patent for Niaspan, Kos’s leading agent; the patent is being contested by Barr Laboratories. Following lawsuits in March 2002, a 30-month stay was issued that prevents Þnal FDA approval of Barr’s generic equivalent of Niaspan until patent expiration of branded Niaspan (2007) or resolution of the lawsuit. Cholesteryl Ester Transfer Protein Inhibitors Overview. One approach to raising HDL levels in the plasma is inhibition of CETP. DeÞciency of CETP is associated with raised HDL and lowered LDL,

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a proÞle that is considered antiatherogenic; heterozygous CETP deÞciency in humans is associated with HDL levels greater than 60 mg/dL and reduced risk of CHD (Barter PJ, 2003). Further data from the European Prospective Investigation into Cancer and Nutrition in Norfolk (EPIC-Norfolk) study indicate that elevated CETP concentration in conjunction with elevated TG levels is associated with increased odds for the occurrence of cardiovascular events (Boekholdt SM, 2004). Researchers reviewing these data concluded that the data highlight the complexity of HDL metabolism and the need for a wide variety of studies (Surtees PG, 2005). CETP inhibitors in late-stage development as monotherapies include JTT-705 (Roche and Japan Tobacco) and CETi-1 (Avant Immunotherapeutics). CETi-1 completed Phase II trials, but its development has been set back because of formulation issues. Mechanism of Action. CETP is a plasma protein that mediates the exchange of cholesteryl esters from antiatherogenic HDL to proatherogenic apolipoprotein B (Apo-B)-containing lipoproteins, such as VLDL, VLDL remnants, intermediatedensity lipoprotein (IDL), and LDL. Theoretically, inhibiting CETP has a positive effect on lipid ratios, increasing HDL and reducing LDL levels. JTT-705. Japan Tobacco licensed exclusive development and commercialization rights to JTT-705 (Figure 19), outside of Japan, to Roche Holding. JTT-705 is in European and U.S. Phase II clinical trials with Roche and in Phase I trials in Japan with Japan Tobacco. JTT-705’s mechanism of action is exactly as described for the CETP inhibitor drug class in general: it inhibits the transfer of cholesteryl esters from HDL to proatherogenic lipid subfractions. Bioavailability studies suggest that the agent is a more effective inhibitor of CETP postprandially. Phase I investigations conducted so far suggest that JTT-705 is well tolerated. Data from two randomized, dose-response Phase II studies in 198 patients with mild dyslipidemia indicate that treatment with 900 mg/day of JTT-705 for four weeks reduced CETP activity by 37%, raised HDL levels by 34%, and reduced LDL by 7% without affecting TGs (de Grooth GJ, 2002). Subsequent data conÞrm the Phase II Þndings with respect to HDL and LDL but indicate an 11.1% reduction in TGs (Schaefer E, 2003). Minor gastrointestinal side effects such as nausea, diarrhea, ßatulence, and constipation were reported in clinical studies.

FIGURE 19. Structure of GW-501516.

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Other Phase II studies show the beneÞts of JTT-705 in combination with pravastatin. In a double-blind, placebo-controlled trial, JTT-705 (300 mg or 500 mg once daily) was administered to 155 patients with type II dyslipidemia who were already using pravastatin (40 mg). After four weeks of treatment with 600 mg of JTT-705, HDL increased by 28% and LDL declined by 5% (Kuivenhoven JA, 2005). Researchers report that JTT-705 was safe and well tolerated. CETi-1. CETi-1∗ is an anti-CETP vaccine in Phase II trials with Avant Immunotherapeutics in the United States for the treatment of atherosclerosis and hypercholesterolemia. Currently, further development has been delayed following company reports in October 2004 that the company is continuing to evaluate several new adjuvants and delivery technologies for its original formulation of its clinical candidate vaccine. Avant Immunotherapeutics hopes that the vaccine can be administered as a single immunization with once- or twice-yearly booster doses to sustain adequate antibody levels against the CETP enzyme. CETi-1 is a unique drug that has the potential to allow long periods between dosing and to offer signiÞcant advantages over existing lipid-lowering drugs, especially with respect to compliance. The vaccine may also offer a favorable gastrointestinal side-effect proÞle, compared with that of other antihyperlipidemics, including other CETP inhibitors. Inhibition of CETP by an antibody prevents the transfer of cholesteryl esters from HDL to proatherogenic lipid subfractions, thereby increasing HDL and reducing LDL. Avant reports that in preclinical studies in rabbits, CETi-1 reduced atherosclerotic lesions in blood vessels by 40%, compared with controls. Antibody response to vaccination was titrated during a Phase I, double-blind, placebo-controlled study in 48 healthy adult volunteers; dose response was demonstrated (with maximal response to 250 µg injections), and no signiÞcant adverse events were reported. Antibody response was observed as early as one week following repeat vaccination and was sustained for up to ten weeks in some volunteers treated with the highest dose. The company reports a Phase II, placebo-controlled, dose-escalating clinical trial that is still under way in the United States; 200 patients with low HDL were administered four vaccinations over six months. The trial is designed to evaluate the safety, immunogenicity, and dose/response relationship of CETi1. Initial injections were followed at four and eight weeks, and a Þnal booster was administered after six months; the primary trial end point is the change in HDL measured after the Þnal six-month booster. More data from this study were presented at the Emerging Cardiovascular Therapeutics meeting in Cambridge, Massachusetts, in June 2003; no treatment-related adverse events were apparent at that point (Ryan U, 2003). Avant reported preliminary data from the study in October 2003. The agent was shown to increase HDL by 8.4% in patients who were not receiving statin therapy. However, the effects observed were not signiÞcantly different from effects seen in the placebo group. No effect on HDL was seen in patients who were receiving a statin.

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Reverse Lipid Transport Pathway Activators Overview. Following the acquisition of Esperion Therapeutics in 2004, PÞzer is developing a series of compounds that target the reverse lipid transport pathway for the treatment of atherosclerosis in both the acute and chronic settings. Agents have reached Phase II clinical trials for atherosclerosis and acute coronary syndrome (ACS), and the results to date have been promising. These agents include ETC-216, a recombinant form of apolipoprotein A-1 Milano (ApoA-IM), and ETC-588, a large unilamellar vesicle (LUV) formulation of a phospholipid liposome. In addition, ETC-642, a peptide/phospholipid complex in Phase I development that mimics the structure and function of HDL, is under investigation for reducing atherosclerotic events in patients with ACS and patients with stable atherosclerosis. ETC-216, ETC-588, and ETC-642 all require intravenous administration in a hospital setting and will generally be restricted to acute or short-term treatment of patients with established CHD. (Because these agents are not prescribed for chronic lipid modulation, they are not discussed further here.) However, PÞzer is undertaking preclinical investigations of orally active small molecules that increase HDL and have the potential to be used for chronic treatment of atherosclerosis and lipid abnormalities. Other companies with oral Apo-A1 mimetics include Novartis, whose D-4F—licensed from Bruin Pharma—is in Phase I development for hyperlipidemia, and Avanir Pharmaceuticals, which is investigating AVP-26452, the lead compound from a series of orally active ApoA-1 hypolipidemic agents, as a potential treatment for atherosclerosis and dyslipidemia. Mechanism of Action. All of PÞzer’s candidate molecules stimulate the RLTP, a four-step process responsible for removing excess cholesterol and other lipids from the walls of arteries and other tissues and transporting them to the liver for elimination from the body. The Þrst step is the removal of cholesterol from the walls of arteries by HDL. In the second step, cholesterol is converted to a new form that is more tightly associated with HDL as it is carried in the blood. The third step is the transport and delivery of that converted cholesterol to the liver. The Þnal step is the transformation and discarding of cholesterol by the liver. ETC-1001. ETC-1001∗ is an oral RLTP inhibitor in development by PÞzer following its acquisition of Esperion Therapeutics in 2004. ETC-1001 entered Phase I trials in 2003 in the United States for atherosclerosis and lipid abnormalities. Other compounds in the series, ESP-15228∗ and ESP-24232∗ , remain in discovery for the treatment of atherosclerosis. All these agents mimic the function of the ApoA-1 protein, the primary component of HDL, by interacting with phospholipids, thus mimicking the actions of HDL particles. While ETC-1001 was in development by Esperion Therapeutics, a company press release reported results from a double-blind, placebo-controlled, Phase I study in 36 healthy volunteers (Esperion Therapeutics, press release [electronic],

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December 4, 2003). The researchers found ETC-1001 to be safe and well tolerated in single doses over a broad range of dose levels. Esperion also reported that ESP-15228 reduced non-HDL cholesterol and serum TG levels by 90% in the female obese Zucker rat (fed on a cholesterolrich diet); also, HDL levels doubled after one week of therapy. Another study in the same model demonstrated a 50% reduction in non-HDL cholesterol at low doses when ESP-15228 was administered in combination with atorvastatin. Preclinical studies of ESP-24232 in both Sprague-Dawley rats and obese Zucker rats demonstrate a reduction in TGs and an increase in HDL; positive effects on the arterial wall were also observed and found to be independent of the agent’s lipid-lowering actions (Brousseau ME, 2002).

Acyl-CoA Cholesterol Acyltransferase Inhibitors Overview. Researchers have investigated Acyl-CoA cholesterol acyltransferase (ACAT) inhibitors as potential lipid-lowering therapies for some time, but in recent years, the number of ACAT inhibitors in development has declined rapidly because of problems associated with drug-induced adrenal toxicity, together with the failure of early clinical trials to meet the expectations raised by studies in animal models. Two ACAT enzymes are responsible for cholesterol ester formation in tissues, ACAT1 and ACAT2. Each enzyme has different cell locations, membrane orientations, and metabolic functions. Gene disruption experiments suggest that ACAT1 is responsible for cholesterol homeostasis in the brain, skin, adrenal glands, and macrophages (ACAT action in macrophages is important in atherosclerosis development) (Rudel L, 2001). By contrast, ACAT2 mice have limited cholesterol absorption in the intestine and decreased cholesterol ester content in the liver and plasma lipoproteins. ACAT inhibitors in late-stage development for dyslipidemia include Pierre Fabre/Eli Lilly’s eßucimibe. Sumitomo Pharmaceuticals has SMP-797 in Phase I development for this indication. Much of the research on ACAT inhibitors is now directed at the development of these agents as antiatherosclerotics—in Japan, for example, Kyoto Pharmaceutical Industries and Nisshin Seifun Group have discovery programs under way. Mechanism of Action. ACAT catalyzes cholesterol esteriÞcation, regulates intracellular free cholesterol levels, and promotes both cholesterol absorption in the small intestine and assembly of very-low-density lipoprotein (VLDL) in the liver. Ultimately, ACAT activity increases total serum cholesterol (TSC) and LDL levels in the bloodstream. ACAT inhibitors demonstrate cholesterollowering and antiatherosclerotic activities by blocking intestinal absorption of dietary cholesterol, inhibiting hepatic secretion of VLDL, and preventing accumulation of cholesterol esters in the cytoplasm of macrophages, thereby reducing the formation of foam cells (Alegret M, 2004).

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Eflucimibe. Pierre Fabre, in collaboration with Eli Lilly, is developing eßucimibe∗ . The drug entered Phase II trials in 2002 and has yet to progress to Phase III. Development may have been stalled by the low solubility of eßucimibe, but recent data show that when eßucimibe is complexed with gamma cyclodextrin, its solubility is increased 44-fold (Gil A, 2004). Peroxisome Proliferator Activated Receptor Agonists Overview. The PPAR family, including PPAR-α, PPAR-γ , and PPAR-δ, comprises ligand-activated transcription factors that regulate gene transcription in lipoprotein metabolism, immune response, and insulin resistance. The therapeutic potential of PPAR-α is currently realized in the hypolipidemic action of Þbrates and the potential of PPAR-γ in the antidiabetic effects of the thiazolidinediones (rosiglitazone [GlaxoSmithKline’s Avandia] and pioglitazone [Takeda Chemical Industries’ Actos]). Less is known about the potential of PPAR-δ as a target for modifying cardiovascular risk factors, but ongoing research of ligands that are selective for this receptor will help to clarify its therapeutic role. Some investigators propose that PPAR-δ controls the inßammatory status of macrophages, making this receptor a natural target for atherosclerosis (Lee CH, 2003). Current research into PPAR agonists is focused on agents that act as single agonists of one of the PPAR family of receptors and on agents that combine the effects of the PPAR family in dual-acting agonists PPAR-γ /δ and PPAR γ /α. The metabolic beneÞts induced by PPAR ligands would indirectly beneÞt cardiovascular outcomes by addressing both dyslipidemia and non-insulin-dependent diabetes. Because of their concomitant action on lipids and insulin sensitization, dual-acting agents would be useful as treatments for the metabolic syndrome. Additionally, because this family of receptors is also expressed in endothelial cells, vascular smooth-muscle cells, and macrophage cells, gene expression can be altered in the vascular wall directly, making agonists of these receptors useful in the treatment of atherosclerosis. The side-effect proÞles of PPAR agents have raised some concerns. Reports of hepatic abnormalities were followed by the withdrawal of Warner-Lambert’s antidiabetic thiazolidinedione, troglitazone (Rezulin), from the market in 2000. (Warner-Lambert is now part of PÞzer.) In 2004, the FDA introduced guidelines stating that two-year rodent toxicity studies must be completed for all PPAR compounds and reviewed by the FDA prior to the initiation of clinical trials of more than six months’ duration. This requirement will likely delay the launch of all PPAR agonists in development. Concerns about the role of PPAR in cancer development and progression are poorly deÞned. Research into the oncogenic effect of PPAR agonists indicates they have both oncogenic and anti-oncogenic effects. Research into mammary carcinogenesis found that PPAR-γ agonists exhibited a moderate delay in tumor formation, while PPAR-δ agonists accelerated tumor formation (Yin Y, 2005). In models of tumor progression in human colon cells, PPAR-α showed potential as a therapeutic target to prevent adenoma formation (Matthiessen MW, 2005).

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One PPAR agonist in late-stage development for dyslipidemia is a dual-acting PPAR γ /α, AstraZeneca’s tesaglitazar (Galida). GlaxoSmithKline has two PPAR agonists in Phase II development: the PPAR-α GW-590735 and the PPAR-δ GW501516, which is being developed in collaboration with Ligand Pharmaceuticals. Kyotorin Pharmaceutical is investigating a PPAR-α in Phase II development (KRP-101), but lack of information precludes further discussion of this agent at this time. Several agents are in early development; most activity is focused on PPAR-α agonists. SanoÞ-Aventis has two agents in Phase I development: AVE8134 and AVE-0847. Other Phase I agents include Eisai’s E-3030; Dr. Reddy’s Laboratories’ DRF-10945; and GlaxoSmithKline’s 641597. Merck, MaxoCore Pharmaceuticals, Nippon Shinyaku, Sigma-Tau Industries, Farmaceutical Riunite SpA, and GenÞt also have discovery programs investigating PPAR agonists for dyslipidemia. Mechanism of Action. The PPAR family, including PPAR-α, PPAR-γ , and PPAR-δ, comprises ligand-activated transcription factors that regulate gene transcription in lipoprotein metabolism, adipogenesis, and insulin resistance. The Þbrate and PPAR-α agonist fenoÞbrate (Fournier’s Lipantil, Abbot’s Tricor) (Figure 14) reduces atherosclerotic lesion size in animal models (Duez H, 2001), and PPAR-α agonism seems to reduce foam-cell formation (Argmann CA, 2001). Recently, researchers have demonstrated that PPAR agonists increase the expression of the ATP-binding cassette transporter-A1 (ABCA1 ) gene (Spijkers JA, 2001; Chinetti G, 2001), thereby increasing reverse cholesterol transport (lowering LDL and raising HDL levels). In animal models, PPAR agonists also reduce the expression of endothelial adhesion molecules, thereby encouraging the attachment of circulating monocytes (Zhang L, 2001). Other important roles of the PPAR family include involvement in the development of diabetic nephropathy in type 2 diabetes (Guan Y, 2004). Tesaglitazar. AstraZeneca is developing the dual PPAR-γ /α agonist tesaglitazar (Galida)∗ for the treatment of dyslipidemia and type 2 diabetes. Phase III clinical trials for both indications are under way in the United States. In October 2004, after the FDA call for two-year rodent studies, the company announced that it would delay Þling until 2007. The PPAR family comprises ligand-activated transcription factors that regulate gene transcription in lipoprotein metabolism, adipogenesis, and insulin resistance. The activation of PPAR-γ is associated with improved insulin sensitivity, while PPAR-α activation reduces serum TGs and increases serum HDL. Company reports of Phase II data state that tesaglitazar improves both dyslipidemia and glycemic control. Tesaglitazar is reported to lower blood-glucose levels and improve patients’ lipid proÞles in a dose-dependent manner after 12 weeks. One study involving approximately 500 patients found that with a one milligram dose, triglycerides were cut by 41% and LDL was cut by a statistically insigniÞcant 6%. On average, patients gained about 2.2 pounds, and adverse events like edema occurred at a rate of 1–4% in patients receiving tesaglitazar. Phase I clinical studies investigating the actions of tesaglitazar in healthy persons

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show tesaglitazar to be well tolerated, with 100% bioavailability, at daily doses of 1 mg (Ericsson H, 2004). In animal models, including the obese Zucker rat, tesaglitazar produced signiÞcant reductions in TGs and large increases in HDL, with glucose control similar to that of the marketed, insulin-sensitizing thiazolidinediones, another class of PPAR agonists. Other animal studies in Wistar rats show that tesaglitazar is an effective insulin-sensitizing agent in a mild dietary model of insulin resistance (Hegarty BD, 2004). Because tesaglitazar’s primary effects are seen in beneÞcial changes to insulin, TG, and HDL levels, this agent can be used in conjunction with the wellestablished statin class to treat patients who have metabolic syndrome. AstraZeneca is likely to promote this agent as an add-on therapy to its superstatin, rosuvastatin (Crestor). Ultimately, the commercial success of tesaglitazar will be determined by its efÞcacy compared with that of other PPAR-α/γ agonists in development; its lipid-lowering effects for use as a single therapy for modiÞcation of lipid levels; and the results of toxicity studies. GW-501516. Ligand Pharmaceuticals and GlaxoSmithKline are codeveloping the PPAR-δ agonist GW-501516 (GSK-516). This PPAR ligand is in Phase II development in the United Kingdom for the treatment of dyslipidemia. GW590735∗ —reported to be a derivative of GW-501516—is also in Phase II development for dyslipidemia, but it is not discussed further here because of a lack of data. GW-501516 is one of the Þrst PPAR-δ agonists in development, and unlike other PPAR agents, it modulates LDL in addition to TGs and HDL. Researchers report that GW-501516 increases the expression of the ABCA1 gene, thereby increasing reverse cholesterol transport and, consequently, lowering LDL levels and raising HDL levels (Oliver WR, 2001). Clinical data evaluating GW-501516 in dyslipidemic patients are unavailable. Ligand and GlaxoSmithKline report that in a primate model of metabolic syndrome, GW-501516 at a twice-daily dose of 3 mg/kg reduced LDL and TGs by 29% and 56%, respectively, and raised HDL by 79%. Fasting insulin levels also fell by 48% from baseline. In primate models of atherosclerosis, GW501516, placebo, and fenoÞbrate (Tricor) increased HDL by 43%, 5%, and 20%, respectively. Other animal data in obese ob/ob mice provide evidence for increases in glucose metabolism—independent of insulin—following administration of GW-501516, demonstrating its potential for use as an antidiabetic therapy (Kramer DK, 2005). Microsomal Triglyceride Transfer Protein Inhibitors Overview. Some experts had considered microsomal triglyceride transfer protein (MTTP) inhibitors one of the more promising areas of lipid-lowering research, but doubts about their potential toxicity have been raised. Some experts express concern that MTTP inhibitors are likely to have a relatively narrow therapeutic window and that they may cause signiÞcant adverse effects, including fat malabsorption with subsequent loss of fat-soluble vitamins and fatty liver deposits. If

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such toxicity issues can be overcome, these agents are most likely to be used as adjunct therapies to existing Þrst-line treatments in patients unable to reach lipid goals; patients with severe dyslipidemia will be candidates for such therapy. R&D within this drug class is rather limited. Bayer’s implitapide, in Phase II, is the most advanced MTTP inhibitor agent. Japan Tobacco’s JTT-130 is in Phase I clinical trials. Tanabe Seikayu and Janssen International (part of Johnson & Johnson) have preclinical programs. PÞzer was developing CP-346086 for the treatment of dyslipidemia, but it recently terminated the program. Data presented in March 2003 at the Fourth International Cholesterol-Lowering Conference in Cambridge, Massachusetts, indicate that in humans, CP-346086 lowers LDL, does not consistently lower TGs, and does not signiÞcantly affect HDL (Chang G, 2003). Despite these disappointing data, PÞzer continues to report on a preclinical development program of MTTP inhibitors. Mechanism of Action. MTTP localized in the endoplasmic reticulum of hepatocytes and enterocytes catalyzes the transport of TGs, cholesteryl ester, and phosphatidylcholine between membranes. MTTP is involved in the production of several lipoproteins and is required for the synthesis and cellular secretion of ApoB. Inhibitors of MTTP block the hepatic secretion of VLDL and intestinal secretion of chylomicrons, ultimately reducing the levels of circulating LDL (Jamil H, 1996; Chang G, 2002). Implitapide. In 2003, worldwide rights to Bayer’s implitapide (BAY-139952)∗ for atherosclerosis and lipid-related diseases were granted to MRL International (PPD Development). Bayer planned to develop this product outside of these indications. PPD was conducting proof-of-principle studies of implitapide as an adjunct to statin therapy. Plans included enrolling 200 patients with familial hypercholesterolemia or hypertriglyceridemia for three Phase II studies, including dose titration studies. However, in the Þrst quarter of 2005, for unstated reasons, MRL International elected to terminate the implitapide program and the collaboration with Bayer. Implitapide’s mechanism of action is as described for the MTTP class in general: inhibitors of MTTP block the hepatic secretion of VLDL and intestinal secretion of chylomicrons, ultimately reducing the levels of circulating LDL. In Phase I studies in patients with LDL above 130 mg/dL and TGs above 200 mg/dL, 80 mg implitapide signiÞcantly reduced postprandial lipid responses following high-fat meals. Data from 61 dyslipidemic patients treated with 20, 40, 80, or 160 mg/day of implitapide, presented at the Drugs Affecting Lipid Metabolism (DALM) meeting in September 2001 in New York, demonstrated a 12% reduction in TSC at the lowest dose and a 54% reduction at 160 mg/day. LDL cholesterol was reduced by 8% and 58% with the 20 mg/day and 160 mg/day doses, respectively. Apo-B fell 2–55% across the dose range. Importantly, nausea and diarrhea were noted at the 80 and 160 mg/day doses, and serum depletion of vitamin E was observed in preclinical studies.

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Antiatherosclerotic mechanisms of implitapide were shown in preclinical models. Following an eight-week treatment period of 3.2 mg/kg/day, implitapide reduced atherosclerotic lesion area by 83% in addition to reducing TGs and total cholesterol levels (Ueshima K, 2005). Squalene Synthase Inhibitors Overview. Inhibition of squalene synthase as a potential treatment approach for dyslipidemia is an idea that has circulated for some time. In the early 1990s, many large pharmaceutical companies, including Bristol-Myers Squibb and GlaxoSmithKline, had R&D programs focused on this mechanism of action. However, most of these research programs have been discontinued. Companies still pursuing this area of research include Takeda, which has TAK-475 in Phase II clinical trials, and Eisai, whose lead compound, ER-119884, is undergoing preclinical evaluations. Mechanism of Action. Squalene is a metabolite of mevalonate, which is involved in an advanced stage of cholesterol synthesis. Squalene synthase inhibitors interfere with cholesterol synthesis by halting the action of liver enzymes. In addition, because squalene synthase may be involved in cell growth, these agents may also slow or stop the proliferation of several cell types that contribute to atherosclerotic plaque formation. TAK-475. Takeda is developing TAK-475 for the treatment of dyslipidemia. European and U.S. Phase III trials are ongoing for this indication. This agent’s mechanism of action is as described for squalene synthase inhibitors in general. Squalene is a metabolite of mevalonate, which is involved in an advanced stage of cholesterol synthesis. Squalene synthase inhibitors interfere with cholesterol synthesis by halting the action of liver enzymes. Clinical data evaluating the actions of TAK-475 are unavailable, but the company reports that several long-term studies, involving hundreds of patients, are ongoing. Phase II proof-of-concept studies were conducted in patients with higher than normal LDL cholesterol; decreases in LDL and TG were observed. Effects on HDL are established in preclinical models but have yet to be proved in humans. Studies in several animal models, carried out by Takeda, suggest that TAK-475’s hypolipidemic actions occur via inhibition of hepatic TG secretion and an upregulation of LDL receptors, thus increasing HDL by lowering TGs (Nishimoto T, 2003[a]). In a rabbit model of homozygous familial hypercholesterolemia (HoFH), four weeks of treatment with 100 mg/kg per day TAK-475 lowered TSC by 17% and TG and phospholipids levels by 52% and 26%, respectively (Amano Y, 2003). Squalene synthase inhibitors act on the same cholesterol synthesis pathway as statins. Proponents of TAK-475 have suggested that inhibition downstream of mevalonate and its derivatives, dolichol and ubiquinone, may confer a greater safety advantage than inhibition of HMG-CoA reductase, the object of statins’ activity. These claims have yet to be established, but Takeda has conducted in vitro studies of TAK-475 combined with statins in human rhabdomyosarcoma

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models, and these studies show that TAK-475 in combination with atorvastatin or simvastatin may in fact lower statin-induced myotoxicity (Nishimoto T, 2003[b]).

Lipoprotein Lipase Activators Overview. Direct activation of lipoprotein lipase (LPL) is a novel approach to treating dyslipidemia. Otsuka/TAP Pharmaceuticals’ ibrolipim (formerly NO1886) is the member of this drug class that is most advanced in development. Mechanism of Action. LPL is an insulin-sensitive enzyme that promotes the breakdown of the fat portion of lipoproteins, speeding natural cholesterol degradation in the body. Activation of this enzyme increases serum HDL and lowers TGs. Ibrolipim. Otsuka and TAP Pharmaceuticals are codeveloping ibrolipim (NO1886)∗ , an LPL activator. Phase II clinical trials for dyslipidemia are ongoing in the United States. The agent’s mechanism of action is as described for the LPL activator class in general. Activation of LPL increases serum HDL and lowers TGs. Clinical trial data evaluating the actions of ibrolipim in dyslipidemic patients are unavailable; however, several studies have investigated its effects in animal models. Results show that increased LPL activity in response to NO-1886 administration signiÞcantly increased serum HDL and lowered serum TGs (Tsutsumi K, 1993). These effects are also seen in the diabetic state: single and repeated administrations of NO-1886 to streptozotocin-induced diabetic rats increased LPL activity, resulting in a reduction in plasma TGs and an elevation in plasma HDL levels. No effects on plasma glucose or insulin were observed in this study (Tsutsumi K, 1993). A recent study investigating the actions of NO-1886 in minipigs fed on a high-fat/high-sucrose diet found that supplementing food (1% NO-1886) inhibited adipocyte enlargement, had anti-inßammatory actions reducing TNF-α, reduced free fatty acids, and improved glucose metabolism by reducing insulin resistance (Yin W, 2004). Positive effects on vascular endothelial function have also been reported in rat studies (Kusunoki M, 2002). Studies show positive effects of ibrolipim in rat obesity models; although the mechanism is not well understood, researchers postulate that ibrolipim accelerates the expression of fatty-acid-oxidation-related enzymes (Doi M, 2003). Evidence from preclinical studies suggests that ibrolipim will be used predominantly in patients with combined dyslipidemia who have raised TGs, low HDL, and moderately elevated LDL. The positive effects on insulin resistance and obesity observed in some animal studies also point to potential use of this agent in patients with metabolic syndrome as outlined in the National Cholesterol Education Panel Adult Treatment Panel (NCEP ATP III) guidelines.

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Lipoprotein (a) Antagonists Overview. Elevated lipoprotein (a) (Lp[a]) is an independent risk factor for the development of various cardiovascular disorders. At this writing, PÞzer’s gemcabene is the only Lp(a) antagonist in development for the treatment of dyslipidemia. Mechanism of Action. The mechanism of action of this drug class has yet to be completely established. A reduction in Lp(a) and other key cholesterol levels (TSC, LDL, and TG) has been observed in animal models. In human studies, elevations in HDL have also been observed. Gemcabene. PÞzer’s gemcabene (formerly PD-79253)∗ , an Lp(a) antagonist, is in Phase II clinical trials in the United States. The agent’s mechanism of action has yet to be established. In preclinical studies, gemcabene demonstrated dose-dependent reductions in Lp(a) of 9–87% at doses of 3–300 mg/kg per day; TSC levels were also reduced (Auberbach BJ, 1999). The Þrst published study to investigate gemcabene’s actions in humans focused on its actions on serum HDL. One hundred and sixtyone dyslipidemic patients with HDL ≤35 mg/dL and serum TGs ≥200 mg/dL (n = 94) or <200 mg/dL were randomized to receive gemcabene (150, 300, 600, or 900 mg/day) or placebo for 12 weeks. In patients with TGs ≥200 mg/dL, gemcabene increased HDL by 18% at the 150 mg dose. A 12% increase was also seen at the 300 mg dose, but this result was not statistically signiÞcant. No increases in HDL were seen in patients with TGs ≥200 mg/dL who were receiving the 600 or 900 mg doses or in patients with TGs <200 mg/dL. At these higher doses, reductions in LDL of 15–25% were observed in both TG groups. Despite variations in effects on individual lipoproteins, a consistently favorable effect on atherogenic serum non-HDL cholesterol was seen across the dose range for this agent. Gemcabene was well tolerated in this study; its adverse-event proÞle was similar to that of placebo (Bays HE, 2003).

Bile Acid Reabsorption Inhibitors Overview. The discovery of the sodium (Na+ )/bile acid cotransporter has given researchers the opportunity to pursue development of speciÞc bile acid reabsorption inhibitors (BARIs; often referred to as Na+ /bile acid cotransporter [IBAT] inhibitors), potentially leading to the introduction of a novel class of antihyperlipidemic agents. Previously, both Aventis and Shionogi had Phase II BARI compounds in development. Development of Aventis’s BARI-1453 was stopped in 2004, and in 2005, Shinogi reported discontinuation of the development program for S-8921. SanoÞAventis’s BARI-1741 and AstraZeneca’s AZD-7806 are in Phase I development. PÞzer is investigating a series of candidate molecules in preclinical development.

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Mechanism of Action. BARIs inhibit the intestinal epithelial uptake of bile acids. These agents block the reabsorption of bile acid that is cycled from the liver to the gut and increase bile acid elimination, resulting in depletion of the cholesterol supply to hepatocytes and upregulation of LDL receptors. S-8921. Shionogi was developing the BARI S-8921 for treatment of dyslipidemia in European and Japanese Phase II clinical trials. However, in 2004, the company reported that Japanese trials were suspended, and in 2005, development in Europe was also stopped because data from Phase II studies failed to show any greater beneÞt of S-8921 compared with other compounds in this group. This agent’s mechanism of action is as described for the bile acid reabsorption inhibitor class in general. It inhibits IBAT, interrupting the ßow of bile acids cycling between the liver and the small intestine. In vivo data from several animal models were impressive; in normolipidemic hamsters, seven days of treatment at 77 mg/kg per day reduced LDL and VLDL by 49% and 60%, respectively, and raised HDL by 169% (Hara S, 1997). Data from a study using the Watanabe heritable hyperlipidemic rabbit reveal that dietary supplementation with S-8921 (0.01–0.1%) reduces serum cholesterol by 29–37% and increases bile acid excretion by 60–180%. Further investigations in cholesterol-fed New Zealand white rabbits reveal that dietary supplementation with S-8921 (0.003%−0.1%) dose-dependently inhibited the development of hypercholesterolemia, prevented the accumulation of cholesterol in the aortic arch, and reduced the severity of coronary atherosclerosis (Higaki J, 1998).

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Metabolic Syndrome

ETIOLOGY AND PATHOPHYSIOLOGY Definition Clinicians recognize metabolic syndrome as a cluster of comorbidities that are risk factors for cardiovascular disease. This cluster includes abdominal obesity, insulin resistance, hypertension, and dyslipidemia. Proinßammatory and prothombic states are also thought to contribute to the disorder and the risk of mortality from coronary heart disease (CHD) and other cardiovascular complications. However, no clear deÞnition of metabolic syndrome has been established, and the etiology and pathophysiology of the syndrome are still poorly understood. Furthermore, many basic questions regarding the development of metabolic syndrome and the interactions between the various risk factors remain unanswered. The lack of consensus and confusion surrounding the deÞnition of metabolic syndrome will likely be a tremendous obstacle to the development of therapeutics for this emerging indication. Without generally accepted diagnostic guidelines and clear deÞnitions of therapeutic and clinical trial end points, it is unlikely that any novel agents speciÞcally labeled for the treatment of metabolic syndrome will ever be approved by drug regulatory agencies. In addition, until a clear consensus is reached by the medical community and regulatory bodies regarding treatment guidelines for patients with metabolic syndrome, treatments for the disorder will continue to center on the individual components of metabolic syndrome rather than the indication as a whole. The uncertainty regarding the deÞnition of metabolic syndrome is evident in the varied nomenclature describing the syndrome. Gerald Reaven of Stanford University coined the term syndrome X to describe a cluster of cardiovascular risk factors connected to insulin resistance and obesity (Reaven GM, 1988). The Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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syndrome has also been referred to by a number of different names: dysmetabolic syndrome, insulin resistance syndrome, Reaven syndrome, cardiovascular dysmetabolic syndrome, dyslipidemic hypertension, and ”the deadly quartet.” Most researchers now commonly refer to this cluster of cardiovascular risk factors as metabolic syndrome. Key features of metabolic syndrome include the following: • • • •

Abdominal obesity. Insulin resistance. Hypertension. Dyslipidemia.

Although not formally included in the diagnostic criteria for metabolic syndrome (described later), a proinßammatory state (subclinical inßammation marked by elevated C-reactive protein [CRP] levels) and a prothrombotic state (marked by raised Þbrinogen levels) are also believed to contribute to the pathogenesis of metabolic syndrome and cardiovascular disease (CVD). However, despite the increasing attention placed on inßammation and thrombosis, their roles in the etiology and pathophysiology of metabolic syndrome are unclear. Questions remain about whether these factors have fundamental roles in the development of metabolic syndrome, or whether they are consequences of the disorder. Furthermore, measurements of these proteins in blood are not routinely obtained in clinical settings, and their relevance in the diagnosis or treatment of metabolic syndrome remains uncertain. In addition to the many names assigned to metabolic syndrome, a host of diagnostic criteria have been published by various sources. The most widely used diagnostic criteria for metabolic syndrome are from the Third Report of the Adult Treatment Panel of the National Cholesterol Education Program (NCEPATP III) and the World Health Organization (WHO). Two other deÞnitions of metabolic syndrome have been released by the American Association of Clinical Endocrinologists (AACE) and the European Group for the Study of Insulin Resistance (EGIR); however, these latter two sets of criteria are not widely recognized by physicians. The NCEP-ATP III deÞnition is used most often by PCPs, cardiologists, and endocrinologists in the United States, Europe, and Japan—three of the Þve diagnostic criteria in this deÞnition are markers for cardiovascular health that are easily measured in a clinical setting. The NCEP-ATP III criteria will likely form the basis of any future consensus deÞnition. Under the NCEP-ATP III deÞnition, patients must meet three of the following Þve criteria for a diagnosis of metabolic syndrome: abdominal obesity, elevated triglyceride levels, low levels of high-density lipoprotein cholesterol (HDL-C), hypertension (elevated systolic or diastolic blood pressure), and insulin resistance (elevated fasting plasma glucose levels) (Table 1). Although both decreased HDL-C and elevated triglycerides are components of dyslipidemia, they are considered to be independent components of metabolic syndrome under these guidelines (Expert Panel on Detection, Evaluation, and Treatment of High Blood Cholesterol in Adults, 2001).

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TABLE 1. Definitions of Metabolic Syndrome According to the National Cholesterol Education Program (NCEP) Adult Treatment Panel III (ATP III), a patient with three or more of the following traits is defined as having metabolic syndrome (Expert Panel on Detection, Evaluation, and Treatment of High Blood Cholesterol in Adults, 2001): • Abdominal obesity: waist circumference >102 cm in men and >88 cm in women. • Hypertriglyceridemia ≥150 mg/dL (1.7 mmol/L). • Low high-density lipoprotein (HDL) cholesterol: <40 mg/dL (1.04 mmol/L) in men and <50 mg/dL (1.29 mmol/L) in women. • Hypertension, defined as elevated blood pressure (≥130 mm Hg systolic or ≥85 mm Hg diastolic). • Elevated fasting plasma glucosea ≥110 mg/dL (≥6.1 mmol/L). According to the World Health Organization (WHO), a patient with insulin resistance as defined by a diagnosis of type 2 diabetes, impaired fasting glucose (IFG), impaired glucose tolerance (IGT), or impaired glucose uptake (measured by the hyperinsulinemic euglycemic clamp technique) and two of the criteria described below has metabolic syndrome (Alberti KG, 1998): • Hypertension, defined as antihypertensive treatment and/or elevated blood pressure (>140 mm Hg systolic or >90 mm Hg diastolic). • Dyslipidemia, defined as elevated plasma triglyceride (≥150 mg/dL, ≥1.7 mmol/L) and/or low HDL cholesterol (<35 mg/dL, <0.9 mmol/L in men and <39 mg/dL, <1.0 mmol/L in women). • Obesity, defined as a high BMI (≥30 kg/m2 ) and/or a high waist:hip ratio (>0.90 in men and >0.85 in women). • Microalbuminuria, defined by urinary albumin excretion rate ≥20 µg/min or albumin:creatine ratio ≥30 mg/g. a The cutoff for diagnosis of hyperglycemia was lowered to 100 mg/dL in 2004 by the American Diabetes

Association. The National Heart, Lung, and Blood Institute and the American Heart Association have not yet revised the NCEP-ATP III guidelines to reflect this change but may do so in the near future (Grundy SM, 2004). BMI = Body mass index. Note: Full source citations appear in ‘‘References.’’

In contrast, the WHO deÞnition was written by a panel of endocrinologists and centers on insulin resistance; a diagnosis of metabolic syndrome using these guidelines requires an initial diagnosis of insulin resistance as deÞned by type 2 diabetes, impaired glucose tolerance (IGT), impaired fasting glucose (IFG), or, in patients with normal fasting glucose levels, “glucose uptake below the lowest quartile for background population under hyperinsulinemic, euglycemic conditions” (Grundy SM, 2004) (Table 1). The WHO criteria are not widely used by physicians because the diagnostic tests for insulin resistance and impaired glucose tolerance are cumbersome to administer. Insulin resistance can be measured only via the hyperinsulinemic euglycemic clamp technique, which requires specialized equipment and continuous monitoring of serum glucose levels. Oral glucose tolerance testing involves the drawing of multiple blood samples following oral administration of a bolus of glucose and requires a two-hour stay in a doctor’s

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ofÞce so that periodic samples can be taken. By comparison, measurement of FPG (the criterion used by the NCEP-ATP III deÞnition) requires only a single blood sample. The WHO criteria further differ from the NCEP-ATP III deÞnition by including albuminuria (a marker for advanced diabetes characteristic of reduced kidney function) as a diagnostic criterion and by utilizing different cutoff values for HDL-C and hypertension. The American Association of Clinical Endocrinologists and American College of Endocrinology (AACE/ACE) guidelines for diagnosis of metabolic syndrome are an amalgam of the NCEP-ATP III and WHO criteria (Table 2). Formal diagnosis is left to the discretion of the individual clinician based on an assessment of the following risk factors: family history of type 2 diabetes, hypertension, or cardiovascular disease; polycystic ovary disease; sedentary lifestyle; age; ethnicity at risk for type 2 diabetes; or cardiovascular disease (Einhorn D, 2003). Of note, the focus on insulin resistance in the AACE/ACE guidelines excludes patients already diagnosed with type 2 diabetes (fasting plasma glucose greater than 126 mg/dL) from diagnosis of metabolic syndrome (Grundy SM, 2004; Einhorn D, 2003). The EGIR deÞnition of metabolic syndrome is similar to the WHO deÞnition (Balkau B, 2002) (Table 3). A key difference between these two guidelines is the consideration of hyperinsulinemia as the underlying diagnosis in the EGIR deÞnition—its inclusion offers a distinct advantage, given that insulin resistance is more difÞcult to measure. Like the AACE guidelines, the EGIR deÞnition excludes patients already diagnosed with type 2 diabetes from diagnosis of metabolic syndrome. A patient is diagnosed with hyperinsulinemia if his or her fasting plasma insulin concentration exceeds the top quartile of values compared with nondiabetic individuals. For a diagnosis of metabolic syndrome under the EGIR deÞnitions, patients must also meet at least two of four additional criteria: hyperglycemia, hypertension, dyslipidemia, or abdominal obesity. These additional criteria were chosen based on the ease by which they can be measured in a clinical setting. They directly address some of the limitations of the WHO deÞnition of metabolic syndrome, which has been criticized as being useful only as an academic tool. Etiology Lifestyle factors such as poor diet, lack of physical exercise, and smoking contribute to the development of obesity and insulin resistance. However, both of these conditions can also arise independently and cause the other metabolic defects that constitute the syndrome. Although obese individuals are predisposed to developing insulin resistance, dyslipidemia, and hypertension, not all obese individuals are insulin resistant. Nevertheless, these two disorders form the core fcof metabolic syndrome and are inextricably linked. This section addresses some of the theories connecting lifestyle factors to the etiology and pathophysiology of metabolic syndrome.

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Lifestyle Factors. Lifestyles have changed signiÞcantly over the past 50 years. In particular, increased adoption of Western diets and decreased physical activity due to the rising use of automobiles and other modern conveniences have led to a rise in the prevalence of obesity, diabetes, and metabolic syndrome throughout the world. Smoking. Smoking has been implicated as a signiÞcant factor contributing to insulin resistance, diabetes, and cardiovascular disease and is increasingly recognized as a factor in the pathogenesis of metabolic syndrome as well (Dzien A, 2004; Eliasson B, 2003). In addition to contributing to inßammation and oxidative stress by damaging endothelial tissue, smoking has been shown to have deleterious effects on lipoprotein and cholesterol levels. Data from the Prospective Cardiovascular M¨unster Heart Study (PROCAM), which enrolled more than 20,000 men and 10,000 women, demonstrated that smokers had, on average, 7% lower HDL-C and 15% higher triglyceride (TG) levels compared with nonsmokers (Cullen P, 1998). Low-density lipoprotein cholesterol (LDL-C) levels were only a modest 1.4% greater in smokers enrolled in the PROCAM study relative

TABLE 2. AACE/ACE Diagnostic Guidelines for Insulin Resistance Syndrome Under the American Association for Clinical Endocrinology (AACE) and American College of Endocrinology (ACE) guidelines, diagnosis of insulin resistance syndrome is based on the following risk factors: Overweight/obesity: • BMI ≥25 kg/m2 . Lipid abnormalities: • • • •

Elevated triglyceride levels ≥150 mg/dL (1.7 mmol/L). Low HDL cholesterol. Men <40 mg/dL (1.04 mmol/L). Women <50 mg/dL (1.29 mmol/L).

Elevated blood pressure: • ≥130 mm Hg systolic or ≥85 mm Hg diastolic. Insulin resistance: • Elevated fasting glucose between 110 mg/dL (6.1 mmol/L) and <126 mg/dL (7.0 mmol/L). or • Serum glucose between 140 mg/dL (7.8 mmol/L) and 200 mg/dL (11.1 mmol/L) 120 minutes after a 75 gram oral glucose challenge. BMI = Body mass index. HDL = High-density lipoprotein. Note: Full source citations appear in ‘‘References.’’ Source: Based on Einhorn D, 2003, and Grundy SM, 2004.

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TABLE 3. European Group for the Study of Insulin Resistance (EGIR) Definitions of Metabolic Syndrome Patients must exhibit hyperinsulinemia (defined as fasting insulin concentration above the upper quartile for nondiabetic subjects) and at least two additional criteria to be diagnosed as having metabolic syndrome: Hyperglycemia • Fasting plasma glucose (FPG) >110 mg/dL (6.1 mmol/L). Elevated blood pressure • ≥140 mm Hg systolic. or • ≥90 mm Hg diastolic. or • Under treatment for hypertension. Lipid Abnormalities • Triglycerides >180 mg/dL, and/or (2.02 mmol/L). or • HDL cholesterol <40 mg/dL (1.04 mmol/L). or • Under treatment for dyslipidemia. Central obesity • Waist circumference • Men >94 cm. • Women >80 cm. HDL = High-density lipoprotein. Note: Full source citations appear in ‘‘References.’’ Source: Based on Balkau B, 2002.

to nonsmokers (Cullen P, 1998). Similar results were obtained in a meta-analysis of 54 published studies on the effects of smoking: in this analysis, smokers were shown to have 5.7% lower HDL-C, 9.1% higher triglycerides, and 1.7% higher LDL-C compared with nonsmokers (Craig WY, 1989). The Data from the Epidemiological Study on the Insulin Resistance Syndrome (DESIR) study enrolled more than 5,000 men and women; 22.5% of male smokers in this study were diagnosed with metabolic syndrome at baseline (based on the WHO guidelines) compared with only 15.3% of nonsmokers. Interestingly, there was no signiÞcant difference in this study in the prevalence of metabolic syndrome between female smokers (6.3%) and female nonsmokers (6.0%) (Geslain-Biquez C, 2003). Another study, enrolling 3,804 nondiabetic men, revealed that smokers had lower HDL-C levels and higher fasting glucose and triglyceride levels but no differences in other metabolic parameters compared with nonsmokers (Dzien A,

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2004). However, male smokers with preexisting CVD had signiÞcantly poorer metabolic proÞles than their nonsmoking counterparts; this Þnding supports the notion that smokers often make other poor lifestyle choices (such as poor diet and inactivity) that further contribute to the development of metabolic syndrome and increased cardiovascular risk. Smoking is also strongly associated with elevations in both CRP and Þbrinogen, which are markers of systemic inßammation and increased thrombotic activity, respectively (Fr¨ohlich M, 2003). These two factors are important in the pathogenesis of metabolic syndrome and cardiovascular disease given their prominent roles in atherogenesis, but they are not speciÞcally included in any of the metabolic syndrome deÞnitions. Poor Diet. Public health ofÞcials worldwide implicate Western diets and eating patterns in the rising epidemic of obesity, insulin resistance, and metabolic syndrome. Western diets–which are high in calories, fat, and reÞned sugar–have been shown to give rise to insulin resistance even before obesity develops. The availability of fat-laden and calorie-rich foods in the United States and Europe has resulted in substantial increases in the prevalence of obesity and hyperlipidemia in these countries. The inßuence of Western dietary patterns has been further borne out by studies of immigrant populations, which by the second generation exhibit most of the health problems observed in their new countries (Fujimoto WY, 2000). An interventional study of 180 Italian patients with metabolic syndrome demonstrated that individuals instructed to maintain a traditional Mediterranean diet (which includes high amounts of fruits, whole grains, and olive oil) exhibited reduced body weight, improved insulin sensitivity, less systemic inßammation, and improvements to clinical signs of metabolic syndrome during two years of follow-up compared with those who received no dietary intervention. Only 40 patients still met the criteria for metabolic syndrome (based on the NCEP-ATP III deÞnition) after two years, compared with the 78 patients in the control group who did not modify their diets (Esposito K, 2004). Lack of Physical Activity. Lack of physical activity is a strong contributor to metabolic syndrome. In one study of 612 middle-aged Finnish men without metabolic syndrome at baseline (as deÞned by the WHO criteria), those who exercised regularly (greater than three hours per week) had 50–75% lower risk of developing metabolic syndrome during the four-year follow-up period relative to sedentary men (less than 10 min of exercise per week) (Laaksonen DE, 2002). Similar results were observed in a survey of 7,104 women—those in the highest quintile of cardiovascular Þtness who also exercised the most were 60–75% less likely to have metabolic syndrome than sedentary individuals in the least Þt quintile (Farrell SW, 2004). Physical activity has indirect beneÞcial effects on the individual components of metabolic syndrome, predominantly by facilitating weight loss. In one metaanalysis of 90 clinical studies, researchers demonstrated that a 1 mg/dL increase in HDL-C accompanied every 7 pounds of body weight lost through diet and

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exercise (Dattilo AM, 1992). In another study, 131 overweight sedentary males were randomized to receive instructions on diet, exercise, or no intervention for one year. Patients who dieted lost an average of 7.8 kg of bodyweight compared with controls (no intervention), while those who exercised lost an average of 4.6 kg. Both treatment groups exhibited improvements to HDL-C and triglyceride levels compared with the control population, indicating that both diet and exercise were similarly effective, regardless of the amount of weight lost (Wood PD, 1988). The role of inactivity alone in the etiology of metabolic syndrome is unclear—lack of physical activity is strongly associated with poor physical outcomes such as obesity and insulin resistance. However, other unhealthy lifestyle attributes, such as poor diet and smoking, contribute to poor physical conditioning that leads to inactivity, making it difÞcult to distinguish the impact made by each of these components alone. Nevertheless, physicians treat these three lifestyle attributes as modiÞable risk factors. Genetics. Scientists believe that some individuals are genetically predisposed to develop metabolic syndrome. Some South Asian and Native American populations are particularly susceptible to developing insulin resistance and metabolic syndrome (Tan CE, 2004; North KE, 2003). A genetic link is further supported by analysis of data from the third National Health and Nutrition Examination Survey (NHANES III), which indicates that the prevalence of metabolic syndrome in the United States is highest in the Mexican-American population and lowest among African Americans (Park YW, 2003). Few candidate genes for metabolic syndrome have been identiÞed so far—the most recently found mutation involves a maternally transferred gene involved in mitochondrial protein synthesis that is linked to hypercholesterolemia, hypertension, and hypomagnesemia (low serum magnesium levels) (Wilson FH, 2004). Pathophysiology Obesity. The connection between lifestyle factors and obesity is clear—except in rare individuals with genetic defects that predispose them to accumulate fat mass, obesity arises from poor diet and inactivity. The link between obesity and metabolic syndrome is less clear. Nevertheless, data from NHANES III indicate that the prevalence of comorbidities such as dyslipidemia, hypertension, and diabetes increases in both men and women as a function of rising body mass index (BMI), demonstrating that BMI is strongly associated with the components of metabolic syndrome (Must A, 1999). Much of the focus on obesity and its connection to metabolic syndrome has centered on increased abdominal (visceral) fat mass, which is composed of adipocytes (fat cells) that are more metabolically active than those found in subcutaneous fat. Adipocytes release hormones and cytokines (adipocytokines) that contribute directly to insulin resistance and the development of other components of metabolic syndrome. Adipocytes are also a source of free fatty acids (FFAs), which are released as a result of aberrant insulin signaling and further contribute

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to dyslipidemia, insulin resistance, and the declining function of pancreatic beta cells (described in greater detail later). Adipocytes were once thought to function only as storage cells for excess FFAs but are now widely recognized as important sources of signaling molecules that regulate metabolic processes. These highly metabolically active cells synthesize and secrete molecules that communicate the status of energy stores to the brain and other tissues. Excess endocrine activity by fat cells, either from adipocyte dysfunction or overabundance of fat tissue, has been cited as a trigger for metabolic syndrome (Bays H, 2004; Kershaw EE, 2004). Adipocytes in visceral fat release high amounts of inßammatory cytokines such as TNF-alpha, interleukin-6, and interleukin-8; hormones such as leptin, resistin, estrogen, and angiotensin; polyunsaturated fatty acids; and cholesterol ester transfer protein (CETP) (Table 4). Prolonged exposure to these factors desensitizes liver and muscle cells to the effects of insulin, resulting in insulin resistance (described in greater detail later in this section). The combined effects of chronically elevated glucose, FFAs, and cytokines in the circulation are hypothesized to lead to dyslipidemia through their effects on the metabolism of lipoproteins in the liver and to hypertension because of associated stiffening of the vascular-endothelial walls. Subsequent activation of macrophages and other inßammatory processes, combined with the presence of atherogenic lipoprotein particles and thrombosis, leads to a high risk of poor cardiovascular outcomes (Figure 1). Insulin Resistance and Diabetes. Insulin resistance and type 2 diabetes are both insidious processes that take many years to develop. Insulin resistance often stems from obesity but can develop independently as a consequence of aberrant signaling from adipocytes or disruptions in intracellular signaling pathways. Normal skeletal muscle tissue is responsible for the uptake of approximately 75% of postprandial serum glucose. However, adipocytokines may directly desensitize the liver and skeletal muscle tissue to the effects of insulin. Insulin resistance

FIGURE 1. Etiology and pathophysiology of metabolic syndrome.

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TABLE 4. Key Proteins Secreted by Adipocytes with Roles in Metabolic Syndrome Protein Type Hormones

Cytokines

Fibrinolytic proteins

Lipid metabolism proteins

Molecule Leptin

Function in Metabolic Syndrome

Signals levels of fat stores to the hypothalamus; functions upstream of several regulators of appetite in the brain; communicates with leptin receptors on adipocytes and other tissues. Adiponectin Regulates fat and glucose metabolism, possibly by modifying insulin sensitivity and stimulating uptake of free fatty acids. Adiponectin receptors are found in skeletal muscle tissue and the liver. Resistin Found in high circulating levels in both obese and insulin-resistant individuals. The normal function of resistin is unknown. TNF-α Impairs insulin signaling by inhibiting the function of insulin receptor substrate 1 (IRS-1); strong inflammatory cytokine implicated in atherosclerosis. Interleukin-6 Associated with glucose (IL-6) intolerance; triggers inflammation; stimulates CRP secretion. Plasminogen Inhibits urokinase and activator tissue plasminogen inhibitor 1 activator to block (PAI-1) fibrinolysis (dissolving of blood clots). Cholesterol Transfers cholesterol ester transfer esters from HDL-C protein (CETP) particles to VLDL and LDL-C in exchange for triglycerides; HDL-C particles are lost in the process.

Comment Was under development by Amgen as an antiobesity agent; withdrawn because of lack of efficacy.

Obese and insulin-resistant individuals have low levels of adiponectin; lean individuals have high levels; structurally similar to TNF-α.

Leads to insulin resistance in liver cells but not muscle cells; promotes inflammation.

TNF-α receptors are also found on adipocytes.

IL-6 receptors are also found on adipocytes.

High circulating levels of PAI-1 seen in obese individuals may contribute to a prothrombotic state. Predominantly produced by the liver.

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TABLE 4. (continued) Protein Type Steroid metabolism proteins

Molecule

Function in Metabolic Syndrome

11β-hydroxyste- Converts cortisone to its roid active form, cortisol. dehydrogeCortisol, and other nase glucocorticoids, (11β-HSD) stimulate liver gluconeogenesis, inhibit glucose uptake by muscle cells and adipocytes, and mobilize free fatty acids from adipocytes.

Comment Patients with Cushing’s disease (arising from cortisol excess) have metabolic profiles similar to those of obese patients.

CRP = C-reactive protein; HDL-C = High-density lipoprotein cholesterol; LDL-C = Low-density lipoprotein cholesterol; TNF = Tumor necrosis factor; VLDL = Very-low-density lipoprotein. Note: Full source citations appear in ‘‘References.’’ Source: Adapted from Kershaw EE, 2004, and Bays H, 2004.

also occurs following reductions in fatty-acid metabolism and oxidation in muscle tissue—these changes occur when cells are chronically exposed to high circulating levels of free FFAs. Lipid-Þlled muscle cells become inefÞcient at taking up circulating glucose and become insulin resistant as a result of concomitant alterations in insulin-signaling pathways. Normal blood glucose levels in insulin-resistant individuals can be maintained for several years thanks to the production of excess insulin (hyperinsulinemia) by pancreatic beta cells (Figure 2). Adipocytes respond to hyperinsulinemia by releasing nonesteriÞed FFAs into the circulation, where they are taken up by muscle and liver cells. FFAs are elevated in both type 2 diabetic patients and obese individuals and can amplify glucose-stimulated insulin secretion from pancreatic beta cells to further contribute to hyperinsulinemia (Itoh Y, 2003). Additional studies report that short-term exposure to FFAs enhances insulin secretion under acute conditions and maintains the secretion of insulin by beta cells in response to glucose long after a prolonged fasting. Over time, the beta cells are no longer able to produce sufÞcient insulin to maintain proper glucose levels—the resultant failure of the β-cells subsequently causes hyperglycemia, glucose toxicity, and hypoinsulinemia. It is the glucose toxicity associated with advanced diabetes that is most dangerous in terms of cardiovascular risk. Thus, it is imperative to detect and treat metabolic syndrome and insulin resistance early to prevent the development of diabetes, which is considered to be a cardiovascular risk equivalent based on the Framingham global risk scoring system. Large-scale prospective studies consistently demonstrate that insulin resistance (as measured by fasting insulin levels) and diabetes contribute signiÞcantly to risk of heart attack and other poor cardiovascular outcomes (Table 5). Dyslipidemia. Researchers have found strong inverse associations between plasma HDL-C levels and the risk of CVD. This phenomenon has been documented for both men and women and for both asymptomatic persons and patients

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TABLE 5. Studies Demonstrating the Relationship of Insulin Resistance, Diabetes, and Cardiovascular Disease Study Name Helsinki Policemen Study ¨ al ¨ a¨ M, 1998) (Pyor

Study Design

Primary Outcomes

Prospective study of 970 men Subjects in the highest quintile of aged 34–64 free of circulating insulin levels had cardiovascular disease and the highest risk of heart attacks diabetes; subjects were throughout the 22-year followed over a 22-year follow-up period. period. The increased risk was highest during the first five years of follow-up and decreased over time. Over 90% of subjects in the lowest quintile of circulating insulin levels remained free of heart attack over the 22-year study period, compared with 75% of subjects in the highest quintile. ARIC-Atherosclerosis Risk Prospective study of 13,790 Over 95% of subjects without In Communities (Lee African American and diabetes or prior history of CD, 2004) Caucasian men and women heart attack remained free of aged 45–64; subjects were CVD events over the 11-year followed for up to 11 years. follow-up period, compared with 85% of diabetics without history of MI, 75% of nondiabetics with prior MI, and less than 65% of diabetics with prior MI. Patients with a history of heart attack and no diabetes had 1.8 times the risk CVD mortality compared with diabetics without prior history of CVD. MRFIT-Multiple Risk Survey of 347,978 men aged Type 2 diabetes is a stronger Factor Intervention Trial 34–57, with and without cardiovascular risk factor than (Stamler J, 1993) diabetes; participants were hypertension, total serum followed over a period of 12 cholesterol, or smoking. years. Type 2 diabetics with no other risk factors had significantly higher rates of CVD death than nondiabetics with one or two other risk factors. Type 2 diabetics with multiple risk factors had more than twice the rate of CVD death than their nondiabetic counterparts with the same number of risk factors.

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TABLE 5. (continued) Study Name

Study Design

Quebec Cardiovascular 2,103 men aged 45–76 Study (Depres JP, 1996) without ischemic heart disease; participants were followed over a period of five years.

Primary Outcomes High fasting insulin levels in nondiabetic men were associated with four- to sevenfold higher odds ratios for a primary ischemic event (angina pectoris, heart attack, or death from coronary heart disease). Elevated triglyceride levels were associated with increased cardiovascular risk only in patients with both high fasting insulin levels and elevated apolipoprotein B levels.

CVD = Cardiovascular disease; MI = Myocardial infarction. Note: Full source citations appear in ‘‘References.’’

with conÞrmed CVD (Table 6). Low HDL-C is considered an independent risk factor for CVD; NCEP-ATP III guidelines for the treatment of dyslipidemia put increased emphasis on treating low HDL-C levels (less than 40 mg/dL). However, the relationship between HDL-C and CVD is not entirely understood. Concentrations of HDL-C tend to be low when TG concentrations are high, and HDL-C may, to a large extent, be a reciprocal measure of atherogenic lipoproteins such as very-low-density lipoprotein (VLDL) cholesterol. HDL-C actively transports cholesterol out of foam cells in an atherosclerotic plaque and delivers it to the liver, where HDL-C particles are selectively taken up. Studies suggest that the antioxidant and anti-inßammatory properties of HDL-C may protect against atherosclerosis. Low plasma HDL-C concentrations may also identify people with risk factors for metabolic syndrome; HDL-C is relatively low in smokers, obese individuals, and sedentary persons. Research suggests that TGs may play a more important role in CVD than previously recognized. These fats are a component of atherosclerotic plaques and may contribute to atherosclerosis by promoting coagulation and interfering with Þbrinolysis. Serum TG levels can vary in a given person from 50 mg/dL to more than 500 mg/dL but are typically less than 100 mg/dL. The NCEP-ATP III guidelines consider levels below 150 mg/dL to be desirable. Meta-analyses and epidemiological studies have more clearly identiÞed elevated levels of TGs as a risk factor for CVD (Austin MA, 1998). ATP III considers elevated TGs as a marker for other lipid and nonlipid abnormalities and as a target for lipid-lowering therapy. Hypertension. Patients with complicated hypertension are distinguished from those with uncomplicated hypertension by the presence of comorbidities or additional risk factors such as diabetes or dyslipidemia. Alterations in the properties of the blood vessels exacerbate disturbances in blood ßow inside the large vessels and can lead to sustained dysfunction of the endothelium, which can

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TABLE 6. Clinical Studies Assessing Dyslipidemia and the Risk of Cardiovascular Disease Study Name

Study Design

Primary Outcomes

Framingham Heart Study (Kannel WB, 1983; Kannel WB, 1987; Castelli WP, 1986)

Prospective study of 5,209 men and women aged 30–62; subjects were followed for more than 50 years.

• Risk of CHD is directly related to HDL-C levels. • The relative risk of CVD doubles with each 20 mg/dL decrease in HDL-C levels. • Total cholesterol level is also an important risk factor—risk of CVD rises along with increased total serum cholesterol/HDL-C ratios. • Low HDL-C increases the risk of CHD even in subjects with total serum cholesterol levels <200 mg/dL. These subjects had 14-year incidence rates as high as those with HDL-C <40 mg/dL and total serum cholesterol >260 mg/dL.

PROCAM—Prospective Cardiovascular Munster Study (Assman G, 1992; Assman G, 1993)

Prospective study of 4,407 German men aged 40–65; subjects were followed for a period of six years.

• The incidence of CHD was 3.7–5.2 times higher in subjects with HDL-C levels below 35 mg/dL compared with those with HDL-C levels between 35–55 mg/dL or greater than 55 mg/dL. • Patients with hypertriglyceridemia (TG >200 mg/dL) had significantly higher risk of CVD if they also had high LDL-C/HDL-C ratios.

Framingham Heart Study, Lipid Research Clinics Prevalence Mortality Follow-up Study, Lipid Research Clinics Coronary Primary Prevention Trial, and MRFIT—Multiple Risk Factor Intervention Trial (Gordon DJ, 1989)

Meta-analysis of four published studies.

• Each 1 mg/dL increase in HDL-C levels results in 2% reduced risk of fatal CVD events in men and 3% reduced risk in women. • HDL-C levels were statistically unrelated to noncardiovascular causes of mortality.

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TABLE 6. (continued) Study Name Meta-analysis (Austin MA, 1998)

Study Design Meta-analysis of 17 published prospective studies of TG and CVD covering over 57,000 Caucasian men and women over a period of 8–11 years.

Primary Outcomes • Men with at least 1 mmol/L (89 mg/dL) increase in TG compared with normal individuals had a 14% increased risk of CVD. • The risk of CVD increased by 37% in women with 1 mmol/L elevated triglyceride levels.

CHD = Coronary heart disease. CVD = Cardiovascular disease. HDL-C = High-density lipoprotein cholesterol. LDL-C = Low-density lipoprotein cholesterol. TG = Triglycerides. Note: Full source citations appear in ‘‘References.’’

foster the development of atherosclerosis and complications such as angina, myocardial infarction, peripheral arterial disease, and stroke. Risk factors such as smoking and dyslipidemia increase the likelihood of endothelial dysfunction and atherosclerosis. The Framingham Heart Study Þrst demonstrated that people with hypertension are at greater risk than the general population for cardiovascular disease (Kannel WB, 1969). Because hypertension can be asymptomatic for long periods (10–20 years or longer), it is often described as a “silent killer.” Cardiovascular events such as myocardial infarction or stroke are common primary manifestations of hypertension in undiagnosed and untreated individuals. Historically, elevated diastolic blood pressure (BP) has been associated with increased cardiovascular risk, but more recent epidemiological data stress the importance of systolic BP as a cardiovascular risk factor, particularly in patients older than 50 years. The Multiple Risk Factor Intervention Trial (MRFIT) was a 12-year follow-up investigating cardiovascular risk in 316,000 middle-aged men. In that study, death from coronary heart disease was almost linearly related to systolic BP, at all levels of diastolic BP (Vidt DG, 2004). CURRENT THERAPIES At this time, no pharmacological agents are formally approved for metabolic syndrome. Lifestyle modiÞcations such as diet and exercise are the cornerstones of therapy for metabolic syndrome patients. Most physicians recognize metabolic syndrome as a legitimate disorder that merits the effort to treat; however, only a fraction of physicians (PCPs, endocrinologists, and cardiologists) make formal diagnoses of metabolic syndrome. Furthermore, because there are no pharmacotherapeutic agents speciÞcally approved for metabolic syndrome, the only treatments currently available are drugs that are marketed for the individual diseases that make up the syndrome.

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FIGURE 2. Clinical progression from insulin resistance to type 2 diabetes.

To date, the lack of cost-effective therapies for obesity and unclear guidelines for the treatment of insulin-resistant borderline diabetics have limited drug treatment for metabolic syndrome. Although many agents can be used to treat each of the underlying components of metabolic syndrome, the most promising drugs are those with multimodal effects on at least two comorbid parameters. Table 7 lists the agents with at least some favorable activity on multiple components of

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metabolic syndrome that are currently approved for individual underlying indications. Table 8 details the advantages and disadvantages of these agents for the treatment of metabolic syndrome. The main rationale for treating patients with metabolic syndrome is that they are at high risk for the development of coronary heart disease (CHD). The longterm studies that are presented here focus on the prevention of cardiovascular end points or the prevention of progression to type 2 diabetes—the selected studies involve the use of drugs with effects on more than one component of metabolic syndrome. The patient inclusion criteria for many of these trials are reßective of the controversy surrounding different viewpoints on the diagnostic criteria for metabolic syndrome—many of them do not match either the Third Report of the Adult Treatment Panel of the National Cholesterol Education Program (NCEPATP III) or the World Health Organization’s (WHO) deÞnition of metabolic syndrome. However, these studies do provide rational clinical evidence that many currently marketed drugs may prove useful in the treatment of metabolic syndrome. This section reviews currently marketed therapies, their efÞcacy in their respective FDA-approved indications, and the clinical trials that suggest they may also improve other abnormalities associated with the metabolic syndrome. Antidiabetic Agents Overview. Insulin resistance, impaired glucose tolerance, and type 2 diabetes are all cornerstones of diagnostic criteria in the different deÞnitions of metabolic syndrome. Although diet and aerobic exercise can improve glycemic control in metabolic syndrome (by lowering body weight and improving insulin sensitivity), most patients need pharmacotherapies to effectively control serum glucose levels. This section discusses the antidiabetic agents that offer the most beneÞt to patients with metabolic syndrome—agents that have at least a minor effect on one other disease component of the syndrome beyond addressing glycemic control or insulin resistance. Biguanides Overview. Metformin (Bristol-Myers Squibb’s Glucophage, Glucophage XR, generics) is the only biguanide that has been developed for the treatment of type 2 diabetes. Biguanides have been a mainstay of treatment for type 2 diabetes since their introduction in Europe in 1950, and they continue to be the Þrst-line antidiabetic agent of choice among physicians in the United States and Europe. Metformin is sufÞciently safe and effective to treat prediabetic patients, many of whom fall under the deÞnition of metabolic syndrome, with fasting plasma glucose levels between 110 and 126 mg/dL (the American Diabetes Association’s [ADA’s] current deÞnition of prediabetes is fasting plasma glucose [FPG] greater than 100 mg/dL, but this value has not yet been adopted for metabolic syndrome). Importantly, in contrast to other antidiabetic agents, metformin is not associated with weight gain and is often associated with modest weight loss.

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TABLE 7. Current Therapies Used for Metabolic Syndrome Agent Biguanides Metformin

Company/Brand Bristol-Myers Squibb’s Glucophage, generics

Thiazolinediones Rosiglitazone GlaxoSmithKline’s Avandia Pioglitazone Takeda/Eli Lilly’s Actos Antidiabetic fixed-dose combinations Metformin/ GlaxoSmithKline’s Avandamet rosiglitazone Pancreatic lipase inhibitors Orlistat Roche’s Xenical

Daily Dose 500 tid, 850 mg bid

US, F, G, I, S, UK, J

4–8 qd or 2–4 mg bid

US, F, G, I, S, UK

15–45 mg qd

US, F, G, I, S, UK, J

500 mg/1 mg, 500 mg/2 US, UK mg, or 500 mg/4 mg bid

120 mg tid

Serotonin-norepinephrine reuptake inhibitors Sibutramine Abbott’s Meridia, 10–15 mg qd AstraZeneca’s Reductil HMG-CoA reductase inhibitors Atorvastatin Pfizer’s Lipitor/Tahor/Sortis/ Torvast/Cardyl Simvastatin Merck’s Zocor/Sinvacor, Boehringer Ingelheim’s Denan, generics Pravastatin Bristol-Myers Squibb’s Pravachol/Pravasin/Selectin/ Lipemol/Lipostat/Elisor, Sankyo’s Mevalotin/Sanaprav Fluvastatin Novartis’s Lescol/Locol, Solvay’s Digaril Rosuvastatin AstraZeneca’s Crestor Lovastatin Merck’s Mevacor/Mevinacor Pitavastatin Nissan/Kowa Kogyo/Novartis/ Sankyo’s Livalo

Availability

US, F, G, I, S, UK US, F, G, I, S, UK

10–80 mg qd

US, F, G, I, S, UK, J

10–80 mg qd

US, F, G, I, S, UK, J

10–40 mg qd

US, F, G, I, S, UK, J

20–80 mg qd, up to 40 mg US, F, G, I, S, UK, J tid 5–40 mg qd US, UK 10–80 mg qd US, G, S 1–4 mg qd

J

Selected statin fixed-dose combinations Lovastatin/ Kos’s Advicor Lowest dose is 500 mg US niacin niacin/20 mg lovastatin qd; maximum dose is 2,000 mg/40 mg qd. Atorvastatin/ Pfizer’s Caduet Lowest dose is 5 mg US amlodipine amlodipine/10 mg atorvastatin qd; maximum dose is 10 mg amlodipine/80 mg atorvastatin qd

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TABLE 7. (continued) Agent Simvastatin/ ezetimibe

Fibrates Fenofibrate

Bezafibrate

Gemfibrozil

Company/Brand

Daily Dose

Merck/Schering-Plough’s Lowest dose is 10 mg Vytorin/Inegy ezetimibe/10 mg simvastatin qd; maximum dose is 10 mg ezetimibe/80 mg simvastatin qd

Availability US

Fournier’s Lipan67 mg tid, 200 or 267 mg US, F, G, I, S, UK, J til/Lipidil/Fulcro/Secalip, qd Abbott’s Tricor Roche’s 200 mg up to tid F, G, I, S, UK, J Bezalip/Cedur/Befizal, generics Pfizer’s Lopid/Lipur, 450–750 mg bid US, F, G, I, S, UK, J generics

Angiotensin-converting enzyme inhibitorsa Enalapril Merck’s Vasotec/Renitec, 5–40 mg qd or bid generics Lisinopril Merck’s Prinivil, 5–40 mg qd or bid AstraZeneca’s Zestril, generics Ramipril King 1.25–20 mg qd or bid Pharmaceuticals/SanofiAventis’s Altace/Delix/Tritace Angiotensin II receptor antagonistsa Losartan Merck’s Cozaar/Lozaar Valsartan Novartis’s Diovan/Tareg, Sanofi-Aventis’s Nisis Irbesartan Bristol-Myers Squibb/ Sanofi-Aventis’s Avapro/Aprovel/Karvea Candesartan AstraZeneca’s cilexetil Atacand/Amias, Takeda’s Blopress Olmesartan Sankyo’s Benicar/Olmetec

US, F, G, I, S, UK, J US, F, G, I, S, UK, J

US, F, G, I, S, UK

25–100 mg qd or bid 40–320 mg qd

US, F, G, I, S, UK, J US, F, G, I, S, UK, J

150–300 mg qd

US, F, G, I, S, UK

4–32 mg qd or bid

US, F, G, I, S, UK, J

10–40 mg qd

US, G, UK, J

a According to the package inserts for the drugs in this class, these agents can be taken once daily or twice

daily, depending on the needs of the patient. In the case of twice-daily administrations, the maximum daily dose provided here is not intended to be taken twice a day. bid = Twice daily; qd = Once-daily; tid = Three times daily. US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan.

Mechanism of Action. Metformin’s precise mechanism of action is unknown. However, research shows that its primary action is to reduce hepatic glucose production—which is elevated in diabetes patients—by inhibiting the activity of glucose-6-phosphatase (Hundal RS, 2000; Mithieux G, 2002; Stumvoll M, 1995) (Figure 3). Other studies indicate that unlike two other antidiabetic classes that target pancreatic β-cell insulin secretion (the sulfonylureas and meglitinides),

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TABLE 8. Comparison of Current Therapies for Metabolic Syndrome Class

Advantages

Disadvantages

Biguanides

• Not associated with weight gain. • Does not cause hypoglycemia. • Beneficial effect on lipid subfractions.

• Gastrointestinal irritation in some patients during the first few weeks of therapy. • No major effects on lipid levels.

Thiazolinediones (PPARγ agonists)

• Beneficial effects on lipid levels and lipoprotein particle size with pioglitazone. • Short-term benefit on pancreatic beta cell function.

• Associated with weight gain, fluid retention, and hemoglobin decline.

Antidiabetic fixed-dose combinations

• Coadministration of two agents can result in a greater decrease in blood glucose levels compared with singleagent therapy. • Convenience.

• Fixed dosing is limiting.

Pancreatic lipase inhibitors

• Weight loss. • Reduction in serum lipid levels. • Reduction in incidence of type 2 diabetes.

• Gastrointestinal side effects include fecal incontinence, flatulence, and oily spotting.

Serotonin and norepinephrine reuptake inhibitors

• Weight loss. • Reduction in serum lipid levels. • Minor improvements to glycemic control.

• Elevated blood pressure. • Increased heart rate. • Dry mouth, nausea, insomnia.

HMG-CoA reductase inhibitors

• Reduction in LDL-C levels. • Minor reductions in TG levels and minor increases in HDL-C when taken at high doses. • Possible anti-inflammatory effect when taken at high doses.

• High doses may cause rhabdomyolysis (muscle breakdown).

Selected statin fixed-dose combinations

• Convenience. • Possible synergistic effect from dual mechanisms of action. • Improved compliance.

• Fixed dosing is limiting.

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TABLE 8. (continued) Class Fibrates

Angiotensin-converting enzyme inhibitors

Angiotensin II receptor antagonists

Advantages • Reduction in TG levels. • Modest improvement HDL.

Disadvantages

to

• Effective antihypertensive agent. • Well tolerated by patients. • May have beneficial effects on endothelial dysfunction. • Effective antihypertensive agent. • Fewer side effects compared with ACE inhibitors.

• Myositis (muscle inflammation) in patients with renal impairment. • May reduce libido or cause impotence. • Minor side effects include nausea, loss of appetite, and stomach pain. • Persistent dry cough. • Possible risk of angioedema.

• Possible risk of angioedema and orthostatic hypotension. • May not have the same secondary benefits as ACE inhibitors.

ACE = Angiotensin-converting enzyme. HDL-C = High-density lipoprotein cholesterol. LDL-C = Low-density lipoprotein cholesterol. TG = Triglycerides.

metformin also has an effect on insulin sensitization (Lupi R, 1999; Mather KJ, 2001). Peroxisome Proliferator-Activated Receptor Agonists (Thiazolidinediones) Overview. Peroxisome proliferator-activated receptor (PPAR) agonists (also called thiazolidinediones, or TZDs) are the newest class of insulin sensitizers—they increase insulin sensitivity by enhancing its effects at peripheral target sites. Rosiglitazone (GlaxoSmithKline’s Avandia) (Figure 4) and pioglitazone (Takeda/Eli Lilly’s Actos) (Figure 5) are the only TZDs currently available in the major pharmaceutical markets. Troglitazone (Warner-Lambert’s Rezulin, GlaxoSmithKline’s Romozone, Sankyo’s Noscal) (Figure 6) was withdrawn from the market in Europe in 1998 and in the United States and Japan in 2000 because of the potential for fatal liver toxicity. Clinical studies demonstrate that rosiglitazone and pioglitazone offer comparable improvements to glycemic control; however, pioglitazone causes reductions in triglyceride and low-density lipoprotein cholesterol (LDL-C) levels and a modest increase in high-density lipoprotein cholesterol (HDL-C), giving it a more favorable proÞle for use in metabolic syndrome patients (Olansky L, 2003; Boyle PJ, 2002). These Þndings were recently conÞrmed in a head-to-head study involving 735 type 2 diabetic patients who were randomized to receive either

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FIGURE 3. Mechanism of action of metformin.

FIGURE 4. Structure of rosiglitazone.

FIGURE 5. Structure of pioglitazone.

pioglitazone (30 mg/day) or rosiglitazone (4 mg/day) for 24 weeks (Goldberg RB, 2004). Patients in the pioglitazone and rosiglitazone treatment arms saw similar reductions to HbA1c levels (−0.7% and −0.6% reduction, respectively). However, patients receiving pioglitazone experienced, on average, 12% lower TG, 15% increased HDL-C, and 16% increased LDL-C, whereas those receiving rosiglitazone saw a 15% increase in TG, an 8% increase in HDL-C, and a 23% increase in LDL-C. Both rosiglitazone and pioglitazone are associated with weight gain, ßuid retention, and hemoglobin reduction, as the labeling indicates. Guidelines released by the National Institute for Clinical Excellence (NICE) in the United Kingdom indicate that TZDs may be used only in combination with metformin or a

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FIGURE 6. Structure of troglitazone.

FIGURE 7. Mechanism of action of thiazolidinediones.

sulfonylurea in diabetic patients. However, due to the hepatotoxicity observed with troglitazone, the guidelines recommend that patients be routinely monitored for liver toxicity during the Þrst year of treatment. Mechanism of Action. Activation of PPAR-γ improves glycemic control by enhancing insulin sensitivity, as illustrated in Figure 7. In the presence of endogenous or exogenous insulin, PPAR-γ agonists increase glucose uptake and utilization in skeletal muscle, boost glucose uptake and reduce fatty-acid output in adipose tissue, and reduce gluconeogenesis, glucose output, and triglyceride synthesis in the liver. Key Clinical Trials. Small, dense LDL-C particles are gaining acceptance as an important risk factor for cardiovascular disease (CVD). In addition to the beneÞcial effects on glycemic control exerted by different oral antidiabetic agents (pioglitazone, metformin, and gliclazide [a sulfonylurea]), the effects of these agents on lipid levels and lipoprotein subfractions (e.g., small, dense LDL-C particles) were examined in 60 overweight patients with type 2 diabetes (Lawrence JM, 2004). Although no signiÞcant improvements to lipid levels were seen, the ratio of small, dense LDL-C particles decreased in patients taking pioglitazone (30–45 mg daily) or metformin (500–1,000 mg twice daily). No effect on particle density was observed in patients taking gliclazide (80–160 mg twice daily). In addition, the ratio of dense HDL3 particles to the less dense HDL2 particles decreased in patients treated with either metformin or pioglitazone. The decrease

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observed in the pioglitazone arm resulted from an increase in the HDL2 level and an increase in total HDL-C levels. In the metformin arm, a decrease in HDL 3 level was observed. The importance of lipid subfractions in metabolic syndrome may alter screening and treatment paradigms. Alpha-Glucosidase Inhibitors Overview. The primary effect of alpha-glucosidase inhibitors (AGIs) is to lower postprandial plasma glucose (PPG) levels and to improve glycemic control without causing weight gain (a problem with many antidiabetic agents) or increasing the risk of hypoglycemic events. However, the AGIs generally exert only a modest effect, reducing glycosylated hemoglobin HbA1c levels by 0.5–1.0% (Inzucchi SE, 2002). These agents are used as alternatives for patients, particularly the elderly, who have mild to moderate hyperglycemia and for patients who are prone to hypoglycemia or lactic acidosis. Acarbose (Bayer’s Precose/Glucobay, generics), miglitol (Bayer/PÞzer/SanoÞ-Aventis’s Glyset), and voglibose (Takeda’s Basen) are the only AGIs currently available in the major markets. Voglibose is available only in Japan. Mechanism of Action. AGIs hinder carbohydrate digestion at the brush border of the intestinal epithelium, effectively delaying carbohydrate absorption and allowing the delayed insulin secretion characteristic of type 2 diabetics to catch up with carbohydrate absorption (Figure 8). Consequently, AGIs reduce PPG levels. These agents concentrate in the intestinal enterocytes, where they act locally, minimizing the risk of hypoglycemia. Key Clinical Trials. The effects of one alpha-glucosidase inhibitor, acarbose, on CVD risk in patients with impaired glucose tolerance (IGT) was examined in the Stop—Non-Insulin-Dependent Diabetes Mellitus (STOP-NIDDM) trial. In this double-blind trial, 1,368 patients with IGT were randomized to receive acarbose (Bayer’s Precose; 100 mg three times daily) or placebo (Chiasson J-L, 2003). Patients were followed for a mean of 3.3 years. The primary end point of the trial was the frequency of major cardiovascular events. Acarbose treatment reduced the risk of cardiovascular events by 49% and the risk of developing hypertension by 34%. These results suggest that reducing postprandial hyperglycemia is associated with a signiÞcant reduction in cardiovascular risk—a beneÞt with important implications for patients with metabolic syndrome. Antidiabetic Fixed-Dose Combinations Overview. For patients who require therapy with more than one agent, singlepill, Þxed-dose combination agents provide a convenient alternative to taking two separate pills. All available combination agents contain metformin plus an additional oral antidiabetic agent, thus reßecting metformin’s core position in the type 2 diabetes therapeutic regimen. The only single-pill combination agent currently on the market that may be effective in metabolic syndrome patients is metformin/rosiglitazone (GlaxoSmithKline’s Avandamet). In addition to metformin, which is associated with a minor weight loss, this combination includes

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FIGURE 8. Mechanism of action of alpha-glucosidase inhibitors.

the insulin-sensitizing agent rosiglitazone. The greater convenience of the onepill regimen may lead to an increase in patient compliance, which may in turn improve glycemic control, particularly in metabolic syndrome patients who are on multiple medications for the various disease components of the disorder. Mechanism of Action. The metformin/rosiglitazone combination (GlaxoSmithKline’s Avandamet) possesses two mechanisms of action courtesy of its single-pill formulation of two individual antidiabetic agents. For further details, see the ”Mechanism of Action” section for the biguanides and the PPAR agonists. Antiobesity Agents Overview. Obesity is believed to be a major factor contributing to the development of metabolic syndrome. Although lifestyle modiÞcations such as diet and exercise are effective treatments for metabolic syndrome, patient compliance tends to be poor. Therefore, many patients require pharmacotherapy to aid in weight loss. Weight loss can also prevent the onset of metabolic syndrome in borderline patients and effect improvements in each of the disease components that make up metabolic syndrome. Clinical trials show that the risks of developing complications such as cardiovascular disease and diabetes can be reduced by weight loss. In addition to prescribing a comprehensive program of dieting, physical exercise, and behavioral therapy, some physicians are including pharmacological agents as an adjunctive therapy in some cases of obesity. Only orlistat (Roche’s Xenical) (Figure 9) and sibutramine (Abbott’s Meridia) (Figure 10) have been approved for long-term (one year or longer) treatment of obesity.

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FIGURE 9. Structure of orlistat.

FIGURE 10. Structure of sibutramine.

Pancreatic Lipase Inhibitors Overview. Orlistat is the only pancreatic lipase inhibitor for the treatment of obesity currently available; it has been launched in both the United States and Europe. Unlike systemically acting appetite suppressants, pancreatic lipase inhibitors act locally in the intestinal lumen to decrease the absorption of dietary fats. Although their side effects are unpleasant (e.g., oily spotting, bowel urgency, diarrhea), this class of drugs is safer than the centrally acting weight-loss drugs (i.e., sibutramine [Abbott’s Meridia; AstraZeneca’s Reductil]). Mechanism of Action. Gastric and pancreatic lipases transform dietary triglycerides into free fatty acids and monoglycerides so that the small intestine can absorb them during digestion. Pancreatic lipase inhibitors inhibit gastric, pancreatic, and carboxyl ester lipases to decrease dietary free fatty acid absorption by 30%. The decreased absorption results in a concomitant increase in fat excretion. This class of drugs is believed to cause weight loss by reducing the number of calories derived from ingested fat. As illustrated in Figure 11, these inhibitors act peripherally within the intestinal lumen rather than centrally in the brain. Key Clinical Trials. In addition to causing moderate weight loss, orlistat has been associated with a reduction in serum lipid levels. In a study of 444 obese subjects with hyperlipidemia, weight loss induced by treatment with orlistat in conjunction with a reduced-calorie diet resulted in statistically signiÞcant decreases in total cholesterol, LDL-C, and fasting insulin levels compared with patients who lost weight by dieting alone (Lucas CP, 2003). Results from several clinical trials

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FIGURE 11. Mechanism of action of orlistat.

demonstrate that patients exhibit, on average, a 10% reduction in total cholesterol and LDL-C levels, independent of weight loss (Hollander PA, 1998; Davidson MH, 1999; Rossner S, 2000; Finer N, 2000). In addition, several orlistat trials have shown that in obese patients with hypertension, blood pressure is lowered proportionally to weight loss (Davidson MH, 1999; Rossner S, 2000). In 2004, following completion of a large-scale study known as Xenical in the Prevention of Diabetes in Obese Subjects (XENDOS), both the FDA and the European Commission approved the labeling of orlistat for the reduction of the risk of developing type 2 diabetes. In this double-blind, placebo-controlled prospective study, 3,305 obese, nondiabetic patients were followed over a fouryear period. All patients underwent lifestyle changes in addition to receiving either placebo or orlistat (120 mg three times daily). The two primary end points were the time to onset of type 2 diabetes and the change in weight after four years. Patients receiving orlistat not only lost more weight than those receiving placebo (on average, 5.7 kg versus 3.0 kg, respectively) but also exhibited a 37% reduced incidence of type 2 diabetes over the course of the study (Torgerson JS, 2004). In addition, treatment with orlistat improved risk factors of metabolic syndrome by lowering blood pressure, decreasing waist circumference, and lowering LDL-C. Although weight loss is known to improve lipid values, patients in the study who took orlistat were able to lower their LDL levels but exhibited no improvements to HDL levels compared with those taking placebo. Other studies have shown that orlistat can reduce the severity of several diagnostic markers for type 2 diabetes in obese diabetic patients. P.A. Hollander and colleagues reported that patients taking orlistat improved their insulin sensitivity

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Obesity

ETIOLOGY AND PATHOPHYSIOLOGY Introduction Beyond the simple explanation for the cause of obesity—an imbalance of energy intake and expenditure—is a range of factors affecting energy consumption. These factors include complex signals that inßuence food intake by regulating feelings of hunger and satiety. In addition, genetic predisposition, environmental variables, and cultural norms all contribute to the development of obesity. Researchers have made considerable progress in identifying pathophysiological processes that lead to obesity, but much remains unknown in this complex Þeld. Etiology The etiology of obesity involves complex polygenic factors that are not yet well understood. Some cases of obesity arise from genetic alterations or damage to regions of the brain that are important in controlling feeding behavior. Table 1 lists some of the disorders that cause obesity. It is important to note, however, that the majority of obesity cases are driven by environmental and behavioral factors acting in concert with a genetic predisposition to maintaining and gaining fat mass. Before discussing the etiology of obesity, this section provides an overview of some of the factors that regulate the storage and expenditure of energy in the human body as well as the normal physiology of hunger and satiety signaling. Regulation of Energy Storage. Much of the food humans eat is broken down into glucose molecules, which are used to support the body’s metabolic Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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TABLE 1. Disorders Causing Obesity Disorder Congenital abnormalities (e.g., Prader-Willi syndrome) Cushing’s syndrome Hypothalamic tumor, infection, or injury

Hormonal disorders (hypogonadism, growth hormone deficiency, insulinoma) Hypothyroidism

Relationship to Weight Gain Increased food intake triggered by a chromosomal disorder that results in hypothalamic impairment. Fat deposition in truncal area of body. Increased food intake caused by hypothalamic changes (e.g., increased parasympathetic activity or decreased sympathetic activity). Increased food intake.

Weight gain caused by decreased energy expenditure.

activities. A portion of the glucose is stored as glycogen in the liver and in muscle tissue. Excess glucose and fatty acids are stored in the form of triglycerides in adipocytes, which are specialized fat storage cells. When the body requires energy beyond that supplied by the available glycogen stores, adipocytes release triglycerides as fatty acids into the bloodstream. This process, also known as lipolysis, results in the shrinking of individual adipocytes and a decrease in overall adipose tissue deposits. Most cases of adult-onset obesity are associated with enlarged fat cells (hypertrophic obesity) rather than an increase in the number of fat cells (hyperplastic obesity). White adipocytes are the most common type of fat cells found in humans. These cells serve as storage depots for fat and also provide insulation and cushioning for the internal organs. White adipose tissue (WAT) is found throughout the body and forms deposits of fat under the skin and over the organs. Scientists have discovered that WAT, which was previously thought to have only a simple storage function, is hormonally active and performs important endocrine functions, including the release of leptin, tumor necrosis factor, angiotensin, estrogen, and cholesterol ester transfer protein. A second type of fat cells found in humans is brown adipose tissue (BAT). BAT is composed of highly active cells that are sparse in adult humans but plentiful in animals and newborn humans. The primary role of BAT, although not fully understood by researchers, appears to be the production of heat (thermogenesis) rather than energy storage. Leptin is the product of the obesity gene ob (also called the LEP gene) and is a key factor in both long- and short-term appetite regulation. This hormone has attracted signiÞcant attention from researchers and the media over the past decade because of its proposed role as a central regulator of body weight and adiposity. Leptin is released by WAT and is transported across the blood-brain barrier to the hypothalamus, where it helps the brain monitor how much energy is stored in the body as fat. As would be expected, obese humans, who possess large amounts of WAT, have high levels of circulating leptin. In humans of normal weight, leptin works well to allow individuals to avoid over- or undereating. Leptin-sensitive nerve

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cells in the medial hypothalamus release anorectic neuropeptides that act to suppress appetite through mediators in the lateral hypothalamus. At the same time, leptin suppresses the release of the appetite stimulant neuropeptide Y (NPY). An emerging theory of obesity claims that this system malfunctions in obese people, leading to leptin resistance. A tenet of this hypothesis is that leptin receptors are saturable, such that once a certain level of leptin is reached in the blood, additional leptin cannot be transported to the hypothalamus. Neurons produce the protein SOCS3 (suppressor of cytokine signaling-3) in response to leptin, and this suppressor contributes to leptin resistance by rendering leptin receptors insensitive to additional leptin signaling (Clapham JC, 2001). Other molecules that researchers believe play a part in the leptin-signaling pathway include ciliary neurotrophic factor (CNTF), melanin-concentrating hormone (MCH), and melanocyte-stimulating hormone (MSH). Drug research is now focusing on direct delivery of leptin, on analogues that may stimulate receptors more effectively than endogenous leptin, and on drugs that target the CNTF, NPY, MCH, and MSH pathways. Regulation of Energy Expenditure. Three basic components account for daily energy expenditure. The largest, accounting for 60–70% of all energy expenditure, is the resting metabolic rate (RMR), or the rate at which a person uses energy to maintain bodily processes such as respiration, circulation, and brain activity while in a resting state. RMR varies by individual and depends on the amount of lean body mass. Because of alterations in body composition, obese people have more of both lean and fatty body mass. Hence, obese individuals generally have a higher RMR than normal-weight individuals (contrary to the popular belief that obese people have “slow metabolisms”). Another component of daily energy expenditure is diet-induced thermogenesis, the process by which food intake triggers an increase in metabolism. This metabolic increase occurs following each meal and is required to maintain digestion. Much of the energy used during digestion is lost in the form of heat. This process accounts for approximately 20% of daily energy expenditure. The Þnal component of daily energy expenditure, physical activity, accounts for approximately 10–20% of daily energy expenditure. The adrenergic system plays a major role in controlling energy expenditure. β3 -adrenergic receptors are located in both WAT and BAT. The nervous system is thought to act on BAT cells through β3 -adrenergic receptors, which, when activated, stimulate mitochondrial uncoupling protein-1 (UCP-1) to release stored energy as heat. The proteins UCP-2 and UCP-3 are found in skeletal tissue and other sites in the body, and researchers are investigating their role in energy consumption. In WAT, activated β3 -adrenergic receptors stimulate lipolysis. Some researchers propose that body weight might be inßuenced by manipulating the energy-expending activities of BAT or by causing WAT to behave more like BAT by burning off fat stores to create heat. Regulation of Food Intake. A network of interacting systems controls shortterm (meal-to-meal) feeding behavior: signals originating in the brain and the

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FIGURE 1. Neural and hormonal mediators of hunger and satiety signals.

peripheral nervous system provide feedback to one another, and the central nervous system (CNS) monitors divergences from balanced equilibrium and adjusts eating behavior accordingly. Figure 1 illustrates the many chemical factors that inßuence normal food intake. Because of the myriad mechanisms involved in the development of obesity, any drug targets that emerge will likely be effective for only certain patient groups. This section describes some of the most inßuential hunger and satiety signaling pathways in weight regulation, many of which remain to be fully explored in the search for a cure. Central Satiety and Hunger Signals. The brain stem monitors satiety and hunger signals with receptors that are sensitive to nutrients as well as to signals sent by the stomach, liver, and duodenum. The hypothalamus is responsible both for inducing and for inhibiting food intake. In particular, the lateral hypothalamus (LH) and the ventromedial nucleus (VMN) are responsible for hunger and satiety,

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FIGURE 2. Hypothalamic structures of the brain involved in food intake regulation.

respectively. The neurosecretory cells of the paraventricular nucleus (PVN) have also been shown to be involved in food intake, although the mechanisms are still poorly understood. These three regions affect behavioral aspects of feeding and regulate the body’s growth and other metabolic activities; they also affect peripheral regulation by mediating signals from the CNS. Figure 2 details the role these regions play in regulating food intake. •

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Serotonin. Researchers have found that the neurotransmitter serotonin (5-HT) triggers a feeling of satiety. Administration of serotonin to the brain, or the use of drugs to enhance its activity, reduces food intake. Serotonin achieves this response by acting on the noradrenergic and endogenous opioid pathways. Researchers have proposed that serotonin helps determine food preference and that elevated serotonin levels suppress carbohydrate intake and increase protein intake. Some appetite suppressants work by triggering the release of serotonin, by inhibiting its reuptake, or both. Norepinephrine. Norepinephrine (also called noradrenalin) is a catecholamine neurotransmitter that can cause increased or decreased food intake depending on the receptor to which it binds. Activation of the α2 adrenergic receptor in the PVN stimulates food intake (Wellman PJ, 2000), while stimulation of the α1 - and β2 -adrenergic receptors in the perifornical area (adjacent to the LH) has the opposite effect. Norepinephrine increases glycogenolysis (conversion of glycogen to glucose) in the liver and lipolysis (conversion of triglycerides into fatty acids) in adipose tissue. Sibutramine (Abbott’s Meridia) is a currently available antiobesity

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agent that functions by inhibiting norepinephrine reuptake by postsynaptic neurons. Opioids. Endogenous opioids (including dynorphins and endorphins) are morphine-like substances that are produced in the pituitary gland and induce a preference for sweets and fatty foods. Excessive feeding behavior in genetically obese rodents can be alleviated by administration of naloxone, an opioid receptor antagonist. Researchers have found raised endorphin levels in the plasma of obese people and have hypothesized that opioids may increase feeding behavior by stimulating the reward centers in the brain, which reinforces the pleasurable feelings associated with eating certain foods. Neuropeptide Y. NPY, a 36-amino-acid peptide found in the hypothalamus, is the most potent orexigenic (appetite-stimulating) peptide. NPY encourages the intake of carbohydrates, acting independently of norepinephrine to block satiety signals and reduce energy expenditure. In addition, NPY has been associated with anxiety and depression, endocrine function, and metabolism. Impaired regulation of NPY gives rise to inappropriate neurohormonal control of metabolism, resulting in persistent obesity. The NPY5 receptor in the hypothalamus has been identiÞed as the receptor most closely linked with eating behavior and is therefore a focus of discovery efforts for drugs to suppress appetite. One of the mechanisms by which the hormone leptin is thought to regulate food intake is by inhibition of NPY synthesis. Galanin. Galanin, a 29-amino-acid peptide, is found in high concentrations in the hypothalamus and the gastrointestinal (GI) tract. Experiments in rats have shown that interaction of this substance with speciÞc receptors in the hypothalamus of satiated rats stimulates food intake (especially of fatty foods). Some researchers suggest that a selective galanin antagonist could be a promising target for drug discovery. Cannabinoids. Endogenous cannabinoids are known to stimulate appetite. The CB1 endocannabinoid receptor is localized mainly in the brain, including regions of the hypothalamus. Like the endogenous opioids, endocannabinoids are believed to enhance appetite by activating the brain’s reward centers, which reinforces the pleasurable feelings associated with eating certain foods. Administration of cannabinoid receptor antagonists has been shown to decrease food intake, thereby causing weight loss in both humans and animals. Orexins. Orexins A and B are peptides in the hypothalamus that stimulate appetite. They are derived from prepro-orexin (or prepro-hypocretin). Some researchers suggest that orexins may be short-term regulators of eating that are stimulated by the falls in glucose that precede eating. Two receptors—OX-1 (selective for orexin A) and OX-2 (selective for orexins A and B)—have been identiÞed. OX-2 receptor defects have been connected with narcolepsy in research animals, and therefore this pathway may not be a

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suitable drug target for obesity because of the possibility of similar side effects in humans (Lin L, 1999). Peripheral Satiety and Hunger Signals. Peripheral chemical regulators secreted throughout the body provide feedback to the brain on metabolism and digestion rates. These regulators provide feedback directly (by stimulating the vagus nerve) or indirectly (by entering the circulatory system to reach the brain). •

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Insulin. Insulin is a well-known peripheral hormone that stimulates the uptake of glucose, amino acids, and fatty acids. Insulin also promotes the synthesis of glycogen, protein, and lipids. Researchers have demonstrated that insulin acts on the CNS to suppress appetite by inhibiting the synthesis and secretion of NPY. Plasma insulin levels fall during fasting, which results in a rise in NPY secretion that prompts a return of appetite. (Obesity is closely related to insulin resistance and type 2 diabetes, as discussed later in this section.) Researchers have shown a direct correlation between insulin levels in plasma and the amount of adipose tissue in the body. Because many obese people also have hyperinsulinemia, it is thought that the feedback system that involves insulin as a regulator of appetite and satiety may be impaired in some obese people. Whether this impairment results from the failure of insulin to reach the brain or from a defect in the relevant receptors remains unknown. Cholecystokinin. The peptide hormone cholecystokinin (CCK) interacts with CCK-A receptors (located in peripheral areas such as the gut) and CCK-B receptors (located in the brain). As a result, CCK has a dual action: it works both peripherally and centrally to slow gastric emptying and inhibit food intake. CCK is released in the gut during mealtimes in response to ingested proteins and fat. Researchers hypothesize that its main purpose is to aid in digestion by limiting further food intake after meals and causing increased secretory activity in the gallbladder and pancreas. Bombesin. Bombesin is found in the GI tract and CNS of mammals and plays a role in regulating appetite. The exact mechanism through which this peptide functions remains unclear; however, researchers believe that it may function both peripherally and centrally. Research in healthy humans has shown that bombesin inhibits calorie intake. Glucagon-Like Peptide-1. The GI tract releases glucagon-like peptide 1 (GLP-1) in response to food intake. GLP-1 receptors are present in the gut, liver, pancreas, and CNS. GLP-1 is an insulin-like hormone that delays gastric emptying, stimulates the release of insulin from the pancreas, and decreases glucose concentration in the blood. GLP-1 has also been shown to exhibit both lipogenic and lipolytic effects on adipocytes, depending on the concentration of the hormone. Amylin. The pancreas releases amylin, which is a potent inhibitor of gastric emptying. In rodent models, an increase in amylin was associated with a

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decrease in food intake, suggesting that this peptide may play a role in satiety signaling. Enterostatin. Enterostatin is a pentapeptide generated in the intestines following consumption of high-fat meals. In rodents, peripheral or central administration of enterostatin has been shown to limit fat intake by reducing food consumption. No speciÞc receptors for enterostatin have yet been identiÞed.

Genetic Factors. The speciÞc contribution of genes to the development of obesity is an area of great interest, not only because of the potential for the development of appropriate therapies, but also for helping researchers to understand why some people have so much trouble losing weight and maintaining weight loss. A variety of studies—examining family groups, sibling pairs, and twins reared together and apart—have attempted to quantify the statistical link between obesity and heredity. Also, studies of people who have managed to keep their weight constant are helping to provide additional information. Throughout these studies, a common Þnding is that shared genes have a greater effect on adult weight gain than a shared environment, with estimates of genetic contribution to variations in weight gain ranging from 40% to 80%. Researchers have studied single-gene defects in rodents and have successfully cloned the genes for several models of obesity in these animals. Five major models of obesity in mice and three in rats have been widely studied. Table 2 describes the Þve mouse models of obesity. Animal models lend important insight into possible interactions between genetics and obesity, and learning the functions of the products of genes allows scientists to examine new pathways in the search for drug therapies. Human obesity is more complex than the obesity found in animal models and likely has multiple heterogeneous etiologies. The connection between genes and obesity is not as clear in humans as in animals, and except in very rare instances, single-gene mutations do not cause human obesity. Table 3 lists select genetic mutations that have been observed in rare cases of extreme obesity or early-onset obesity in humans. Environmental Factors Behavior. Along with recent discoveries of genetic causes of obesity comes the temptation to blame weight gain solely on genetic factors. However, the gene pool has not changed during the past century, while the prevalence of overweight and obesity has increased substantially. Since the spread of industrialization, food has become more abundant and varied. With this emergence has come a trend toward satisfying the desire for high-calorie and fatty foods. Furthermore, economic factors and the increasing pace of modern lifestyles often results in the selection of foods that are cheap and convenient, yet low in nutritional quality. Simultaneously, the development of the automobile, the television, and other

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TABLE 2. Mouse Models of Obesity Resulting from Single Gene Mutations Mutation and Gene Product ag (agouti) or Ay (agouti yellow); agouti signaling protein

ob (obese) or Lepob ; leptin hormone

db (diabetes) or Leprdb ; leptin receptor

fat or Cpefat ; carboxypeptidase E

Tub (tubby); tubby protein

Mechanism Mice carrying this dominant mutation exhibit a characteristic yellow coat, late-onset obesity, and insulin resistance. Overexpression of agouti-signaling protein blocks the action of α-melanocyte-stimulating hormone at the melanocortin-1 and melanocortin-4 (MC-4) receptors. MC-4 receptors are expressed in the hypothalamus where they participate in the regulation of feeding behavior. Disruption of MC-4 receptor function by the agouti protein results in increased feeding. An agouti-related protein (AGRP) with possible roles in obesity has been found. Mice that are homozygous for a mutation in the ob gene display signs of overeating and characteristics of starvation, such as reduced energy expenditure and infertility, caused by defective leptin production. Leptin-deficient ob/ob mice are overweight from the time of birth onward. Mice carrying the db allele of Lepr have a defect in leptin receptor production. db/db mice exhibit characteristics similar to those of ob/ob mice, including overeating and weight gain. Injection of leptin has little or no effect on feeding behavior or weight loss in db/db mice. Researchers discovered that a defect in this gene (involved in the prohormone processing pathway) in mice leads to both late-onset obesity and diabetes. The defect is caused by a mutation in the carboxypeptidase E (CPE) gene, which is required to process peptides that act on melanocortin receptors in the central nervous system. This gene was isolated from the tubby mouse, which becomes overweight after sexual maturity. Mice with this genotype are also blind and deaf. Researchers believe that obesity may arise from a malfunction of signals to the brain’s satiety center. The tubby gene encodes a G protein-coupled receptor-activated transcription factor. Several tubby-like proteins (TULPs) have also been discovered.

inventions has allowed people to indulge in a sedentary way of life. Consequently, the average weight of the population has climbed steadily, along with the prevalence of overweight and obesity. Reduced Physical Activity. Physical activity represents only 10–20% of total daily energy expenditure. However, exercise can stimulate the production of mediators that have additional effects on hunger, weight, and energy storage. With this in mind, a great controversy continues over the effect that exercise

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TABLE 3. Candidate Genes for Obesity in Humans Genetic Defect Leptin (LEP)

Leptin receptor (LEPR)

Prohormone convertase 1 (PC1) Proopiomelanocortin (POMC)

Melanocortin 4 receptor (MC-4R)

Melanocortin 3 receptor (MC-3R)

Peroxisome proliferator-activated receptor-γ 2 (PPAR-γ 2) Neuropeptide Y receptor Y5 (NPY5R) Single-minded (SIM1)

Comments Autosomal recessive mutations in the gene for leptin, leading to leptin deficiency, have been observed in patients with early-onset obesity (Montague C, 1997; Ozata M, 1999). Autosomal recessive mutations in the gene for the leptin receptor have been linked to obesity (Clement K, 1998). Mutations in the gene for PC1 are associated with early-onset obesity (Jackson RS, 1997). Heterozygous deletions of the POMC gene have been observed in patients with severe obesity (Krude H, 2000). A range of mutations in MC-4R results in a common form of severe early-onset obesity (Farooqi IS, 2000; Farooqi IS, 2003; Vaisse C, 2000). A combination of early-onset obesity, high body fat content, and insulin resistance have been observed in members of a family with a single point mutation in MC-3R (Lee YS, 2002). A missense mutation in PPAR-γ 2 results in an autosomal dominant form of obesity (Ristow M, 1998). Polymorphisms in the noncoding region of the NPY5R gene are strongly linked to obesity in nondiabetic Pima Indians (Jenkinson CP, 2000). A translocation of the human homolog of the SIM1 gene from fruit flies is believed to interfere with MC-4 receptor function in an individual exhibiting early-onset obesity due to increased food intake (Holder JL, 2000).

Note: Full source citations can be found in ‘‘References.’’

has on body weight and obesity. Although many studies have found associations between reduced exercise and obesity, some researchers postulate that obesity actually causes reduced activity levels. Pathophysiology The health risks associated with obesity are well known. Body fat distribution is a key predictor of obesity-related health risks. Central obesity, also known as android or abdominal obesity, is deÞned as excess weight around the abdomen. It is characterized by an apple-shaped physique and is more common in men. Excess weight around the abdomen is associated with an increased risk of comorbid disease compared with peripheral or gynecoid obesity (an excess of weight in the gluteofemoral area), which predominates in women and results in a pear-shaped physique. The reason for the heightened risk with abdominal obesity is the excess of visceral fat (fat covering the organs). Visceral fat is metabolically and hormonally more active than fat stored elsewhere, and results in an increased concentration of

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FIGURE 3. Prevalence of comorbidities in obese and overweight U.S. men and women from 1988 to 1994.

free fatty acid in the blood. Increased risk of heart disease, hypertension, peripheral arterial disease, stroke, type 2 diabetes, gallbladder disease, osteoarthritis, and cancer, as well as insulin resistance, hyperinsulinemia, and hyperlipidemia, have all been associated with this type of central body fat distribution. Regardless of fat distribution, however, the relative risk of developing these conditions rises with increasing body weight. A variety of studies have shown that obesity is associated with an increased risk of mortality (Allison DB, 1999;

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Barbagallo CM, 2001; Calle EE, 1999; Katzmarzyk PT, 2001). A report from the U.S. Centers for Disease Control and Prevention (CDC) predicts that obesityrelated deaths will soon supersede deaths from smoking-related illnesses as the top cause of mortality in the United States (Mokdad AH, 2004). Researchers have determined the prevalence of six major comorbidities by class based on the results of the National Health and Nutrition Examination Survey (NHANES) III (Must A, 1999; illustrated in Figure 3). These researchers found strong correlations between excess weight and comorbidities. Compared with the prevalence of these comorbidities in normal weight individuals, even overweight individuals have signiÞcantly elevated health risks. Table 4 describes the major complications related to obesity. The prevalence of type 2 diabetes and hypertension has a particularly strong relationship with overweight and obesity, and increases proportionately by the severity of the obesity. These trends are seen in both men and women. CURRENT THERAPIES The vast majority of obese people never seek the advice of a physician in their efforts to lose weight. Most attempt to lose weight on their own, through calorie reduction, fad diets, meal replacements, exercise programs, over-the-counter (OTC) drugs, herbal products, or group weight-loss programs. Of the patients who do seek medical help, most have already exhausted other methods before approaching a physician. At the most basic level, the object of obesity treatment is weight loss, or the reduction of body mass. A sustained reduction of 5–10% in body weight can often produce clinically signiÞcant improvements in blood pressure, cholesterol, and blood glucose levels. The impact of these improvements on mortality remains unclear, but clinical trials do show that the risks of developing complications such as cardiovascular disease and diabetes can be reduced by weight loss. Patients often hope to lose up to 20–30% of their initial weight to achieve their “ideal” body weight for cosmetic reasons. Patient frustration with treatment approaches—as well as the dip in metabolism that occurs after weight loss, the natural propensity to gain weight with age, and the comfort that calorie-rich foods provide—all combine to make weight loss and weight maintenance an uphill struggle. Successful weight loss requires a comprehensive program of dieting, physical exercise, and behavioral therapy. In a growing number of cases, pharmacological agents are included as an adjunctive therapy. Increasingly, patients are opting for surgical intervention to achieve weight loss. Bariatric surgeries have been popularized by several overweight celebrities, but these surgical options are both expensive and potentially risky, as discussed later in this section. The addition of pharmacological agents to weight-loss programs has gained considerable attention from patients, physicians, pharmaceutical companies, and the media. Until recently, the only agents available were amphetamine derivatives, which can cause chemical dependence. In the mid 1990s, the obesity market

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TABLE 4. Complications of Obesity Complication Endocrine/metabolic Type 2 diabetes

Dyslipidemia

Gout Cardiovascular Coronary artery disease

Hypertension

Stroke Gastrointestinal Gallbladder disease

Musculoskeletal Osteoarthritis Bone malformation

Respiratory

Comments Visceral fat in particular is strongly associated with the development of type 2 diabetes because visceral deposits generate high levels of free fatty acids; these acids interfere with glycolysis (the handling of glucose by the liver) and insulin clearance. Overeating causes high blood glucose levels, and the pancreas responds to these elevated levels by increasing the production of insulin. Over time, cells become less receptive to the insulin secreted, and type 2 diabetes develops. Obese patients frequently have abnormal blood lipid levels. In these patients, high-density lipoprotein cholesterol (HDL-C) levels are typically low, whereas low-density lipoprotein cholesterol (LDL-C) is often normal or elevated. Obesity and associated hypertension can increase uric acid levels in serum due to decreased urinary uric acid clearance. Obesity has been shown to increase the risk of coronary artery disease independent of other factors such as dyslipidemia. Increased body weight causes increases in total body oxygen consumption, leading to an increase in cardiac output. Over time the atria and ventricles may begin to fail. Several studies have demonstrated a correlation between excess weight and increased blood pressure. Excess central and upper body weight leads to an increase in systemic vascular resistance, resulting in a sustained rise in blood pressure. Obese patients have an increased risk of ischemic (not hemorrhagic) stroke as a result of hypertension. Researchers have shown that bile saturation is greater in obese people where there is supersaturation of cholesterol in the gallbladder bile. Efforts to shed weight may aggravate the problem by increasing bile saturation and gallstone formation. Excess body weight increases stress on weight-bearing joints, contributing to the development of osteoarthritis. Obesity is a major risk factor for Blount’s disease, a growth defect observed in some obese children. Symptoms include a progressive bowing of the legs. A heavier chest increases strain on the lungs, and increased fat levels in the chest wall reduce lung volume. Morbidly obese patients suffer decreased arterial oxygenation because of poor ventilation-perfusion volume. Common morbidities include obstructive sleep apnea, asthma, chronic hypoxia, and shortness of breath.

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TABLE 4. (continued) Complication Cancer

Reproductive disorders

Psychological disorders

Comments Increased body weight is associated with increased risk of certain cancers including prostate, endometrial, postmenopausal breast, and colon cancer. The increased risk may be associated with the elevation of free estrogen caused by altered adipose tissue and suppression of sex-hormone-binding globulin in obese patients. Excess body fat may also delay detection of small tumors in the case of breast cancer. The mechanisms leading to the increased risk of colon cancer in obese patients are unclear. Intentional weight loss is associated with reduced cancer mortality. Obesity is associated with pituitary and gonadal dysfunction and thus with decreased fertility. Studies have also linked obesity to erectile dysfunction in men and polycystic ovary syndrome in women. Obesity is associated with psychological disorders such as depression and poor body image. These disorders may be caused by obese patients’ physical incapacities owing to shortness of breath as well as back and joint problems, and by difficulties with sleep, work, and social interactions.

was abuzz with the rapid uptake and subsequent withdrawal of the controversial drugs fenßuramine and dexfenßuramine (Wyeth-Ayerst’s Pondimin and Redux, respectively). Although newer drug options are currently available, no pharmacological “magic bullet” cure for obesity has yet been developed, and much remains to be learned about potential drug targets. Current choices for drug therapy include centrally acting appetite suppressants and peripherally acting agents that limit fat absorption in the GI tract (lipase inhibitors). Table 5 provides a list of currently available antiobesity agents. Only orlistat (Roche’s Xenical) and sibutramine (Abbott’s Meridia) have been approved for long-term (one year or longer) treatment of overweight and obesity. Table 6 compares the clinical expectations and most common side effects of each class of antiobesity drugs available on the market. Pancreatic Lipase Inhibitors Overview. Orlistat is the only pancreatic lipase inhibitor for treatment of obesity that is currently available in the major pharmaceutical markets. Unlike other marketed antiobesity agents, pancreatic lipase inhibitors act locally in the intestinal lumen to decrease the absorption of dietary fats. Although its side effects are unpleasant (e.g., oily spotting, bowel urgency, and diarrhea), this class of drugs is safer than the centrally acting weight-loss drugs. Mechanism of Action. Gastric and pancreatic lipases transform dietary triglycerides into free fatty acids and monoglycerides so that the small intestine can absorb them during digestion. Pancreatic lipase inhibitors inhibit gastric, pancreatic, and carboxyl ester lipases to decrease dietary free fatty acid absorption by

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TABLE 5. Current Therapies Used for Obesity Agent

Company/Brand

Pancreatic lipase inhibitors Orlistat Roche’s Xenical Serotonin and norepinephrine reuptake inhibitors Sibutramine Abbott’s Meridia, AstraZeneca’s Reductil Noradrenergic anorectic agents Phentermine GlaxoSmithKline’s Fastin, UCB’s Ionamin, generics Mazindol Wyeth-Ayerst’s Mazanor, Novartis’s Sanorex

Daily Dose

Availability

120 mg tid

US, F, G, I, S, UK

10–15 mg qd

US, F, G, I, S, UK

15–37.5 mg qd

US

1 mg qd

US, J

Notes: qd = Once daily; tid = Three times daily. US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan.

FIGURE 4. Mechanism of action of orlistat.

30%. The decreased absorption results in a concomitant increase in fat excretion. This class of drugs is believed to cause weight loss by reducing the amount of calories derived from ingested fat. As illustrated in Figure 4, these inhibitors act peripherally within the intestinal lumen rather than centrally in the brain. Orlistat. Orlistat (Roche’s Xenical) (Figure 5) has been available in Germany, the United Kingdom, and France since late 1998 and is now available in all major markets except Japan, where Chugai recently completed Phase II clinical trials. In late 2003, the FDA approved the use of orlistat for treatment of obesity

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TABLE 6. Comparison of Current Therapies for Obesity Compound Pancreatic lipase inhibitors

Serotonin and norepinephrine reuptake inhibitors

Noradrenergic anorectic agents

Advantages

Disadvantages

• Trials show that 5–10 kg or 5–10% body weight is lost over six months to one year. • Up to 30% of patients maintain their weight loss over two years of therapy. • Patients exhibit reduced serum lipid levels and improvements in risk factors for developing cardiovascular disease and diabetes.

• Gastrointestinal side effects include fecal incontinence, flatulence, and oily spotting. • Malabsorption of lipid-soluble vitamins can occur in some patients taking orlistat.

• Trials show that 3–6 kg or 5–10% of body weight is lost over six months to one year. • Up to 30% of patients maintain their weight loss over two years of therapy. • A study demonstrated that obese patients with concurrent binge-eating disorder exhibited improved control of eating behavior while taking sibutramine.

• Common side effects include elevations in blood pressure, increased heart rate, dry mouth, anorexia, insomnia, and nausea. • Sibutramine is contraindicated in patients taking antidepressants or who have uncontrolled hypertension.

• Trials demonstrate a weight loss of 0.5 kg/week or less than 5% of body weight lost over two months of therapy.

• Side effects include overstimulation of the CNS, restlessness, dizziness, insomnia, euphoria, dysphoria, tremor, and headache. • Drugs in this class are approved for short-term use only.

CNS = Central nervous system.

in adolescents (aged 12 to 16), making orlistat the Þrst and only prescription obesity drug approved for nonadult patients. Over-the-counter forms of orlistat are currently being marketed by Roche in Australia and New Zealand. In July 2004, GlaxoSmithKline licensed from Roche the right to market an OTC version of orlistat in the United States (GlaxoSmithKline, press release, July 19, 2004). The efÞcacy rates found in trials of orlistat are somewhat more consistent than those found in studies of another widely marketed antiobesity drug, sibutramine. Clinical trials for obesity drugs are typically designed to include a concomitant reduced-calorie diet. These hypocaloric diets generally allow 1,200-1,500 kcal/day. Therefore, even patients receiving placebo lose some percentage of their initial body weight. In a one-year study of orlistat in 391 obese men and

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FIGURE 5. Structure of orlistat.

women with type 2 diabetes, 49% of patients receiving orlistat lost more than 5% of their initial body weight, compared with 21% of those receiving placebo. Furthermore, 18% of orlistat-treated patients lost at least 10% of their body weight, compared with 9% of those receiving placebo (Hollander PA, 1998). In a different one-year study of 228 obese adults, 35% of patients treated with orlistat lost at least 5% of their initial body weight, compared with 21% of patients receiving placebo. Moreover, 28% of the drug-treated patients experienced at least 10% weight loss, compared with 17% of the placebo-controlled group (Finer N, 2000). In a two-year, randomized, double-blind, placebo-controlled study of 1,187 obese men and women, 66% of patients receiving orlistat lost 5% of their initial body weight, compared with 44% of patients receiving placebo (Davidson MH, 1999). In addition, 39% of the orlistat-treated patients lost 10% or more of their body weight, compared with 25% who received placebo. Patients receiving orlistat lost an average 9% of their body weight by the end of the Þrst year, compared with an average of 6% lost by the placebo group. However, the total amount of weight lost by patients in either group was relatively modest. Moreover, patients who continued taking the drug in the second year of the study regained an average 35% of the weight they had lost compared with placebo-treated patients, who regained an average of 63% of their lost weight. In addition to moderate weight loss, orlistat has been associated with a reduction in serum lipid levels. In a study of 444 obese subjects with hyperlipidemia, weight loss induced by treatment with orlistat in conjunction with a reducedcalorie diet resulted in statistically signiÞcant decreases in total cholesterol, low-density lipoprotein (LDL), and fasting insulin levels compared with patients who lost weight by dieting alone (Lucas CP, 2003). Results from several clinical trials demonstrate that patients exhibit, on average, a 10% reduction in total cholesterol and LDL levels, independent of weight loss (Hollander PA, 1998; Davidson MH, 1999; Rossner S, 2000; Finer N, 2000). Major clinical trials have consistently shown this level of improvement. In addition, several orlistat trials have shown that in obese patients with hypertension, blood pressure is lowered proportionally to weight loss (Davidson MH, 1999; Rossner S, 2000). In 2004, following completion of a large-scale study known as Xenical in the Prevention of Diabetes in Obese Subjects (XENDOS), the European Commission

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approved labeling for the reduction of risks associated with the development of type 2 diabetes. In this double-blind, placebo-controlled prospective study, 3,305 obese patients were followed over a four-year period. Patients receiving orlistat not only lost more weight compared with those receiving placebo (on average, 5.7 kg versus 3.0 kg), but also exhibited a 37% reduced incidence of type 2 diabetes over the course of the study (Torgerson JS, 2004). Other studies have shown that orlistat can reduce the severity of several diagnostic markers for type 2 diabetes in obese diabetic patients. P.A. Hollander and colleagues reported that patients taking orlistat were able to reduce their doses of antidiabetic drugs by 23%, compared with a 9% reduction in patients receiving placebo (Hollander PA, 1998). The negative effects of orlistat are mainly gastrointestinal. Because the net effect of orlistat is to render 30% of dietary fat non-absorbable, the fat travels through the gastrointestinal (GI) tract, where it causes gastrointestinal discomfort prior to being excreted. The most frequently noted adverse effects are minor and include oily spotting, ßatus with discharge, fecal urgency, and oily stools. These events can increase in severity if a patient taking the drug eats a high-fat meal because of the increased volume of fat that must pass through the GI tract. In addition, orlistat has been shown to limit the absorption of fat-soluble vitamins A, D, and E, which depend on pancreatic carboxyl ester lipase for their activation. These vitamins also rely on lipid binding for their absorption by the small intestine. Furthermore, several studies have shown that orlistat can inhibit bioavailability of fat-soluble drugs such as cyclosporine and amiodarone to subtherapeutic levels (Asberg A, 2003; Zhi J, 2002; Zhi J, 2003). Serotonin and Norepinephrine Reuptake Inhibitors Overview. In recent years, scientiÞc research has revealed the important role that serotonin and norepinephrine play in appetite control and food intake. Sibutramine (Abbott’s Meridia; AstraZeneca’s Reductil) is the only serotonin and norepinephrine reuptake inhibitor (SNRI) approved as an antiobesity agent. Other agents in this class are used as antidepressants and have not been approved for use as appetite suppressants. Mechanism of Action. SNRIs produce two metabolites (M1 and M2) that block reuptake of both serotonin and norepinephrine by presynaptic neurons (illustrated in Figure 6). In addition, some animal studies suggest that, because the drug causes an increase in sympathetic nervous system activity, it indirectly activates β3 -adrenergic receptors in the periphery, thereby increasing energy expenditure (Liu YL, 2002). Sibutramine has negligible effects on dopamine reuptake, so it does not have the potential to cause dependence, as amphetamines do. Appetite suppression can be achieved by pharmacological activation of the hypothalamic noradrenergic (or catecholaminergic) and serotonergic pathways. Catecholaminergic activation mimics the effects of norepinephrine to stimulate the sympathetic nervous system, which results in decreased appetite. Activation of

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FIGURE 6. Mechanism of action of sibutramine.

the serotonergic pathway suppresses appetite by stimulating neurons that promote a feeling of satiety. Unlike several other appetite suppressants that stimulate the release of extra serotonin and norepinephrine, sibutramine acts on both the noradrenergic and serotonergic pathways by inhibiting the reuptake of norepinephrine and serotonin in the hypothalamus. Sibutramine also increases thermogenesis, a process involving involuntary energy expenditure to produce heat. Sibutramine. Sibutramine (Abbott’s Meridia; AstraZeneca’s Reductil) (Figure 7) is an SNRI, the Þrst of its kind to be developed for obesity. Sibutramine received approval in the United States in 1997 and was launched in 1998. The agent has since been launched in all of the major European markets. The Italian Health Ministry temporarily withdrew sibutramine in March 2002 after 50 complaints of adverse events and two deaths were reported (Pharmaceutical Business News, March 20, 2002). At the request of the Italian Health Ministry, the European Commission’s Committee for Proprietary Medicinal Products (CPMP) conducted a risk/beneÞt assessment of sibutramine, after which the organization issued a positive opinion in June 2002 (Marketletter, July 8, 2002). Sibutramine is now available in Italy but can be prescribed only by hospital-based specialists. Eisai has licensed the right to market sibutramine in Japan and recently initiated Phase III studies there. Sibutramine is indicated for patients with a body mass index (BMI) greater than 30 or, if comorbidities are present, a BMI of at least 27. Sibutramine is indicated for a maximum duration of one year of therapy. Although sibutramine has a low potential for abuse, the drug is listed as a class IV controlled substance in the United States (along with the noradrenergic anorectic agents phentermine and mazindol).

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FIGURE 7. Structure of sibutramine.

In a study that included 1,001 patients who had achieved at least a 2% weight reduction during an open-label run-in period, patients were randomized to receive continuous sibutramine treatment for 48 weeks, intermittent treatment (consisting of 36 weeks of total treatment and 12 weeks of placebo), or placebo for 4 weeks. At the end of the study period, 65% of the continuously treated patients and 63% of the intermittently treated patients lost 5% or more of their body weight, compared with 35% in the placebo group. Furthermore, 32%, 33%, and 13% of the continuous, intermittent, and placebo groups, respectively, lost at least 10% of their body weight (Wirth A, 2001). The investigators did not report what, if any, dietary advice they provided to patients. One study of sibutramine included obese patients with hypertension. In this study, which included minimal dietary advice to patients, 224 patients were randomized to receive sibutramine or placebo for 52 weeks. At the end of the study period, 40% of the sibutramine-treated patients lost 5% or more of their body weight, compared with 9% of those receiving placebo. In addition, 13% of sibutramine patients lost 10% or more of their body weight, compared with 4% of those receiving placebo. However, patients receiving sibutramine had statistically relevant increases in both blood pressure (2.0 mm Hg) and pulse rate (4.9 beats per minute), compared with patients receiving placebo (Bray GA, 1999; McMahon FG, 2000). The Sibutramine Trial of Obesity Reduction and Maintenance (STORM) study was a randomized, double-blind, placebo-controlled study that examined the ability of sibutramine to sustain weight loss over 18 months in 467 obese patients who had already achieved a 5% weight loss through diet and exercise (James WPT, 2000). Of those who completed the trial, 69% of treated patients (n = 352) maintained at least a 5% weight loss, and 46% sustained at least a 10% weight loss, compared with 40% and 20% in the placebo group (n = 115), respectively. Sibutramine also works to control eating behavior in the subset of obese patients who have a psychological binge-eating disorder. In a double-blind, placebo-controlled study of 60 obese patients previously diagnosed with bingeeating disorder, researchers found that treatment with sibutramine resulted in reduced numbers of binge-eating episodes. Improvement in depression, which is commonly associated with binge-eating behavior, was also observed. In addition to the behavioral improvement, a majority of these patients were also able to achieve signiÞcant weight loss during the course of the 12-week study. Patients

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receiving sibutramine lost an average of 7.4 kg compared with a weight gain of 1.4 kg in patients receiving placebo (Appolinario JC, 2003). Researchers quantiÞed the number of cardiovascular complications and other side effects resulting from sibutramine treatment in a meta-analysis of 2,068 patients enrolled in various clinical trials (Weber MA, 1999). In a minority of patients, sibutramine therapy caused a signiÞcant increase in blood pressure and heart rate. Tachycardia was observed in 3% of treated patients and 1% of those in the placebo group, and an increase in blood pressure occurred in 2% of treated patients, compared with 1% of those who received placebo (Weber MA, 1999). In the STORM trial, 20 patients (3%) withdrew because of elevated blood pressure (James WPT, 2000). Finally, in a study of 31 obese hypertensives, only one patient treated with sibutramine withdrew from the study, although four patients required modiÞcations to their antihypertensive therapy (McMahon FG, 2000). Other side effects observed in sibutramine-treated patients include insomnia and nausea, which have been reported in 1–7% of patients (Apfelbaum M, 1999; Wirth A, 2001) and dry mouth, which occurs in up to 16% of patients on the drug (McMahon FG, 2000). Because of the risk of hypertension and tachycardia, sibutramine is contraindicated in patients with a history of coronary heart disease, cardiac arrhythmias, congestive heart failure, or stroke. According to sibutramine’s labeling, the drug is also contraindicated in patients taking monoamine oxidase inhibitors or selective serotonin reuptake inhibitors. Noradrenergic Anorectic Agents Overview. Most currently marketed noradrenergic anorectic agents are appetite suppressants. The Þrst noradrenergic anorectic agents, amphetamines (including phenmetrazine [Boehringer Ingelheim’s Preludin]), have signiÞcant abuse potential and are now only rarely used in weight management. Other catecholaminergic anorectics, including diethylpropion (Aventis’ Tenuate, also called amfepramone), phentermine (GlaxoSmithKline’s Fastin, UCB Pharma’s Ionamin, generics), and mazindol (Wyeth-Ayerst’s Mazanor; Novartis’s Sanorex), are amphetamine derivatives. These drugs do not cause as substantial an increase in sympathetic activity as does amphetamine, but they are also associated with abuse and have been connected to pulmonary hypertension and heart disease (Scheen AJ, 1999; Kordik CP, 1999; Abenhaim L, 1996). These drugs, where available, are approved only for short-term, intermittent treatment (i.e., up to 12 weeks) and not for chronic use. Common side effects for all drugs in this class include sleeplessness, nervousness, euphoria, tremor, and headache. In 2000, the European Commission attempted to ban the sale of all noradrenergic anorectic drugs (including diethylpropion, phentermine, and clobenzorex [Aventis’s Asenlix]) because of their lack of efÞcacy and the risk of arterial pulmonary hypertension and other side effects (Abenhaim L, 1996). In April 2000, after several companies fought this attempt, the European Union’s Court of First Instance (responsible for interpreting community law and actions brought against the European Commission) suspended the ban (owing to lack of evidence) until

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FIGURE 8. Structure of phentermine.

the main case was heard. On July 2003, the court conÞrmed the earlier ruling that overturned the ban of noradrenergic anorectic drugs by the European Commission. This section discusses phentermine and mazindol, which are the primary noradrenergic anorectic agents marketed in the United States and Japan, respectively. All other noradrenergic anorectic agents are either unavailable or only used minimally in the major pharmaceutical markets. Mechanism of Action. Noradrenergic drugs work by stimulating the release of norepinephrine from synaptic granules or by blocking its reuptake by presynaptic neurons. These mechanisms increase sympathetic activity, resulting in catecholamine release. Subsequent catecholaminergic activation stimulates the sympathetic nervous system, which results in appetite suppression. Increased levels of norepinephrine also stimulate glycogenolysis (conversion of glycogen to glucose) in the liver and lipolysis (conversion of triglycerides into fatty acids) in adipose tissue by activation of peripheral β3 -adrenergic receptors. The addictive properties of amphetamines and amphetamine derivatives stem from their ability to trigger dopamine release in addition to norepinephrine release. Sustained stimulation of the dopamine receptors results in their desensitization. Phentermine. Phentermine (GlaxoSmithKline’s Fastin, UCB Pharma’s Ionamin, generics) (Figure 8) launched in the United States in 1991 and is approved for short-term treatment of obesity. The drug stimulates the noradrenergic pathway, but it has less of an effect on dopamine than amphetamines do; therefore, it presents less abuse potential. In one crossover study, 66 obese diabetic patients were treated daily with 30 mg phentermine resin for six months (n = 34) or with placebo (n = 32). Patients receiving phentermine lost, on average, 5.3 kg of body weight compared with 1.5 kg lost by patients receiving placebo. A portion of the placebo group (n = 14) was then treated with phentermine. After six months, this group lost an average of 6.5 kg of body weight (Campbell C, 1977). Other placebo-controlled clinical trials of phentermine lasting less than six months have demonstrated similar amounts of weight loss (3 to 6 kg) (Truant AP, 1972; Langlois KJ, 1974; Weintraub M, 1984). Phentermine is highly stimulatory, so patients usually have difÞculty tolerating this agent as monotherapy. In the 1990s, physicians began prescribing a combination of fenßuramine and phentermine (fen-phen) because weight loss was greater with the combination, and the lethargy induced by fenßuramine balanced the stimulatory effects of phentermine. Fenßuramine acts as a highly selective serotonin agonist that triggers the release of serotonin and inhibits its reuptake.

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FIGURE 9. Structure of mazindol.

Dexfenßuramine is another form of fenßuramine. The combination of fenßuramine and phentermine exerted a slight synergistic effect, with some studies reporting as much as a 15–20% weight loss in patients taking the combination. As fen-phen and dexfenßuramine’s popularity surged, patients increasingly reported side effects such as neurotoxicity, primary pulmonary hypertension, and valvular heart disease. For these reasons, dexfenßuramine and fenßuramine were withdrawn from the world-wide market in 1997. Phentermine remained on the market because the safety concerns caused by the fen-phen combination were attributed solely to the serotonergic effects of fenßuramine and dexfenßuramine. Mazindol. Mazindol (Wyeth-Ayerst’s Mazanor, Novartis’s Sanorex) (Figure 9) is a noradrenergic compound available in the United States and Japan. This agent inhibits the reuptake of norepinephrine in the lateral hypothalamus but does not inhibit the uptake of dopamine. One clinical trial of mazindol in obese Japanese patients demonstrated that on average, patients experienced a moderate weight loss of 6.8 kg (15 lb) over 60 weeks of treatment. Patients were concurrently treated with a very low calorie diet (VLCD) and were able to maintain weight loss better than patients who were treated with VLCD alone. However, the dropout rate was greater than 50% owing to side effects (thirst, constipation, and fatigue) and patients’ perception of a lack of efÞcacy (Inoue S, 1995). Nonpharmacological Approaches Research shows that the sustained loss of a modest 5% of initial weight can improve comorbidity outcomes by reducing vascular inßammation and insulin resistance (Esposito K, 2003). However, achieving this amount of sustained weight loss through behavioral modiÞcation alone is very difÞcult. Behavioral modiÞcation programs operate on the assumption that obesity results from habitual overeating and the choice of nutritionally poor foods—qualitative lifestyle aspects that can be changed. Obese patients’ dietary histories often reveal a lack of meal structure and a tendency to snack. These habits are associated with the consumption of foods that are high in fats and sugars. Behavioral therapy can take place within a group setting or as part of an individual counseling program and is typically administered by physicians, psychologists, or successful former patients. Self-monitoring plays an important role in behavioral change:

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most programs involve keeping a daily log of food intake and exercise. Behavioral modiÞcation techniques aim to control food intake by reducing the number of external environmental cues or stimuli that lead to overeating. The principal means of achieving weight reduction is calorie restriction. The intake of fewer calories than are expended causes the body to burn off stored calories. Typically, a loss of 1 kg of body fat can be achieved by reducing food intake by 7,700 kcal over a speciÞed period of time. On average, a person needs to consume 30–35 kcal per kilogram of body weight per day for weight maintenance. The association between caloric intake and body weight is well known, and many obese people attempt to restrict calories on their own. Such attempts are often sporadic, and without lasting changes in lifestyle, they usually fail. Physician-prescribed diets range from mild caloric restriction for overweight patients to VLCDs for severely obese patients. The amount of energy consumed during physical activity constitutes only 10–20% of total daily expenditure. However, although exercise plays only a small role in weight loss, regular exercise is essential for weight maintenance. Although exercise can increase appetite, it provides a sense of well-being and can improve regulation of body fat composition, glucose tolerance, blood pressure, and lipid proÞles. Some researchers also believe that exercise can help to lower the theoretical “set point” of body weight to cause lasting beneÞcial changes in metabolism. Experts stress that even light exercise (e.g., brisk walking for 30 minutes), if done regularly, offers synergistic health advantages beyond simply increasing the amount of calories expended. Encouraging patient compliance with regular exercise is a major challenge for physicians. There are many obstacles to introducing a regular exercise regimen: the costs of gym memberships, intimidation, fear of exacerbating comorbid medical conditions, safety concerns (e.g., injury, triggering of acute coronary syndromes). Other well-known obstacles include the reluctance to change long-term habits, fear of failure, and time constraints. Invasive surgical intervention is considered by many to be an aggressive approach to treating obesity. However, the efÞcacy of surgical intervention and its potential for limiting weight regain far exceed those of diet, exercise, and behavior modiÞcation: patients can lose 30–40% of their original weight with surgery, compared with 5–10% with drug therapy. The combined effect of media coverage of dramatic weight loss achieved by celebrities and the promise of a “quick Þx” has led to a signiÞcant increase of these surgical procedures worldwide. Despite the substantial risks associated with bariatric surgeries, demand continues to soar. According to the American Society for Bariatric Surgery, an estimated 100,000 gastric bypass surgeries were performed in the United States alone in 2003. Surgical procedures used in the treatment of obesity fall into two categories: (1) restrictive procedures such as vertical banded gastroplasty (stomach stapling) and gastric banding, which promote weight loss by restricting food intake, and (2) gastric bypass procedures (e.g., Roux-en-Y gastric bypass), which cause malabsorption of food. All of these procedures are performed routinely in Europe,

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where they have been in use for many years. In the United States, although gastric banding was approved in 2001, gastric bypass and gastroplasty remain the more popular options. For the morbidly obese, success in terms of immediate weight loss makes surgery a potentially life-saving intervention. At present, surgery is recommended only to patients with a BMI > 40.0 who are strongly motivated to lose weight. In addition, surgery may be considered for patients with a BMI between 35.0 and 40.0 if they are at a high risk of life-threatening comorbid conditions such as cardiomyopathy, severe diabetes, or obstructive sleep apnea (also called Pickwickian syndrome, which is characterized by obesity, somnolence, and chronic hypoventilation). Surgery is rarely indicated for children and adolescents, and patients older than age 55 are believed to be unlikely to successfully adapt to the radical dietary changes associated with this form of treatment. The efÞcacy of surgery in terms of maintaining weight loss is much greater than for any form of drug or behavioral therapy. Surgical patients typically shed one-third of their preoperative weight (or 50–60% of their excess weight) and are able to maintain the loss longer (approximately 10–12 years after surgery) than pharmacologically treated patients. In addition, surgery has been shown to reduce the risks of many pathological conditions associated with obesity: fasting serum glucose and fasting insulin levels often drop; blood pressure decreases; respiratory problems are alleviated; and joint pain and sleep apnea resolve in many patients (Melissas J, 1998). Nevertheless, surgery has many disadvantages. Although gastric banding allows for a degree of adjustment, gastric bypass and gastroplasty procedures may necessitate corrective intervention in the event of dilation or loosening of staples. In addition, the bands may erode in the stomach. Moreover, all gastric surgery carries the attendant risk of bleeding, heart problems, lung problems, intestinal obstruction, and problems associated with general anesthesia. Death as a result of heart attack, cardiac arrhythmias, or, more commonly, peritonitis or pulmonary embolus occurs in approximately 1% of patients immediately following surgery. Other potential complications include leakage of gastric contents from the site of surgery, injury to the spleen, infections, hernias, and a signiÞcantly increased risk of developing gallstones. Subsequently, ulcers may form in the stomach or small intestine, or obstruction of the opening of the stoma may occur. More serious complications are pouch or esophageal dilation (stretching) and staple disruption, which render the intervention useless because the enhanced feeling of fullness disappears. Depending on the procedure performed, 10–50% of patients may require secondary surgical procedures to repair these complications (Podnos YD, 2003; Martikainen T, 2004). EMERGING THERAPIES Given the increasing patient population of overweight and obese individuals worldwide, vast opportunity exists in the market for weight-loss drugs. Very few

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TABLE 7. Emerging Therapies in Development for Obesity Compound

Development Phase

ATL-962 United States — Europe II Japan I GI-181771 United States — Europe II Japan — HMR-1426 United States — Europe II Japan — Centrally acting agents Recombinant human ciliary neurotrophic factor (Axokine) United States III Europe — Japan — Rimonabant (Acomplia)a United States III Europe III Japan I Bupropion SRa United States II Europe — Japan — Thermogenic agents L-796568 United States — Europe I Japan —

Marketing Company — Alizyme Takeda — GlaxoSmithKline — — Sanofi-Aventis —

Regeneron — — Sanofi-Aventis Sanofi-Aventis Sanofi-Aventis GlaxoSmithKline — —

— Merck —

a Rimonabant is also in Phase III development by Sanofi-Aventis for smoking cessation. b Bupropion SR is currently marketed for smoking cessation as Zyban and for depression as Wellbutrin.

patients are able to achieve signiÞcant weight loss for sustained periods using nonpharmacological means such as diet and exercise. This difÞculty, combined with the modest efÞcacies and signiÞcant side effects of currently available obesity drugs, has resulted in a great need for new pharmacological therapies that deliver improved weight loss with fewer side effects. In addition, the inability of patients to maintain the amount of weight lost using available therapies drives the demand for agents that can help sustain long-term weight control. Table 7 shows the principal agents in development for treating obesity. Peripherally Acting Agents Overview. Peripherally acting agents treat obesity locally; the most common site of action is the gastrointestinal (GI) tract. Therefore, these drugs have relatively favorable side-effect proÞles, giving them a signiÞcant clinical advantage

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over centrally acting drugs that affect the central nervous system (CNS). However, some experts suggest that the localized mechanisms of action that make these agents safer to use may be a trade-off that results in limited efÞcacy. Mechanism of Action. Peripherally acting agents induce weight loss by directly altering fat absorption in the GI tract or by modifying signals that affect food intake. Several agents in this drug class work by inhibiting digestive enzymes or by slowing gastric emptying. Other targets of peripherally acting agents are gastric hormones that are released in response to intestinal distension following meals, thereby triggering neural pathways that ultimately lead to a feeling of satiety. ATL-962. Alizyme’s ATL-962∗ is a synthetic oral lipase inhibitor in Phase II development in Europe for obesity and obesity-related complications such as type II diabetes. Alizyme announced in early 2004 that Takeda has entered a licensing agreement for the exclusive rights to develop and market ATL-962 in Japan (Alizyme, press release, January 4, 2004). ATL-962 works in a similar manner to orlistat, the only lipase inhibitor currently on the market. Lipase inhibitors block the digestion of fat by inhibiting gastric, carboxyl ester, and pancreatic lipases to reduce dietary fat absorption and calorie intake in the GI tract. A Phase IIb study of ATL-962 to treat obesity was recently completed in the United Kingdom and four other European countries. Trial results appear in a company press release. In this randomized, double-blind, placebo-controlled study, 372 obese patients were treated daily with 60 mg, 120 mg, or 240 mg ATL-962 over a three-month period (Alizyme, press release, September 19, 2003). Although no data were presented in the press release, the communication stated that patients in all three dosing groups lost a statistically signiÞcant amount of weight compared with patients taking placebo. In addition to weight loss, serum cholesterol was analyzed as a clinical end point in the Phase IIb study detailed in the press release. Patients given ATL-962 exhibited reduced low-density lipoprotein (LDL) and total cholesterol levels and no change in high-density lipoprotein (HDL) levels, whereas patients receiving placebo saw no change in their LDL levels and increases of both HDL and total cholesterol. The extent of the improvements to patients’ lipid proÞles was not stated in the press release. Side effects were primarily gastrointestinal and mild in severity. Although the average number of adverse events reported by drugtreated patients was nearly twice that of patients in the placebo group (3.8 per patient compared with 2.1), the company reports that the overall prevalence of GI side effects was up to 90% lower than reported for patients taking orlistat. Other early-stage clinical study results released by Alizyme indicate that the efÞcacy of ATL-962 is comparable to that of orlistat, but with an improved sideeffect proÞle even when high single doses of up to 900 mg of the drug are administered. Alizyme reports that a production process has been developed to help lower the cost of the drug and make it more affordable for consumers (Alizyme, press

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release, August 26, 1999). ATL-962 is poised to become a low-cost alternative to orlistat, which would give it a substantial competitive advantage based on current reimbursement policies in the major pharmaceutical markets. However, the reclassiÞcation of obesity as a disease by the U.S. Medicare administration is indicative of changes in pharmaceutical coverage for antiobesity therapies. Should these agents be routinely reimbursed in the future, ATL-962 will lose any market advantage based on its lower price. Furthermore, ATL-962 will likely face competition from generic versions of orlistat in the future. GI-181771. GlaxoSmithKline’s (GSK’s) GI-181771 is a cholecystokinin-A (CCK-A) receptor agonist that is currently in Phase II trials in Europe for treatment of obesity. CCK-A receptors are most abundant in peripheral tissues such as the pancreas and gallbladder but also exist in the brain. Activation of peripheral CCK-A receptors aids digestion at mealtimes by inducing gallbladder contractions and increasing the secretion of pancreatic enzymes. Other effects of stimulating CCK-A receptors include delaying gastric emptying and mediating gastric distension signals, both of which help to promote feelings of satiety. The biological responses following CCK-A receptor activation in peripheral tissues that lead to satiety signaling in the brain are still poorly understood. Preliminary results from a Phase II study of 254 obese patients taking 0.5 mg or 1.5 mg GI-181771 indicate that patients were able to achieve weight loss in a dose-dependent manner. The average amount of weight lost by each patient during the eight-week study was 2 kg. No other details of this study are available (GlaxoSmithKline R&D Day, December 3, 2003). In a Phase I trial reported in 2001, a single 8 mg oral dose of GI-181771 reduced food intake by 20% over a period of 24 hours (IDdb3, 2004). HMR-1426. SanoÞ-Aventis’s HMR-1426∗ , currently in Phase II development in Europe, is a compound that has been shown in rats to delay gastric emptying and limit food intake (Gossel M, 2001). Studies in rodents indicate that HMR-1426 does not act as a CCK-A receptor antagonist; the drug’s actual mechanism of action is unknown (Bickel M, 2004). Despite its unknown mechanism, SanoÞAventis reports some preliminary positive results of HMR-1426 in the treatment of human obesity. In a randomized, double-blind, placebo-controlled, Phase IIa study, 247 obese patients were treated daily with either 400 mg or 1,200 mg of HMR-1426 or placebo over a period of one month (Aventis, corporate presentation, June 18, 2002). No dietary restrictions were imposed on any of the three patient groups in the study. On average, treated patients lost 1.3% and 1.5% of initial body weight in the low- and high-dose groups, respectively, compared with 0.8% in the placebo group. The company reports that HMR-1426 was well tolerated. Centrally Acting Agents Overview. In the past, centrally acting appetite suppressants, such as the combination of fenßuramine and phentermine (“fen-phen”), have been the most

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efÞcacious weight-loss drugs. However, because this class of drugs acts on complex pathways involved in multiple neurological processes, use of centrally acting agents is accompanied by considerable side effects. Nevertheless, the pipeline for antiobesity drugs is currently dominated by centrally acting agents. The latestage pipeline includes agents that target novel mechanisms of action, such as recombinant ciliary neurotrophic factor (Regeneron’s Axokine) and rimonabant (SanoÞ-Aventis’ Acomplia), a cannabinoid receptor antagonist. On rare occasions, some physicians prescribe drugs originally developed for CNS disorders off-label to treat obesity. These agents include antidepressants such as bupropion (GSK’s Wellbutrin) and citalopram (Forest Lab’s Celexa) and the antiepileptic agent zonisamide (Elan’s Zonegran). Bupropion, a marketed antidepressant, is being investigated in Phase II clinical trials in the United States for use as an antiobesity agent. Drugs that target the neuropeptide Y (NPY) and melanocortin (MC-4) receptor pathways are also under development. Bristol-Myers Squibb, Esteve, and Lilly are developing NPY1 antagonists, while Novartis and Synaptic are codeveloping a nonpeptide NPY5 receptor antagonist (CGP-71683A). However, because of conßicting scientiÞc results in laboratory animals, the utility of these receptors as drug targets to treat human obesity remains unclear (Erikson JC, 1996; Heilig M, 1995; Inui A, 1999; Levens NR, 2003; Pedrazzini T, 1998; Schaffhauser AO, 1997). Several companies have initiated development programs for MC-4 receptor agonists, including GSK, Novo Nordisk, Neurocrine Biosciences, and Melacure. GSK’s MC-4 receptor agonist is in preclinical development with Chiron (Chiron, press release, January 8, 2003). Studies suggest that activation of MC-4 receptors triggers the release of anorexigenic signals. Mutations in the MC-4 receptor are commonly seen in obese people (Farooqi IS, 2000; Farooqi IS, 2003; Hinney A, 2003; Vaisse C, 2000). Mutations in the MC-4 receptor have also been linked to binge-eating disorder (Branson R, 2003), which is a primary cause of obesity in a subset of patients. Two other neuropeptide pathways that have gained considerable interest involve the gut hormone PYY, which signals satiety following food intake, and human growth hormone, which alters fat metabolism to induce lipolysis while also inhibiting the synthesis of fat. Intravenous administration of PYY has been shown to reduce appetite in both humans and mice (Batterham RL, 2002; Batterham RL, 2003). Both Nastech Pharmaceuticals and Amylin, in a licensing agreement with Curis, have analogues of PYY in early-stage clinical development. Metabolic Pharmaceuticals has an early-stage molecule in development derived from a portion of the human growth hormone (AOD-9604) that appears to mediate an effect on fat metabolism without affecting growth or triggering insulin resistance. Mechanism of Action. Although the neurophysiological processes of feeding behavior have yet to be fully established, scientists have been able to target several neuropeptides and receptors that regulate hunger and satiety (summarized in Figure 10). Centrally acting appetite suppressants, currently in development,

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FIGURE 10. Key neural and hormonal targets of emerging therapies.

target these CNS pathways by blocking or enhancing key mediators or downstream effecters of hunger and satiety signaling. Two of the most promising pathways being targeted include the endocannabinoid and noradrenergic systems. Recombinant Human Ciliary Neurotrophic Factor. Regeneron’s Axokine∗ is a genetically engineered version of human ciliary neurotrophic factor (CNTF)

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that exhibits enhanced stability compared with the naturally occurring protein. Originally developed for amyotrophic lateral sclerosis (ALS), Axokine was repositioned as an antiobesity agent when it unexpectedly caused signiÞcant weight loss in ALS patients (ALS CNTF Treatment Study Group, 1996). In January 2003, the U.S. Food and Drug Administration (FDA) granted fast-track designation to Axokine for treatment of severe obesity in patients who are unresponsive to currently available antiobesity therapies (Regeneron, press release, January 7, 2003). Initial results of Phase III trials conducted in the United States were released in January 2003 (Regeneron, press release, January 7, 2003). A pegylated version of Axokine is in Phase I development with Nektar Therapeutics, and an oral formulation is in preclinical investigation in collaboration with Emisphere Technologies. CNTF is a naturally occurring protein that functions as an important regulator of neuronal cell differentiation and survival. Although the mechanism by which injections of CNTF lead to weight loss in humans and animals is largely unknown, the protein is believed to work similarly to the hormone leptin, which is released by adipocytes to suppress appetite signaling in the brain. Axokine is administered by injection. The drug travels through the bloodstream and crosses the bloodbrain barrier, ultimately reaching the arcuate nucleus of the hypothalamus. Within the hypothalamus, Axokine binds to neuronal CNTF receptors to trigger satiety centers in the brain (Gloaguen I, 1997). Studies in leptin-deÞcient ob/ob mice and leptin-resistant, diet-induced obese mice showed that CNTF suppresses food intake without increasing NPY levels (which typically occurs during periods of starvation) and without causing cachexia (muscle wasting) (Lambert PD, 2001). Consequently, CNTF-treated mice did not exhibit binge-eating behavior or immediate rebound weight gain, which was observed in untreated mice. Other evidence suggests that the ability of CNTF to induce weight loss is not solely mediated by the CNS. One study showed that CNTF affects adipocyte signaling in vivo in rodents and induces insulin action in vitro (Zvonic S, 2003). Regeneron reported one-year preliminary results of a pivotal double-blind, placebo-controlled Phase III trial in a company press release (March 31, 2003). In this study, investigators enrolled subjects with no obesity-related risk factors and body mass index (BMI) ratios between 30.0 and 55.0 and subjects with obesity-related risk factors (e.g., high blood pressure, high blood lipids) and BMI ratios between 27.0 and 55.0. A total of 1,968 patients received daily subcutaneous injections of placebo (n = 501) or Axokine (n = 1, 467) at a dose of 1 mcg/kg body weight. Early results of the overall treated population were disappointing because weight loss was limited by the development of neutralizing antibodies that arose in 70% of treated patients after three months of therapy. However, outcomes in patients who did not develop antibodies were favorable: this subgroup exhibited a 5.7 kg reduction compared with 2.0 kg in the placebotreated group. In addition, 46% of these patients experienced a 5% body weight reduction compared with 24% of patients in the placebo group. Furthermore, 24% of patients achieved a 10% reduction in body weight, whereas this milestone was

EMERGING THERAPIES

219

FIGURE 11. Structure of rimonabant.

reached by only 7% of patients receiving placebo. Axokine was generally well tolerated; adverse events (including injection site reactions, nausea, and cough) were characterized as mild to moderate. Preliminary results from a Phase I study of a pegylated form of Axokine show that the drug can persist in the bloodstream long enough that injections are needed only once per week. In comparison, the regular formulation of Axokine has a half-life of two to six hours and must be administered daily. The company reports that the pegylated form of Axokine was poorly absorbed into the blood stream following intramuscular administration and caused signiÞcant injection site reactions. Regeneron plans to improve the formulation for pegylated Axokine before continuing further clinical studies of the drug (Regeneron, Form 10-K, December 31, 2002). The company reports that patients treated with Axokine for three months were able to maintain their weight loss over a 36-week period following cessation of therapy (Regeneron, press release, April 9, 2002). Rimonabant. Rimonabant (SanoÞ-Aventis’s Acomplia) (Figure 11) is a selective cannabinoid CB1 receptor antagonist being developed for obesity and smoking cessation. SanoÞ-Aventis has initiated several Phase III clinical trials for the use of rimonabant in treating obesity. RIO-North America (Rimonabant In Obesity) and RIO-Europe are two-year studies enrolling 2,800 and 1,500 patients, respectively. RIO-Lipids and RIO-Diabetes are one-year studies to investigate the use of rimonabant in 1,000 obese patients who have comorbid disorders such as dyslipidemia or insulin resistance. Rimonabant is in Phase I development in Japan. Endocannabinoids are produced naturally by the body and are believed to stimulate appetite and regulate adipogenesis. However, the mechanisms by which cannabinoids modulate food intake have yet to be fully elucidated. High levels of the CB1 receptor are present in the hypothalamus, where they control the release

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of neurotransmitters such as norepinephrine and serotonin (Cota D, 2003). These neurotransmitters are known to mediate satiety and hunger signals in the brain. CB1 receptors are also located on adipocytes in the periphery, where they regulate adipogenesis. Thus, rimonabant likely has both central and peripheral modes of action. SanoÞ-Aventis announced one-year results from the midpoint of the RIOEurope study at the 2004 Congress of the European Society of Cardiology. The RIO-Europe study is a two-year, multicenter, randomized, double-blind, placebocontrolled trial involving 1,507 obese patients at 60 different medical centers across Europe (Belgium, Finland, France, Germany, the Netherlands, and Sweden). All patients were placed on placebo and a hypocaloric diet (600 kcal/day deÞcit) during a one-month run-in period before being randomized to one of three parallel treatment arms (placebo, 5 mg/day rimonabant, or 20 mg/day rimonabant). Patients receiving 20 mg/day rimonabant lost an average of 8.6 kg of body weight after one year, compared with 4.8 kg lost by patients on the 5 mg/day dose and 3.6 kg lost by patients on placebo. Nearly 70% of patients on the 20 mg/day dose lost at least 5% of their initial body weight, compared with 44% of patients receiving 5 mg/day rimonabant and 31% of those receiving placebo. Furthermore, 39% of patients receiving the highest dose of rimonabant lost at least 10% of their initial body weight, compared with 15% of patients receiving the 5 mg/day dose and 12% of patients receiving placebo. The RIO-Europe study also investigated the effects of rimonabant on some of the comorbidities associated with obesity. Patients receiving the 20 mg/day dose of rimonabant demonstrated an average 3.5-inch reduction in waist circumference (a marker for abdominal fat volume), compared with a reduction of 2.0 inches by patients on the 5 mg/day dose and 1.5 inches in patients receiving placebo. Patients treated with rimonabant also showed improvements in lipid proÞles. In particular, patients receiving the 20 mg/day dose of rimonabant had a 27% average increase in HDL cholesterol, an average 11% reduction in triglycerides, and improved insulin sensitivity. Meanwhile, patients receiving placebo or the 5 mg/day dose of rimonabant exhibited smaller effects on their lipid proÞle and modest increases of 5% and 7%, respectively, in their triglyceride levels. Rimonabant was well-tolerated by most patients in the RIO-Europe study. The most common side effects included nausea (13% of patients receiving 20 mg/day rimonabant; 4% of patients on placebo), diarrhea (7% of the 20 mg/day group; 3% of the placebo group), and dizziness (9% of the 20 mg/day group; 5% of those receiving placebo). The company reports that only a small percentage of patients dropped out of the study because of side effects. Dropout rates were similar in all three treatment groups (39% of patients receiving 20 mg/day rimonabant, 37% of the 5 mg/day group, and 42% of patients on placebo). No differences in the frequency of depressive episodes or anxiety were observed in any of the three patient groups. Moreover, patients taking the highest dose of rimonabant did not exhibit increased blood pressure or heart rate compared with patients receiving placebo.

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Results from the one-year RIO-Lipids study were presented in the Þrst quarter of 2004 (SanoÞ-Synth´elabo, company meeting, February 16, 2004, and press release, March 9, 2004) and closely mirror the one-year results of the RIO-Europe study. In this double-blind, placebo-controlled study, 1,036 obese patients who had dyslipidemia were randomized to receive daily treatments of 5 mg or 20 mg rimonabant or placebo. Patients were placed on a reduced-calorie diet for the trial period. After one year, patients treated with the highest dose of rimonabant lost an average of 8.6 kg body weight, whereas those receiving placebo lost only 2.3 kg. Nearly 75% of patients receiving the 20 mg dose of rimonabant lost at least 5% of their body weight compared with 42% of patients on the 5 mg dose and 28% of patients receiving placebo. Furthermore, 44% of patients receiving the highest dose lost more than 10% of their initial body weight, compared with 16% of patients in the low-dose group and 10% of patients receiving placebo. The RIO-Lipids study was also designed to analyze the effects of rimonabant on several comorbidities associated with obesity. Compared with those given placebo, patients who received the 20 mg dose of rimonabant exhibited improvements in many symptoms of metabolic syndrome. On average, patients who completed one year of treatment saw a 3.4-inch reduction in waist circumference, a 23% increase in HDL-cholesterol levels, a 15% reduction in triglycerides, a 27% decrease of C reactive protein (a marker for systemic inßammation), and improved insulin resistance (as measured by oral glucose tolerance testing). The most common side effects associated with rimonabant were nausea and dizziness, which occurred more often in patients receiving the highest dose of the drug. Although the company reports that side effects were primarily mild and transient, nearly 15% of the patients receiving the 20 mg dose of rimonabant—compared with 7% of those on placebo—dropped out of the study because of side effects. The company reports no differences in the frequency of depressive episodes or anxiety in any of the three patient groups. Moreover, patients taking the highest dose of rimonabant did not exhibit increased blood pressure or heart rate compared with patients receiving placebo. The weight loss observed in clinical trials suggests that rimonabant has efÞcacy slightly better than that of orlistat and sibutramine. Furthermore, none of the serious side effects that have plagued sibutramine have been reported in any of the available trial results. Because rimonabant is also being developed for smoking cessation, it will be compared with bupropion (GSK’s Zyban and Wellbutrin), an antidepressant and smoking-cessation drug that is prescribed by some physicians for off-label treatment of obesity and is currently under Phase II investigation for obesity by GSK. Bupropion SR. GSK is investigating sustained-release bupropion (SR) (Figure 12), which the company currently markets for smoking cessation as Zyban and for depression as Wellbutrin, as an antiobesity therapy. Phase II trials for this indication are under way in the United States. Bupropion’s mechanism of action is not well understood. Some researchers believe that bupropion slows the destruction of norepinephrine, dopamine, and serotonin, which results in a greater concentration of these neurotransmitters in

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CH3 CH3 HN

CH3

O CH3

Cl FIGURE 12. Structure of bupropion.

the cerebrospinal ßuid. The elevated levels of these neurotransmitters lead to greater stimulation of receptors, thereby helping people to feel greater satiety after eating smaller portions. In a double-blind, placebo-controlled trial, 327 obese adults were randomized to a daily dose of 300 mg or 400 mg of bupropion SR or placebo over 24 weeks (Anderson JW, 2002). Patients in the 300 mg and 400 mg treatment arms lost an average of 7% and 10% of initial body weight, respectively, compared with 5% in the placebo arm. SpeciÞcally, 59% of patients in the 300 mg dose group experienced a 5% weight loss and 33% experienced a 10% weight loss, compared with 83% and 46% of patients, respectively, in the 400 mg dosing group, and 46% and 20% of patients, respectively, in the placebo group. At the annual meeting of the North American Association for the Study of Obesity (NAASO) in Quebec in October 2000, results from an open-label, 44-week study were presented. Of the 423 patients enrolled, 31% of bupropion-treated participants lost more than 5% of their body weight, compared with 21% of placebo-treated patients. However, 11% of bupropion-treated patients experienced weight gain, compared with 8% in the placebo-treated group. In an eight-week, double-blind, placebo-controlled study of 50 obese women, subjects receiving bupropion experienced greater weight loss at eight weeks compared with those receiving placebo (Gadde KM, 2001). Women in the bupropion group achieved a mean weight loss of 5%, compared with 1.3% for the placebo group. Thermogenic Agents Overview. Thermogenic agents are a novel class of drugs that promote weight loss through an increase in energy expenditure via thermogenesis, a process involving heat production or involuntary energy expenditure. Numerous agents in this drug class have dropped out during the course of development in the past several years. Dainippon and Takeda’s TAK-677 is the latest thermogenic agent to be discontinued because of lack of efÞcacy (Dainippon, press release, September 25, 2003). The departure of TAK-677 from the antiobesity pipeline leaves Merck’s L-796568, which is currently in Phase I development, as the most advanced member of this drug class.

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Mechanism of Action. Some thermogenic agents activate mitochondrial uncoupling proteins (UCPs; i.e., UCP-1, UCP-2, and UCP-3), which are located in different tissues. UCP-2 is ubiquitously expressed; UCP-1 and UCP-3 are expressed in brown adipose tissue and in skeletal muscle, respectively. UCPs provide alternate paths for protons to cross the mitochondrial matrix. Because the energy involved in this process is not coupled to ATP synthesis, the energy is dissipated as heat and contributes to energy expenditure. Other thermogenic agents target β3 -adrenergic receptors located in adipose tissue, which are involved in lipolysis, thermogenesis, and gastrointestinal function. In rodent models, these receptors mediate catecholamine-stimulated lipolysis in white and brown adipose tissue and thermogenesis in brown adipose tissue. Both processes involve lipid oxidation to produce heat and eliminate excess fat (Weyer C, 1999). β3 -adrenergic receptors also activate UCP-1 to increase energy expenditure. Unlike rodents, however, humans have only a small amount of brown tissue, because most of it is lost during the transition from infancy to adulthood. Moreover, the biology and function of β3 -adrenergic receptors have not yet been clearly established in humans. L-796568. Merck’s L-796568 is a selective β3 -adrenergic receptor agonist in development as an antiobesity agent. In a randomized, placebo-controlled, crossover study of 12 obese and overweight male patients, the effects of single doses of 250 mg or 1,000 mg L-796568 were evaluated during a four-hour period. Statistically signiÞcant increases in energy expenditure (calculated by indirect calorimetry) and plasma glycerol and free fatty acid levels were seen in patients who received the highest dose of the drug. However, these increases were also accompanied by an increase in systolic blood pressure (van Baak MH, 2002). The effect of 28 days of continuous daily treatment with 375 mg L-796568 on 20 overweight and obese but otherwise healthy males was evaluated in a double-blind, randomized, parallel-group, placebo-controlled study. No clinically signiÞcant differences in lipolytic or thermogenic effects were observed between the drug-treated and placebo groups (Larsen TM, 2002). REFERENCES Abenhaim L, et al. Appetite-suppressant drugs and the risk of primary pulmonary hypertension. New England Journal of Medicine. 1996;335(9):609–616. Allison DB. Annual deaths attributable to obesity in the United States. Journal of the American Medical Association. 1999;282:1530. Alvarez-Torices JC, et al. Self-reported height and weight and prevalence of obesity. Study in a Spanish population. International Journal of Obesity and Related Metabolic Disorders. 1993;17:663–667. Amyotrophic Lateral Sclerosis (ALS) CNTF Study Group. 1996. Neurology. 46: 1244–1249. Anderson JW, et al. Bupropion SR enhances weight loss: a 48-week double-blind, placebocontrolled trial. Obesity Research. 2002;10:633–641.

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Osteoporosis

ETIOLOGY AND PATHOPHYSIOLOGY Introduction Osteoporosis and osteopenia are diseases of the bone that are characterized by low bone mineral density (BMD), which increases the risk of fragility and fracture. The World Health Organization (WHO) deÞnes osteoporosis in Caucasian women as a BMD score greater than 2.5 standard deviations (SD) below the average BMD of young, healthy adults (T-score < −2.5); the same standard is used to deÞne osteoporosis in men and in other ethnic groups because data for these groups are unavailable. In practice, physicians also view fractures of the hip or vertebrae as evidence of osteoporosis, regardless of BMD measurements. Osteopenia is deÞned by T-scores that range from −1 to −2.5 in patients who have no history of hip or vertebral fracture. Typically, patients develop osteopenia before developing osteoporosis, but because BMD screening is not commonly performed for osteopenia, they are rarely diagnosed before the disease has progressed to osteoporosis. Osteoporosis can be classiÞed as primary or secondary depending on its cause. Primary osteoporosis in women typically arises following the decrease in levels of circulating estrogen that accompanies menopause. The decrease in estrogen levels increases the rate of natural bone resorption—without any change in the rate of new bone formation, the result is a net loss in bone mass and a decline in bone quality during midlife. Primary osteoporosis also occurs as a process of aging. Senile osteoporosis can occur in both sexes after age 70, and it results in decreased bone formation without changes in the rate of bone resorption, thus reducing bone mass and lowering BMD. Secondary osteoporosis most typically occurs Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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following long-term corticosteroid use, but it can also arise as a consequence of other diseases or pharmacological therapies. Anatomy The skeleton is an important reservoir for calcium that can be rapidly transferred to and from the circulation. Bone tissues can be divided into two main types: cortical and cancellous. Cortical bone (otherwise known as compact bone) is extremely dense and forms a hard shell surrounding the shafts of long bones (femur, tibia, Þbula, radius, and ulna). In an adult, 80% of skeletal mass consists of cortical bone. In contrast, cancellous (or trabecular) bone, which makes up 20% of the skeleton, is a thin, interconnected latticework of trabeculae that forms an internal supportive scaffold within the pelvis and other ßat bones, the vertebrae, and the ends of long bones. Cancellous bone has a vastly larger surface area than cortical bone. An estimated 85% of cancellous bone surface is exposed to bone marrow and bone-regulating factors. As a result, cancellous bone is more vulnerable than cortical bone to the metabolic changes (e.g., estrogen deÞciency) that reduce BMD. Therefore, highly cancellous bones (mainly the hip and vertebrae) are more susceptible to fracture in patients with osteoporosis. The organic matrix of bone comprises 95% type I collagen; other, noncollagenous proteins (e.g., osteocalcin, vitronectin) make up the remainder. Calcium salts (mostly in the form of calcium phosphate crystals [i.e., hydroxyapatite]) constitute the inorganic matrix, although other minerals and salts (magnesium, carbonate, and ßuoride) are also present. The deposition of hydroxyapatite crystals onto collagen Þbrils in the form of a Þshbone pattern is an important process, known as mineralization, that is required to increase BMD and that determines the ultimate strength and structural integrity of the skeleton. Decreased BMD, which results from excessive bone resorption and inadequate bone formation, is a major feature of osteoporosis. Measuring BMD is a well-established means of predicting fracture in osteoporosis. Etiology Normal bone remodeling depends on continuous and balanced cycles of bone resorption and bone formation. During the Þrst three decades of life, this process results in a net gain in skeletal bone mass. After age 30, however, bone mass in both men and women begins to decline. As this process continues, the bones may become brittle, porous, and vulnerable to fracture. Osteoporosis, as this condition is known, may have a primary or secondary etiology. Primary Osteoporosis. Postmenopausal osteoporosis, caused by declining estrogen production, is the most common form of primary osteoporosis. Thirty percent of women develop postmenopausal osteoporosis, in which the initial gradual decline in bone mass of 0.5–1.0% per year (observed in both men and women during the fourth decade of life) signiÞcantly accelerates and results in a net annual bone loss of 3–5% for Þve to ten years before slowing.

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The increased rate of bone resorption following menopause greatly exceeds the rate of bone formation, resulting in a higher rate of bone remodeling and decreased BMD. This imbalance is characterized by an increase in the number of active remodeling sites on bone (bone multicellular units, discussed in detail in the upcoming section, “Pathophysiology”) and an extended period of resorption at these sites. The result is decreased bone mass: primary loss of cancellous bone and disruption of cancellous bone microarchitecture. This condition increases susceptibility to fractures at sites in the skeleton that have proportionally more cancellous bone than cortical bone. These sites include the vertebrae (which are subject to crush fractures involving the lower thoracic or lumbar spine), the wrist, and, less frequently, the hip. Fractures may occur spontaneously or result from minimal trauma; they may also be “silent” (asymptomatic) or painful. Age-related osteoporosis (also called senile osteoporosis) affects both men and women aged 70 years or older; it is characterized by the slow, steady loss of bone, primarily cortical bone but also cancellous bone. In contrast to the increased rate of remodeling associated with postmenopausal osteoporosis, agerelated osteoporosis results from continual decline in osteoblast recruitment to bone and, consequently, diminished bone formation. Bone resorption also accelerates because of increased secretion of parathyroid hormone (PTH) in response to hypocalcemia, which results from poor calcium absorption and poor dietary intake of calcium and vitamin D. For women, the effects of age-related osteoporosis typically exacerbate the bone deÞciencies caused by postmenopausal osteoporosis, amounting to a total lifetime reduction of 30–60% of peak bone mass. Men may lose 20–25% of peak bone mass over the course of their lifetimes. Fractures of the proximal humerus (upper arm), proximal tibia (lower leg), ribs, and femoral neck (hip), as well as wedge-type vertebral fractures, predominate in age-related osteoporosis and are generally caused by minimal trauma. Secondary Osteoporosis. As Table 1 outlines, secondary osteoporosis may develop as a consequence of different diseases that predispose the patient to increased bone resorption and fracture risk. Patients with inßammatory diseases for which long-term glucocorticoids are prescribed are notoriously susceptible to secondary osteoporosis owing to the metabolic side effects of these drugs (Table 2). For instance, a retrospective cohort study of more than 244,000 patients in the United Kingdom demonstrated that long-term oral glucocorticoid therapy (e.g., for treating severe asthma and other autoimmune disorders) may be responsible for as much as 47% of all hip fractures and 72% of vertebral fractures (van Staa TP, 2001). In that study, elderly patients taking the highest doses of oral corticosteroids (> 7.5 mg/day) were at greater risk of fracture than untreated patients of all ages. Risk Factors. Table 3 lists risk factors for developing osteoporosis. Of these, premature menopause and hormone deÞciency are the best-recognized predictors for development of osteoporosis later in life. Another well-recognized predictor is long-term glucocorticoid therapy (≥ 7.5 mg/day prednisolone or equivalent);

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TABLE 1. Causes of Secondary Osteoporosis Predisposing Condition

Diseases/Conditions

Cancers

Leukemia Lymphoma Mastocytosis Multiple myeloma

Endocrine disorders

Diabetes mellitus Hypercalciuria Hypercortisolism (Cushing’s disease) Hyperparathyroidism Hyperprolactinemia Hyperthyroidism Hypogonadism

Genetic disorders

Homocysteinuria Osteogenesis imperfecta

Inflammatory diseases

Chronic liver disease Chronic renal disease Inflammatory bowel diseases Marfan’s syndrome Rheumatoid arthritis Transplantation (lung, heart, liver, kidney)

Malnutrition/malabsorption syndromes

Hypocalcemia Low protein intake

Pharmacotherapy

Antiepileptic drugs Cyclosporin A Glucocorticoids Heparin Thyroxin Iatrogenic

patients receiving this therapy are known to be at increased risk of developing osteoporosis. People who have suffered a fracture have twice the chance of developing an additional, osteoporosis-related fracture, regardless of their BMDs. For example, women who suffer a vertebral fracture have a Þvefold higher risk of developing another vertebral fracture within one year. Often, unfortunately, these people are not diagnosed, either because the fracture is asymptomatic or because orthopedists fail to refer them to other specialists for osteoporosis management. Excessive alcohol consumption, cigarette smoking, and inadequate physical activity are also considered major risk factors associated with reduced bone mass and increased fracture incidence. A sedentary lifestyle that includes too little weight-bearing exercise can lead to skeletal resorption and fragility. Similarly, people of small build and low body weight are more likely to be at risk of developing osteoporosis because their skeletons bear less weight. InsufÞcient intake of dietary calcium over prolonged periods may also be an important risk factor—especially during the Þrst three decades of bone development, when attaining a high peak BMD

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TABLE 2. Mechanisms of Corticosteroid-Induced Secondary Osteoporosis Target Tissue/Cell

Activity

Effects on Bone

Intestine

Reduces calcium absorption.

Osteoblast

Increases apoptosis, decreases maturation and activity, decreases bone matrix formation (e.g., collagen). Increases osteoclast recruitment and bone matrix resorption. Decreases estrogen production.

Osteoclast Ovaries Parathyroid gland

Induces secondary hyperparathyroidism.

Decreased availability of calcium and phosphate ions for mineralization; reduced bone density. Reduced bone density, increased bone porosity, fragility, and fracture risk. Same effect as for osteoblast. Decreased beneficial effects of estrogen on bone formation. Increased parathyroid hormone (PTH)-induced bone resorption.

TABLE 3. Risk Factors for Developing Osteoporosis Category Age Dietary Diseases Familial Hormonal

Iatrogenic Lifestyle

Race, gender Miscellaneous

Specific Risk Factors Advanced age (≥65 years) Chronic low calcium intake Chronic inflammatory disease (e.g., rheumatoid arthritis) Malnutrition due to anorexia nervosa, bulimia Expression of COLIA1 and COLIA2 genes Family history of osteoporosis Early menopause Primary hypogonadism Secondary amenorrhea Vitamin D deficiency Chronic high-dose glucocorticoid therapy Chronic thyroxin treatment Excessive alcohol or caffeine consumption Excessive cigarette smoking Sedentary Caucasian (Northern European origin), Asian Female gender Age- or health-imposed chronic immobility Previous fracture Small frame or low body weight

is crucial to preventing osteoporosis later in life. Calcium supplementation is therefore still prescribed for preventing or slowing age-related bone loss. Patients with chronic inßammatory disease (e.g., rheumatoid arthritis) are also at greater risk of developing osteoporosis. Although the predisposing factors are poorly understood, clinical research suggests that the prolonged release of inßammatory cytokines may be responsible for increasing bone resorption. Certain patient groups—especially the elderly and people with cognitive, neuromuscular, or visual impairments—are at serious risk for falls. If these people

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also have low BMDs (osteopenia) or osteoporosis, they are likely to sustain hip fractures. Pathophysiology Bone Remodeling and Turnover. The human skeleton undergoes a constant process of remodeling, requiring cycles of excavation (bone resorption) and reconstruction (bone formation) within a deÞned number of discrete pockets of bone that are called bone multicellular units (BMUs). Any increase in the number of BMUs translates to an increased rate of “bone turnover”—the overall increase in the total volume of bone being remodeled at any given time. The combined activity of multiple BMUs accounts for an annual replacement of 3% of cortical bone and 25% of cancellous bone in adults. This process is accomplished by a complex interaction between functionally different cells (bone-resorbing osteoclasts and bone-forming osteoblasts) under the systemic and local control of hormones, cytokines, and growth factors; the following section describes their molecular mechanisms. In postmenopausal osteoporosis, the osteoclast-induced bone resorption exceeds osteoblast-induced bone formation, resulting in enhanced bone remodeling and bone turnover and a net loss of bone mass. This dramatically accelerated bone resorption is believed to be mediated by a two- to threefold increase in the number of osteoclasts in osteoporotic bone, compared with normal bone. During the Þrst three decades of life, bone remodeling serves several important functions: repairing microfractures resulting from normal use and stress, increasing bone mass, and maintaining high-quality bone tissue. Prior to age 30, bone formation compensates for bone resorption. The importance of this physiological process is underscored by the fact that the disruption of bone remodeling experimentally, or in disease, has been shown to decrease bone mass and to increase bone fragility. Between the ages of 30 and 40, both men and women begin an age-related process of gradual bone loss. From the fourth decade onward, the rate of bone formation fails to keep pace with the rate of bone resorption, resulting in a progressive net bone loss that can lead to osteoporosis. Osteoporosis is characterized by increased bone remodeling, which is associated with a signiÞcant increase in the rates of bone resorption and formation (increased bone turnover). Because bone formation is unable to keep pace with bone resorption at a given site of excavation, a net deÞcit in bone mass develops. Mechanisms of Bone Remodeling. The generation of mature osteoblasts and the process of bone remodeling require tightly coupled interactions between osteoblasts and osteoclasts. Studies have shown that two osteoblast-derived factors—macrophage colony-stimulating factor (M-CSF) and the receptor for activation of nuclear factor κB ligand (RANKL)—are sufÞcient for osteoclast differentiation in vitro (Teitelbaum SL, 2000). M-CSF stimulates the proliferation of the osteoclasts. The binding of RANKL expressed on osteoblasts by the corresponding receptor (RANK) on pre-osteoclasts induces osteoclast differentiation. Osteoprotegerin, a soluble decoy receptor that negatively regulates osteoclast activity, inhibits this interaction by binding and sequestering RANKL.

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FIGURE 1. Bone remodeling.

As Figure 1 shows, multinucleated osteoclasts adhere to bone matrix. This process is mediated by the osteoclast cell membrane αv β3 integrin receptor binding to tissue vitronectin; inhibiting this process has been shown to reduce bone resorption. Multiple stimuli induce osteoclasts to resorb bone during the Þrst two to three weeks of a remodeling cycle. The activation of an adenosine triphosphatase (ATPase) proton pump at the osteoclast-bone surface serves to acidify and dissolve bone mineral, leading to pericellular demineralization. In addition, osteoclasts secrete different classes of enzymes (cathepsin K and matrix metalloproteinases [MMPs]) that digest and remove the surrounding organic bone matrix. When resorption is complete (in two to three weeks), osteoblasts are recruited to the excavated bone site, replacing the osteoclasts. Osteoblasts secrete type I collagen, osteocalcin, and other noncollagenous proteins, laying down new bone matrix at the resorption site—a process that takes three to four months to complete. Systemic Factors Affecting Bone Metabolism. Under normal physiological conditions, bone turnover is intricately controlled through the systemic release of hormones and growth factors that promote bone resorption or bone formation (e.g., PTH, estrogen, calcitonin, insulin-like growth factors [IGFs]). Dysregulation of any one of these mechanisms can contribute to defective bone metabolism and osteoporosis, as seen in postmenopausal women (as a result of low estrogen levels) and in the elderly. The following sections set forth key factors in bone metabolism. Calcium. The systemic effects of hormones on bone cell activity are twofold: maintaining calcium homeostasis and regulating bone formation and resorption.

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Both effects are interdependent in the sense that bone is an important reservoir for calcium that can be transferred rapidly to or from the circulation in the event of, respectively, hypocalcemia or hypercalcemia. Although the hormones PTH and estrogen control bone turnover, their primary function is to maintain calcium homeostasis. Because calcium is the major mineral component of bone, adequate dietary calcium is essential for developing and maintaining normal skeletal tissue. Inadequate calcium intake over prolonged periods predisposes people to develop osteoporosis and can weaken bone structure and function over the long term. Parathyroid Hormone. Falling blood calcium concentrations stimulate the parathyroid gland to secrete PTH. The continued presence of elevated PTH indirectly increases osteoclast activity by stimulating osteoblasts via their cell-surface, PTH-speciÞc receptors. PTH also stimulates calcium reabsorption from the kidney and absorption from the intestine. The net result is to mobilize calcium from bone to restore blood calcium levels. Chronically elevated PTH levels result in net bone loss due to constant stimulation of osteoclasts. However, intermittent dosing with PTH can stimulate overall bone turnover, thereby increasing bone mass (Jilka RL, 1999). Evidence suggests that the body’s responsiveness to PTH decreases with age, resulting in impaired calcium homeostasis and bone metabolism. PTH also stimulates IGF-1, which promotes long-bone growth. The bone-forming properties of PTH led to the development of one currently marketed therapy, teriparatide (Eli Lilly’s Forteo); several others are in various stages of clinical development (for more information, see “Emerging Therapies”). Vitamin D. Vitamin D is produced in the skin in response to sunlight and, under the inßuence of elevated PTH, is metabolized in the kidney to its active form: 1,25-(OH)2 vitamin D3 (calcitriol). Like PTH, calcitriol promotes calcium absorption from the intestine in response to falling blood calcium levels. Calcitriol restores blood calcium levels by activating speciÞc receptors on osteoblasts, thereby stimulating bone resorption. Conditions that affect vitamin D metabolism, including aging, can adversely affect calcium homeostasis and bone mineralization. Estrogen. Estrogen plays a critical role in the control of bone metabolism, partly by increasing intestinal absorption of calcium and partly by modulating bone formation and resorption through local mechanisms. These mechanisms are not yet fully understood. However, researchers have observed a correlation between declining estrogen levels and reduced osteoprotegerin levels (Rogers A, 2002). Osteoprotegerin is a pro-apoptotic cytokine that promotes programmed cell death in osteoclasts. Therefore, reductions in circulating estrogen prolong osteoclast lifespans. At the same time, low estrogen may shorten the life span of osteoblasts, thereby shifting the balance toward net bone resorption, which causes osteoporosis. Calcitonin. In response to a rise in blood calcium levels, thyroid parafollicular C cells secrete calcitonin, which is an inhibitor of osteoclasts that acts through calcitonin-speciÞc membrane receptors. Calcitonin blocks bone resorption, thus

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sparing calcium bone stores and preventing bone loss. Its effectiveness as a treatment for osteoporosis is discussed in “Current Therapies.” Growth Hormone. Growth hormone is produced by the pituitary gland and promotes long-bone growth (i.e., increased length) in children and adolescents and bone thickening in adults, apparently by stimulating osteoblasts. Its effects are mediated by IGFs produced by the liver and other tissues, including bone. In humans, PTH stimulates IGF-1 production, which in turn promotes bone formation. The production of growth hormone and IGFs declines after age 30. CURRENT THERAPIES The primary objective of osteoporosis treatment is the prevention of bone fractures, which are a cause of signiÞcant disability. Currently available therapies are labeled for the prevention of osteoporosis and/or treatment of the disease. Agents used in the prevention of osteoporosis include antiresorptive therapies (e.g., bisphosphonates, hormone replacement therapy [HRT], and selective estrogen receptor modulators [SERMs]). These agents work to maintain or slow the loss of bone mineral density (BMD) in postmenopausal women who have osteopenia by inhibiting bone turnover. Historically, HRT was a mainstay in the treatment of osteopenia (i.e., prevention of osteoporosis) in postmenopausal women. However, data from trials sponsored by the Women’s Health Initiative (WHI) of the National Heart, Blood, and Lung Institute heralded a shift from HRT toward bisphosphonate and SERM therapy instead. Results of the WHI study demonstrated that the modest beneÞts of HRT for BMD maintenance are outweighed by its potential to increase the risk of cardiovascular disease, stroke, breast cancer, and endometriosis. However, low-dose HRT still has a role in treatment of patients who require relief from postmenopausal symptoms such as vasomotor hyperactivity (hot ßashes). Agents used in the treatment of osteoporosis include bisphosphonates (which are also used for disease prevention in the osteopenia subpopulation), novel bone-forming agents, and calcitonin. These agents are used to treat patients who have already lost considerable BMD. In addition to these pharmacological therapies, physicians commonly prescribe vitamin D and calcium supplements to both osteopenic and osteoporotic patients to help maintain BMD and provide components necessary to mineralize bone. With the introduction of the bone-forming agent teriparatide (Eli Lilly’s Forteo/Forsteo) in 2003, physicians had access for the Þrst time to an anabolic therapy that works not only to inhibit loss of bone mineral density but also to increase bone mass. The agents listed in Table 4 have been shown to stabilize or increase BMD, two major goals of osteoporosis therapy. Another major goal is prevention of fractures; relevant clinical trial data on vertebral and nonvertebral fracture risk associated with these currently marketed therapies are discussed in the following sections. The beneÞts provided by each of the available classes of osteoporosis treatment and their documented risks are presented in Table 5.

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243

TABLE 4. Current Therapies Used for Osteopenia and Osteoporosis Agent

Company/Brand

Parathyroid hormone analogues Teriparatide Eli Lilly’s Forteo/Forsteo Bisphosphonates Alendronate Merck’s Fosamax Risedronate

Sanofi-Aventis/ Procter & Gamble’s Actonel Etidronate Procter & Gamble/ Sumitomo’s Didronel Selective estrogen receptor modulators Raloxifene Eli Lilly’s Evista Hormone replacement therapy Estrogens/progestogens Conjugated Wyeth’s Premique/ estrogen/MPA Prempro/ Premphase Estrogens Conjugated equine Wyeth’s Premarin, estrogen others Calcitonins Novartis’s Calcitonin (intranasal) Miacalcic/Karil, generics Synthetic steroids Tibolone Organon’s Livial/Boltin Synthetic isoflavones Ipriflavone Takeda’s Iprosten/Osten, generics

Dose

Availability

20 µg injection qd

US, F, G, S, UK

70 mg qw, or 10 mg qd 35 mg qw, or 5 mg qd

US, F, G, I, S, UK, J US, F, G, I, S, UK, J

400 mg qd for 14 days every three months

F, G, S, UK

60 mg qd

US, F, G, I, S, UK, J

0.625 mg/2.5 mg qd, or 0.625 mg/5 mg qd

US, F, G, I, S, UK, J

0.625 mg qd, or 1.25 mg qd

US, F, G, I, S, UK, J

200 IU qd

US, G, I, S, UK, J

2.5 mg qd

F, G, I, S, UK

600 mg qd

J

qd = Once daily; qw = Once weekly; IV = Intravenous; MPA = Medroxyprogesterone acetate; IU = International unit. US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan.

Parathyroid Hormone Analogues Overview. The only bone-forming agent currently approved for treatment of osteoporosis is teriparatide (Eli Lilly’s Forteo/Forsteo), a recombinant form of the Þrst 34 amino acids of human parathyroid hormone (PTH). Lilly is collaborating with Emisphere Technologies to develop an oral formulation of teriparatide; Lilly is also in a partnership with Nektar Therapeutics to develop an intranasally administered form of the drug. The oral formulation of teriparatide is in Phase I clinical development, but no news is available about the intranasal formulation. Several other companies are developing different formulations of full-length or truncated

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TABLE 5. Comparison of Current Therapies for Osteopenia and Osteoporosis Compound Parathyroid hormone analogues

Bisphosphonates

Selective estrogen receptor modulators

Hormone replacement therapy

Calcitonin (intranasal) Synthetic steroids

Advantages • Decrease the risk of vertebral fractures by 65% in patients with osteoporosis after 18 months of therapy. • Decrease the risk of nonvertebral fractures by 53% in patients with osteoporosis after 18 months of therapy. • Reduce the risk of both vertebral and nonvertebral fractures by 40–50% over three years of therapy. • Can be used in both male and female patients. • Agents available in onceweekly oral formulations. • Reduce risk of vertebral fracture by 30% in patients with prior fractures over three years of therapy. • Reduce risk of vertebral fracture by 55% in patients without prior fractures over three years of therapy.

• Can reduce the risk of vertebral and hip fractures by 34%. • Has beneficial effects on postmenopausal symptoms (e.g., hot flashes).

• Generally well tolerated. • May provide relief of bone pain. • Can increase BMD by 2–7%. • Agents offer some benefits of HRT without increasing the risk of cardiovascular complications or cancer.

Disadvantages • High cost. • Require daily subcutaneous injection. • Long-term use can cause rare bone cancers in rodents (Vahle JL, 2002). • Many patients exhibit upper GI discomfort (e.g., heartburn, stomach ulcers). • Inconvenient dosing requirements: i.e., drug must be taken 30 minutes prior to first meal and patient must remain upright. • May increase the frequency of hot flashes and other postmenopausal symptoms. • Like estrogen, can increase the risk of thromboembolic events. • Cannot be used in male patients. • No evidence of efficacy in preventing hip fractures. • Increases risk of cardiovascular problems (e.g., heart attack, stroke, deep venous thrombosis) and breast cancer over five years of treatment. • Cannot be used in male patients. • Runny nose. • Nose bleeds. • Efficacy unproven. • Poor efficacy. • Cannot be used in male patients. • May cause vaginal bleeding in up to one-third of patients.

BMD = Bone mineral density; GI = Gastrointestinal; HRT = Hormone replacement therapy.

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245

PTH (see “Emerging Therapies”). Although still in early-stage development, these products are expected to offer signiÞcant improvements in convenience and patient compliance compared with teriparatide, which is currently available only as a daily subcutaneous injection. Mechanism of Action. Unlike antiresorptive therapies, which prevent BMD loss by slowing the rate of bone turnover, bone-forming agents work by increasing the rate of bone production. The primary physiological role of PTH is to maintain serum calcium homeostasis (Figure 2). Low serum calcium levels trigger the release of PTH, which raises serum calcium levels by three mechanisms: stimulating the release of calcium from bone, decreasing renal excretion of calcium, and increasing intestinal absorption of calcium. Whereas chronically elevated PTH levels result in net bone loss due to constant stimulation of osteoclast activity, intermittent administration of low doses of PTH has been found to stimulate osteoblast activity, thereby increasing bone mass. The stimulation of osteoblast activity has been attributed in part to the direct activity of PTH in inhibiting osteoblast apoptosis, thereby increasing the number of these bone-forming cells (Jilka RL, 1999). In contrast, continuously elevated levels of PTH result in hypercalcemia and increased rates of bone resorption via increased expression of the receptor for activation of nuclear factor κB ligand (RANKL), which stimulates osteoclast differentiation and activity (Ma YL, 2001). The human PTH protein consists of 84 amino acids; scientists have discovered that the Þrst 34 amino acids of PTH (PTH 1-34) are sufÞcient to exert all the effects of the full-length PTH molecule (Ejersted C, 1993; Kimmel DB, 1993). NPS Pharmaceuticals is developing Preos/Preotact, a PTH compound that contains the full 84 amino acids of human PTH (PTH 1-84); unpublished clinical studies indicate that PTH 1-84 is nearly indistinguishable from PTH 1-34 in its safety and efÞcacy proÞle (NPS Pharmaceuticals, press release, March 30, 2004). (For more information on Preos/Preotact, see “Emerging Therapies”.) Teriparatide. Teriparatide (Eli Lilly’s Forteo/Forsteo)∗ is a recombinant form of the Þrst 34 amino acids of human PTH (PTH 1-34). Teriparatide was approved in the United States in November 2002 for treatment of osteoporosis in postmenopausal women who have a history of fracture, who are at high risk of fracture, or who have failed to increase BMD following other osteoporosis therapies (Eli Lilly, Forteo package insert; Eli Lilly, press release, November 26, 2002). Teriparatide is also approved for use in men with primary or hypogonadal osteoporosis who are at high risk of fracture. The U.S. labeling for teriparatide includes a “black-box” warning about the increased risk of osteosarcoma, although teriparatide has not been found to increase the risk of this rare cancer in human patients; the black-box warning was prompted by the results of preclinical studies in rodents treated with high doses of teriparatide (Schneider BS, 2001; Vahle JL, 2002). Teriparatide is therefore contraindicated in patients who are at increased risk of osteosarcoma (e.g., patients with Paget’s disease). The European Union’s Committee for Proprietary Medicinal Products (CPMP) recommended in June 2003 that teriparatide be appr

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FIGURE 2. Effect of parathyroid hormone on serum calcium hemostasis.

oved, but only for postmenopausal women at high risk of fracture. In 2000, Asahi Kasei initiated Phase III clinical trials of teriparatide as a once-weekly injection. In 2001, researchers published results from a pivotal Phase III trial that measured the reduction in incidence of new fractures following intermittent teriparatide administration (Neer RM, 2001). The trial included 1,637 women who were given either placebo or a daily injection of 20 µg or 40 µg teriparatide. Teriparatide increased lumbar spine BMD by 9–13%, reduced the risk of vertebral fracture by 65–69%, and lowered the risk of nonvertebral fragility fracture by 53–54%. Teriparatide also increased BMD in the hip by 4–6% and total body BMD by 2–4% more than placebo. Side effects of teriparatide included nausea, dizziness, leg cramps, and transient hypercalcemia; their incidence was found to increase with use of the higher dose (40 µg) of teriparatide (Neer RM, 2001). The risk of transient hypercalcemia requires monitoring of patient serum calcium levels. The effects of teriparatide on male patients were examined in a study that enrolled 437 men with secondary osteoporosis caused by hormonal insufÞciency (Orwoll ES, 2003). Patients were randomized to receive daily injections of either 20 µg or 40 µg of teriparatide or placebo for a median duration of 11 months. Patients treated

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with 20 µg or 40 µg teriparatide had signiÞcant increases in vertebral BMD (5.9% and 9.0%, respectively), compared with an increase of less than 1% in patients treated with placebo. More than 80% of the men were followed up after treatment for an additional 18 months to assess vertebral fracture incidence (Orwoll ES, 2002). Men treated with teriparatide had a 50% reduction in the risk of developing new vertebral fractures and an 83% reduction in the risk of suffering a moderate or severe fracture. Another study involving male patients examined the effect of combination therapy with synthetic PTH 1-34 and the bisphosphonate alendronate (Merck’s Fosamax) on 83 men, aged 46–85, who had low BMD (Finkelstein JS, 2003). Patients were randomized to receive alendronate (10 mg/day orally) or PTH 1-34 (40 µg/day by subcutaneous injection) or both agents. Patients who received alendronate were treated for 30 months; patients treated with PTH 1-34 began therapy six months after the start of the trial and were treated for a total of 24 months. The delayed PTH 134 treatment allowed for six months of bisphosphonate use in patients who were treated with both agents to simulate the switching of patients from alendronate to PTH 1-34. Additionally, all patients were maintained with 1,000–1,200 mg calcium and 400 IU vitamin D daily. In this study, combination therapy was not as effective as PTH 1-34 monotherapy. Patients treated with PTH 1-34 exhibited signiÞcantly greater increases in lumbar spine BMD (18%) compared with patients treated with alendronate (8%) or combination therapy (15%). The apparent attenuation of teriparatide’s anabolic effect on BMD in patients treated with combination therapy was even more pronounced in quantitative computerized tomography (QCT) scans of vertebral BMD. Patients treated with PTH 1-34 experienced a 48% increase in vertebral BMD, compared with a 3% increase in patients taking alendronate and a 17% increase in patients treated with combination therapy. Preliminary data showing similar results in female patients were reported at the 2004 annual meeting of the Endocrine Society in New Orleans (Neer R, 2004). The effect of sequential treatment with an antiresorptive agent and teriparatide was investigated in female patients in a small uncontrolled study that enrolled 59 postmenopausal women (aged 60–87) who had been treated previously with alendronate or raloxifene (a SERM) for 18–36 months (Ettinger B, 2004). All patients were also treated with 1,000 mg calcium and 400 IU vitamin D daily. Patients who had received prior treatment with raloxifene exhibited 10% increased lumbar spine BMD after 18 months of teriparatide treatment, compared with a 4% increase in patients who had been treated with alendronate. Moreover, total hip BMD increased 1.8% in the raloxifene group following teriparatide treatment; patients in the alendronate group exhibited no improvements in total hip BMD. Although this study was small and not placebo-controlled, it suggests that the lingering effects of bisphosphonate treatment may impair the anabolic activity of teriparatide—an effect not observed with raloxifene. Bisphosphonates Overview. Bisphosphonates have been used for the treatment of osteoporosis for more than 30 years. As antiresorptive agents, bisphosphonates work by slowing down bone turnover; the inhibition of osteoclast activity allows enhanced

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osteoblast activity, thereby increasing BMD. In addition to osteoporosis treatment, many agents in this class are used to treat such conditions as hypercalcemia, osteomalacia, Paget’s disease, and cancer-related bone degeneration. This section proÞles the three most popular bisphosphonates approved for the prevention and treatment of osteoporosis: alendronate (Merck’s Fosamax), risedronate (SanoÞ-Aventis/Procter & Gamble’s Actonel), and etidronate (Procter & Gamble’s Didronel). Alendronate and risedronate are available in once-daily and once-weekly oral formulations; etidronate offers only once-daily oral administration. Ibandronate (Roche/GlaxoSmithKline’s Boniva/Bonviva) is a key emerging agent that was approved in a daily oral formulation for treatment of osteoporosis by the FDA in May 2003 and by the European Commission in February 2004 (Roche, company press releases, May 19, 2003, and February 27, 2004). Roche and GlaxoSmithKline have also Þled for approval in the United States and Europe to market once-monthly oral and quarterly injectable formulations of ibandronate as treatments for osteoporosis (see “Emerging Therapies” for details). Other bisphosphonates that are used off label or to a lesser extent in the treatment of osteoporosis include pamidronate (Novartis’s Aredia), neridronate (Abiogen Pharma’s Nerixia), clodronate (Boehringer Ingelheim’s Bonefos), and zoledronate (Novartis’ Zometa), which is in development for osteoporosis as a once-yearly injectable formulation (see “Emerging Therapies”). These agents are available only in speciÞc markets, and in some cases, only for speciÞc indications, such as treatment of osteomalacia and Paget’s disease. However, many physicians use these agents off label for treatment of osteoporosis because of their convenient long-acting, injectable formulations or, in the case of the oral formulation of clodronate, their low cost. The improved convenience offers a tremendous beneÞt to patients who cannot tolerate the gastrointestinal discomfort associated with oral bisphosphonates or are not compliant on oral therapies that require more frequent dosing. Mechanism of Action. Bisphosphonates are stable pyrophosphate analogues that bind to hydroxyapatite in bone and inhibit bone resorption by decreasing the number and activity of osteoclasts. By inhibiting osteoclast activity, bisphosphonates promote bone formation by osteoblasts, thereby increasing bone mass and strength. Bisphosphonates exert their inhibitory effect on osteoclasts by inhibiting farnesyl pyrophosphate synthase, an enzyme in the cholesterol synthesis pathway that is required for the activation of signaling molecules that trigger osteoclast recruitment, adhesion to bone matrix, and resorptive activity. Bisphosphonates may also activate apoptotic pathways in osteoclasts, thereby reducing their numbers in the body (Benford HL, 2001). Alendronate. Alendronate (Merck’s Fosamax) (Figure 3) is a second-generation bisphosphonate with greater efÞcacy in treating osteoporosis than Þrstgeneration agents in this class. The addition of an amino group in the molecule increases the drug’s potency by increasing its half-life. Once-daily alendronate has been approved in the United States since 1995 for the prevention and treatment of osteoporosis in women, in Europe since 1996, and in Japan since late

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PO3H2 H2N

OH PO3H2

FIGURE 3. Structure of alendronate.

2001. A once-weekly oral formulation of alendronate is available in all markets except Japan. Alendronate was approved for use in male patients in the United States during the second half of 2000 and is used to treat men in all of the major pharmaceutical markets. The Fracture Intervention Trial (FIT) was a large, four-year, placebo-controlled Phase III study that investigated the effect of alendronate on 4,432 postmenopausal women diagnosed with osteopenia or osteoporosis. The treatment arm received 5 mg alendronate daily for the Þrst two years; the dose was increased to 10 mg daily for the Þnal two years. BMD measurements for the spine, femoral neck, and total hip increased by 8.3%, 3.8%, and 3.4%, respectively, in the alendronate-treated patients; in the placebo group, BMD increased 1.5% in the spine but decreased 0.8% in the femoral neck and 1.6% in the total hip (Cummings SR, 1998). Although no signiÞcant decrease in overall incident fractures occurred, the study did demonstrate that women who had the lowest BMD (more than 2.5 standard deviations (SD) below the normal adult mean) beneÞted the most from alendronate therapy. These patients exhibited a 44% reduced risk for a new vertebral fracture and a 36% reduced risk for femoral fractures. FIT also found alendronate to be beneÞcial for osteoporotic women who had previously experienced vertebral fracture. The Þrst arm evaluated 2,027 women aged 55–80 who were randomized to receive alendronate (5–10 mg daily) or placebo for three years. Results revealed that relative to placebo, alendronate reduced the incidence of new vertebral fractures by 47%, new wrist fractures by 48%, and new hip fractures by 51% (Black DM, 1996). A subsequent analysis of FIT data from 3,658 women who had existing fractures or had osteoporosis (T-score < −2.5) but no fracture found similar reductions in fracture risk (Black DM, 2000). The effect of continuous, long-term use of alendronate in the original cohort from FIT was investigated in the FIT Long-Term Extension (FLEX) study. FLEX enrolled 1,099 women between the ages of 60 and 86 who were treated with alendronate (by this time, for an average of Þve years) in FIT. Patients were randomized to receive continued treatment with alendronate (5 or 10 mg/day) or placebo for Þve more years. All patients were also given supplemental calcium (500 mg/day) and vitamin D (250 IU/day). Interim results at the three-year midpoint indicate that patients who continued treatment with alendronate experienced 2.0% and 2.5% increases in hip and spine BMD, respectively, compared with patients who were switched to placebo. Patients on long-term alendronate therapy also exhibited a slower rate of bone turnover compared with patients in the placebo group, which was demonstrated by analysis of two biochemical

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markers of bone turnover. However, all patients demonstrated decreased bone turnover and increased BMD compared with baseline values obtained prior to initial treatment with alendronate in the original FIT study, which indicates that alendronate treatment has long-lasting effects (Ensrud KE, 2004). Similar results afÞrming alendronate’s long-term efÞcacy were obtained in the extension of the Alendronate Phase III Osteoporosis Treatment study (Bone HG, 2004). In this randomized, placebo-controlled study, 247 postmenopausal women with osteoporosis (out of a total of 994 enrolled in the original study [Liberman UA, 1995]) were treated with 10 mg alendronate and 500 mg calcium daily for a total of 10 years. Women enrolled in the trial extension averaged 63 years of age at the outset of the original study. Patients on sustained alendronate therapy exhibited increased BMD at the lumbar spine (13.7%), femoral neck (5.4%), and total proximal femur (6.7%), compared with baseline values obtained prior to initial treatment in the earlier study. Furthermore, as observed in the FLEX study, patients had lower levels of biochemical markers of bone turnover while treated with alendronate; levels of these markers increased within a year after cessation of therapy. One major advantage of the bisphosphonates is their usefulness for treating male osteoporosis patients (for obvious reasons, HRT and SERMs are not an option for this group). Although very few clinical studies of alendronate in male patients with primary osteoporosis have been reported in the literature, treatment with alendronate has been proved to increase BMD and prevent incident fractures in men. One study of 241 men found that two years of alendronate treatment (10 mg daily) increased BMD by 5.3% in the spine, 2.6% in the femur, and 2.6% in the hip, compared with placebo (Orwoll E, 2000). The incidence of vertebral fracture in alendronate-treated men was 0.8%, compared with 7.1% in men who received placebo. In another study, 77 patients (average age 57) were randomized to receive either 10 mg alendronate and 1,000 mg calcium daily, or treatment with only 1,000 mg/day calcium for three years. At the conclusion of the study, patients in the alendronate arm exhibited increased BMD at the lumbar spine (8.8%), femoral neck (4.2%), and total hip (3.9%). In comparison, patients treated with calcium alone had minor decreases in BMD (−0.3% to −1.2%) at each of these three locations (Gonnelli S, 2003). Fracture incidence was not examined in this trial. One open-label, prospective study investigated fracture risk in 134 male patients with primary osteoporosis who were given daily treatment with either 10 mg alendronate or 1 µg alfacalcidol (a precursor of vitamin D). Results from this trial demonstrated a dramatic reduction in fracture risk with alendronate treatment. Over three years, the incidence of new vertebral fractures was 10.3% in patients treated with alendronate and 24.2% in patients receiving alfacalcidol. Furthermore, patients receiving alendronate exhibited increased BMD at the lumbar spine (11.5%) and the femoral neck (5.8%). In comparison, patients on alfacalcidol had modest 3.5% and 2.3% increases in BMD at these two locations respectively (Ringe JD, 2004). Once bound to bone, bisphosphonates have a long half-life, so researchers think it logical that intermittent therapy might be just as effective as daily

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treatment. The efÞcacy of different formulations of alendronate (daily 10 mg, twice-weekly 35 mg, and once-weekly 70 mg) was investigated in a nonplacebocontrolled trial that enrolled 1,258 postmenopausal women. Regardless of the treatment regimen, each treatment group exhibited elevated BMD of approximately 5% at the hip, femoral neck, wrist, and whole body, thereby demonstrating the equivalence of the once-weekly and once-daily formulations of alendronate (Schnitzer T, 2000). The once-weekly formulation of alendronate was developed to alleviate one of the agent’s major side effects: upper gastrointestinal (GI) events (e.g., gastroesophageal reßux or esophageal erosion). While many trials of daily alendronate have shown no difference in the frequency of adverse GI events between treatment and placebo groups, these studies have not represented the general population because they often excluded patients with preexisting upper GI problems and/or patients at risk for such events. In the Schnitzer study, 22–24% of patients in the three treatment groups suffered from at least one adverse upper GI event. The frequency of serious GI side effects (e.g., gastric ulcer, gastritis, or esophageal stricture) was only 1.4% in the daily treatment group, but no patients in the once-weekly group exhibited severe GI problems, thus demonstrating the improved side-effect proÞle of the once-weekly formulation of alendronate (Schnitzer T, 2000). Risedronate. The third-generation bisphosphonate risedronate (SanoÞAventis/Procter & Gamble’s Actonel) (Figure 4) launched as a once-daily treatment for osteoporosis in the United States and Europe in 2000. Risedronate is marketed by Takeda and Ajinomoto in Japan, where it launched in 2002. Like alendronate, risedronate is available in once-daily and once-weekly oral formulations in the United States and Europe; the once-weekly formulation is not yet available in Japan. Risedronate offers greater potency than alendronate and the Þrst-generation bisphosphonates thanks to the addition of a heterocyclic ring to the basic pyrophosphate structure. Three large clinical trials have examined the use of once-daily risedronate to decrease the risk of fracture in postmenopausal, osteoporotic women. Under the auspices of the Vertebral EfÞcacy with Risedronate Therapy (VERT) study group, two separate three-year trials were conducted, one in North America and another in Europe and Australia. In the North American study (VERT-NA), 2,458 women with evident fracture were treated with placebo or 2.5 or 5 mg risedronate daily (Harris ST, 1999). After one year, the 2.5 mg arm of the study was discontinued

PO3H2 OH PO3H2 N FIGURE 4. Structure of risedronate.

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as a result of protocol amendment. In the Þrst year of treatment, researchers observed a 65% reduction in the risk of vertebral fracture with risedronate. During the three-year period, 5 mg risedronate decreased the incidence of new vertebral and nonvertebral fracture by 41% and 39%, respectively, compared with placebo. Similar results were obtained in the European and Australian arms of the VERT trial (VERT-MN), in which 1,226 women who had two vertebral fractures were treated with placebo or 2.5 or 5 mg risedronate daily. As in the North American study, the 2.5 mg treatment arm was discontinued. Treatment with 5 mg risedronate decreased the risk of new vertebral fracture by 49% and the risk of nonvertebral fracture by 33% (Reginster J-Y, 2000). The results of a two-year extension of the VERT-MN trial demonstrated that the reduction in incidence of new vertebral fracture was maintained during the entire Þve-year period (Sorensen OH, 2003). An analysis of pooled data from the two VERT trials revealed that patients at greatest risk of new fractures (because of advanced age, higher prevalence of previous fractures, or lower BMD [T-score ≤ 2.5 SD below young adult mean]) who were treated with 5 mg/day risedronate for one year exhibited a 62% reduction in the risk of new vertebral fracture, compared with patients receiving placebo. Moreover, the risk of suffering multiple new vertebral fractures was reduced by 90% in the 5 mg/day treatment arm (Watts NB, 2003). In the third major clinical trial with risedronate, the Hip Intervention Program (HIP) study, the primary efÞcacy end point was incidence of hip fracture. A total of 9,331 postmenopausal women aged 70 or older enrolled in this multicenter study. The study was divided into two groups: patients aged 70–79 who had osteoporosis and patients aged 80 or older who had clinical risk factors or hip fracture. Patients were randomized to receive 2.5 or 5 mg risedronate or placebo daily. Although the group deÞned by risk factors had no signiÞcant reduction of incident hip fracture, a 40% hip fracture risk reduction relative to placebo was seen in the 70–79 age-group with osteoporosis (McClung MR, 2001). The difference between once-daily risedronate and once-weekly alendronate in terms of upper GI events is not as striking (Merck, press release, May 12, 2002) as the difference between once-daily risedronate and once-daily alendronate (Lanza FL, 2000). Results of a randomized, placebo-controlled trial involving 550 patients from ten countries were presented at the International Osteoporosis Foundation World Congress on Osteoporosis in Lisbon, Portugal, May 10–14, 2002. Over a six-month period, researchers observed no differences in adverse events between once-weekly alendronate and once-daily risedronate. These results were conÞrmed in a randomized, double-blind, placebo-controlled study that compared the efÞcacy of daily risedronate (5 mg) to that of once-weekly risedronate (35 or 50 mg) in 1,456 postmenopausal osteoporotic women; this study demonstrated no differences in the frequency of adverse events in any of the treatment groups (Harris ST, 2004). Etidronate. Etidronate (Procter & Gamble/Sumitomo’s Didronel, generics)∗ , a Þrst-generation bisphosphonate, was the Þrst bisphosphonate to be marketed for

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osteoporosis in Europe. In the United States, the drug is approved for treatment of Paget’s disease and malignancy-associated hypercalcemia, but it is sometimes prescribed off label for osteoporosis treatment. Although alendronate and risedronate have become more popular for treatment of osteoporosis because of their efÞcacy and convenient once-weekly formulations, etidronate is an inexpensive drug and therefore has a role in cost-conscious health care systems. The Þrst reports of etidronate’s efÞcacy came from a two-year, double-blind, placebo-controlled trial of 429 women who had osteopenia and at least one prevalent vertebral fracture (Watts NB, 1990). The study population was randomized to receive either cyclical etidronate (400 mg daily for two weeks, followed by 10 weeks off therapy) or placebo. Both groups were further subdivided to receive either 2 g phosphate daily or placebo during the “off therapy” days. In the two groups receiving etidronate, vertebral BMD increased 4.2% and 5.2% respectively. The combined rate of new vertebral fracture fell by 50% compared with the rate in patients who did not receive etidronate. Many additional trials of etidronate have conÞrmed the efÞcacy of this agent. A meta-analysis of 13 randomized, placebo-controlled trials that investigated the efÞcacy of etidronate in postmenopausal women found a 37% reduction in vertebral fracture and increased BMD in the spine (4.1%), femoral neck (2.4%), and total body (1%) relative to placebo (Cranney A, 2001). Like other bisphosphonates, etidronate is characterized by poor GI absorption and is associated with GI side effects. However, etidronate’s lower GI side effects (nausea and diarrhea) occur rarely and are better tolerated than the upper GI side effects (gastric and esophageal ulcers) associated with once-daily alendronate. A disadvantage unique to etidronate is its potential to induce osteomalacia (bone mineralization defects) because of its combined effects: it inhibits both bone formation and bone resorption. However, this side effect can be prevented by administering etidronate in cycled regimens such as those described earlier. Selective Estrogen Receptor Modulators Overview. Selective estrogen receptor modulators (SERMs) are nonsteroidal benzothiophenes that trigger some of the beneÞcial effects of estrogen on bone metabolism without activating estrogen’s undesirable effects on the endometrium or breast tissue. SERMs were initially developed for treatment of breast cancer because of their antiestrogenic effect on breast tissue, but they have become an important class of agents for treatment of osteoporosis. Raloxifene (Eli Lilly’s Evista) is the only SERM approved in the major pharmaceutical markets for the prevention and treatment of osteoporosis. Mechanism of Action. Like estrogen, SERMs decrease bone resorption and the rate of bone turnover. By selectively activating certain subtypes of estrogen receptors, SERMs promote the beneÞcial effects of estrogen without elevating the risk of breast and endometrial cancer. However, currently marketed SERMs can trigger increased vasomotor activity in patients who are already prone to hot

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FIGURE 5. Structure of raloxifene.

ßashes and may also increase the risk of thromboembolism. Unlike HRT, SERMs have been shown to have no effect on reproductive tissues (Bryant HU, 2001). The binding of SERMs to estrogen receptors results in the activation of genes containing estrogen response elements (Riggs BL, 2003). One mechanism by which SERMs exert their effects on bone is increased production of osteoprotegerin, a soluble decoy receptor for RANKL. Sequestration of RANKL by osteoprotegerin inhibits the development of osteoclasts from precursor cells, thereby promoting the activity of osteoblasts (Viereck V, 2003). Raloxifene. Raloxifene (Eli Lilly’s Evista) (Figure 5) is a metabolite of tamoxifen (AstraZeneca’s Nolvadex, generics) and the only SERM approved for prevention and treatment of osteoporosis. Raloxifene has been marketed in most countries for prevention of osteoporosis since 1998 and gained approval for treatment of osteoporosis in 1999 in the United States and in 2000 in Europe. Chugai and Eli Lilly Japan are comarketing raloxifene in Japan (Chugai, press release, May 11, 2004). The largest clinical trial to determine the effects of raloxifene on fracture risk in postmenopausal women is the Multiple Outcomes of Raloxifene Evaluation (MORE) study. This multi-center, placebo-controlled trial enrolled 7,705 women between 31 and 80 years of age from 25 countries who were at least two years postmenopausal and met the World Health Organization (WHO) criteria for osteoporosis and osteopenia. Patients were randomized to receive either 60 mg or 120 mg raloxifene daily or placebo; researchers measured results at three years. Overall, the incidence of new vertebral fracture was lower in the raloxifenetreated patients: 6.6% and 5.4% in the 60 mg and 120 mg groups, respectively, compared with 10.1% in the placebo group (Ettinger B, 1999). The frequency of vertebral fracture decreased with raloxifene treatment regardless of the existence of prevalent fractures at baseline. Additionally, both doses of raloxifene caused signiÞcant increases in BMD (2.1–2.7% more than placebo). Raloxifene’s effects on vertebral fracture risk and BMD were similar at year four of the MORE

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study (Delmas PD, 2002[b]). Analyses of nonvertebral fractures, however, found no statistically signiÞcant differences in fracture incidence rates between the raloxifene-treated groups and placebo. A separate analysis of 3,204 women enrolled in the MORE study who did not have vertebral fractures at the outset of the trial was done to compare the efÞcacy of raloxifene in women with osteopenia (N = 2, 557) or osteoporosis (N = 635) as determined by total hip BMD T-scores. Patients included in this analysis were randomized to receive either placebo or 60 mg raloxifene daily for three years. Osteoporotic patients receiving placebo had a 6.4% incidence of new vertebral fracture during the study period, compared with an incidence of 3.6% in patients with osteopenia. In comparison, the overall risk of fracture in osteoporotic and osteopenic patients treated with raloxifene was 2.0% and 1.9%, respectively. Treatment with raloxifene for three years resulted in a 47% reduction in the incidence of new vertebral fracture in osteopenic women and a 69% reduction in osteoporotic women (Kanis JA, 2003). Given the similar mechanisms of action that estrogens and SERMs exert on bone metabolism, much of the interest in SERMs has focused on the reduced risk of adverse effects compared with HRT. However, in the MORE study, the relative risk of developing thromboembolism was three times greater for patients receiving raloxifene than for patients receiving placebo—a slight elevation in risk compared with HRT (Kanis JA, 2003). However, raloxifene’s beneÞts in other areas distinguish it from HRT. For example, Þndings from the MORE study demonstrated that treatment with raloxifene reduced the risk of all invasive breast cancers by 72% over four years and reduced the risk of estrogen-receptor-positive invasive breast cancer by 84% (Cauley JA, 2001). Several studies also suggest that raloxifene is approximately as effective as HRT in reducing total cholesterol and low-density lipoprotein (LDL) cholesterol and increasing high-density lipoprotein (HDL) cholesterol (Delmas PD, 1997; Walsh BW, 1998; Miskic B, 2003). A secondary analysis of four-year data from the MORE study found that raloxifene reduced the number of cardiovascular events by 40%, compared with placebo, in women who were at high risk of acute coronary events (BarrettConnor E, 2002). In the clinical trials mentioned previously, raloxifene was associated with a low incidence of side effects (e.g., hot ßashes, breast pain, vaginal bleeding). Hormone Replacement Therapy Overview. Historically, hormone replacement therapy (HRT) has been the gold standard for preventing osteoporosis in peri- and postmenopausal women. A fall in estrogen levels during menopause causes an increase in bone turnover; as a result, the rate of bone resorption overtakes the rate of bone formation, leading to a decrease in BMD and development of osteoporosis. HRT slows the rate of bone turnover, thereby preserving BMD. An added beneÞt of HRT is the reduction of vasomotor symptoms (hot ßashes) in perimenopausal women. However, recent data from studies by the Women’s Health Initiative (WHI) and the Heart and

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Estrogen/Progestin Replacement Study (HERS II) have shed new light on the attendant risks of HRT use. HRT is commonly prescribed as unopposed estrogen, a cyclic combination of estrogen and progestogen, or as daily, continuous combination therapy. Estrogen alone has been found to increase the risk of endometrial cancer by three- to eightfold in postmenopausal women with an intact uterus (Weiderpass E, 1999). Therefore, unopposed estrogen is used only in women who have undergone a hysterectomy. All other women are treated with a combination of estrogen and a progestogen, which prevents the carcinogenic effects of estrogen on the endometrial lining. A wide variety of HRT products is marketed in the regions under study. HRT is available in oral, transdermal, and intravaginal formulations; some products contain only an estrogen or a progestogen while others provide a combination. Table 4 summarizes a representative sample of HRT products. Mechanism of Action. HRT products bolster women’s waning levels of estrogen, which helps maintain bone mass by inhibiting resorption of bone and increasing calcium absorption by the intestines. Estrogen’s mechanism of action is similar to that of the SERMs. However, the hormone does not offer selective activation of estrogen receptors. Therefore, HRT is associated with many negative side effects, such as the increased risk of breast and uterine cancers, that SERMs have been designed to avoid. Hormone Replacement Therapy. In clinical trials, HRT use for approximately Þve years or more has been shown to increase BMD. However, its effect on the risk of fracture incidence in postmenopausal women had not been conclusively demonstrated until results of the WHI were published. The WHI randomized 16,608 postmenopausal women to receive either estrogen plus progestin or placebo for an average 5.2 years. HRT reduced the risk of hip fracture by 33% and risk of all fractures combined by 24% (Rossouw JE, 2002). These results were conÞrmed by the Million Women Study, a population-based prospective study of 138,737 postmenopausal women between the ages of 50 and 69 who were followed for an average of 2.8 years (Banks E, 2004). The incidence of fracture in all patients during the study period was 3.7%. Patients who received HRT over the course of the study had a 38% reduction in fracture incidence; no protection against fracture was observed in patients who had ceased therapy for one or more years. Moreover, there were no statistically signiÞcant differences in the protection offered by different doses or types of HRT. Earlier studies generally showed that HRT has a beneÞcial effect on BMD and fracture risk. Results of the Postmenopausal Estrogen/Progestin Interventions (PEPI) trial, in which women were treated for three years with 0.625 mg of conjugated equine estrogens (Wyeth’s Premarin), with or without medroxyprogesterone acetate (MPA), demonstrated a 3–5% increase in BMD of the spine and a 1.7% increase in BMD of the hip in the group receiving HRT (Bush TL, 1996). The Iowa Women’s Health Study followed a cohort of 40,000 postmenopausal

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women for six years. Like the results of the Million Women Study, results from the Iowa Women’s Health Study showed that the risk of hip fracture was 47% lower in current users of HRT than in women who had never taken it (Folsom AR, 1995). In addition to protecting bone, HRT provides other beneÞts. It is often prescribed to alleviate menopausal symptoms (e.g., hot ßashes, night sweats, urogenital atrophy, vaginal dryness). Data from the WHI suggest that HRT may also protect against colorectal cancer. During the average follow-up (5.2 years), HRT users’ risk of colorectal cancer decreased by 37% compared with nonusers (Rossouw JE, 2002). Although HRT’s beneÞts for osteopenia and osteoporosis have been known for decades, considerable controversy has accompanied research into its other beneÞts and risks. Recent results from WHI and HERS II have provided the most deÞnitive data on HRT’s risks to date and prompted a sea change in the understanding of what constitutes appropriate use for osteoporosis prevention and treatment. The HERS trial was designed to assess the impact of HRT on cardiovascular events. Following menopause, the average woman’s risk of cardiovascular disease (CVD) increases substantially; researchers have postulated that a decrease in endogenous estrogen production is to blame and that estrogen replacement may offset the CVD risk. HERS began with 2,763 postmenopausal women under the age of 80 who had preexisting coronary artery disease (CAD). Patients were randomized to receive HRT (in the form of conjugated equine estrogens plus medroxyprogesterone acetate) or placebo for 4.1 years. Overall, no signiÞcant differences were found between the groups in terms of frequency of heart attacks or strokes (Hulley S, 1998). However, the HRT group suffered more CVD events during the Þrst year of treatment, but it demonstrated a lower risk during years three through Þve compared with patients on placebo. This discrepancy led investigators to conduct a follow-up study (HERS II) to determine whether the lower risk of CVD in the later years of HRT use would persist over time. Of the surviving women from HERS, 2,321 patients enrolled in HERS II. After the additional 2.7-year follow-up, the investigators found no signiÞcant difference in the rates of CVD, death from CVD, or nonfatal myocardial infarction in the HRT and placebo groups (Grady D, 2002). Investigators concluded that HRT did not confer a beneÞt for women at risk of CVD. Shortly after publication of the HERS II data, another clinical study conÞrmed the potential risks of HRT. The WHI enrolled 163,809 postmenopausal women aged 50–79 years to participate in a set of clinical trials. In one arm, 16,608 women with intact uteruses were randomized to receive conjugated equine estrogens plus MPA or placebo for 5.2 years. In May 2002, this arm of the WHI was stopped early after a safety review showed excess risks compared with beneÞts of treatment, and an early release of trial Þndings was published. The data demonstrate that, compared with placebo, HRT increased the risk of coronary heart disease (CHD; relative risk [RR] = 1.29), stroke (RR = 1.41), venous thromboembolic disease (RR = 2.06), and invasive breast cancer (RR = 1.26) (Rossouw JE, 2002; Cushman M, 2004).

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Another treatment arm of the WHI enrolled 10,739 postmenopausal women who had previously had a hysterectomy and treated them with estrogen only. This treatment arm was also stopped early due to preliminary data showing that the increased risk to patients outweighed the potential beneÞts of treatment. In this patient population, HRT was shown to have lowered the risk of CHD (RR = 0.91), breast cancer (RR = 0.77), and hip fracture (RR = 0.61). However, patients experienced increased risk of stroke (RR = 1.39), pulmonary embolism (RR = 1.34), and colorectal cancer (RR = 1.08) (Anderson GL, 2004). In summary, the Þndings from HERS II and WHI have changed practitioners’ view of the risks of HRT for postmenopausal women. HRT clearly is not effective in reducing CVD risk and actually poses excess risk of CVD (WHI) and thromboembolic events (HERS II and WHI). Furthermore, the link between HRT and breast cancer has been a persistent but controversial concern since the Nurses’ Health Study Þrst demonstrated an increased breast cancer risk in HRT users compared with women who had never used HRT (Colditz GA, 1992). Although the risk is small for any individual user, it aggregates in a large population. The decision to prescribe HRT, therefore, must be assessed in terms of each individual woman’s risk factors. Low-dose HRT may provide an alternative; several small-scale studies have shown that low-dose HRT may be effective in providing both relief of vasomotor symptoms and preservation of bone mass (Ettinger B, 2004; Gambacciani M, 2003). Currently, however, very few data on the safety of long-term use of low-dose HRT are available. Calcitonins Overview. Calcitonin is a hormone that plays an important role in bone metabolism and in the regulation of serum calcium and phosphorus levels. Although considered by many physicians to be less effective than bisphosphonates and SERMs in treating osteoporosis, calcitonin has the advantage of relieving bone pain, a symptom found in many osteoporotic patients. Synthetically prepared salmon calcitonin (which is more potent than human calcitonin and has a longer duration of action) is available in all markets under study for the treatment of osteoporosis. Because the GI tract absorbs calcitonin poorly, it is marketed in injectable (Novartis’s Miacalcin/Miacalcic, Asahi Kasei’s Elcitonin, generics) and intranasal (Novartis’s Miacalcic/Karil, generics) formulations. Mechanism of Action. Calcitonin, a 32-amino-acid peptide hormone produced by the thyroid gland, inhibits osteoclast activity by binding to calcitonin receptors on the surface of these cells, leading to a decrease in bone resorption. However, calcitonin also inhibits reabsorption of calcium and phosphorus in the kidney, thereby increasing their rates of urinary excretion. Calcitonin. Calcitonin’s efÞcacy in osteoporosis patients has been controversial during the three decades it has been available. Much of the early data on calcitonin measured increases in total serum calcium and BMD but not its ability to prevent

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fractures. Early clinical trials were often unblinded, and placebo controls were not usually included. A meta-analysis of 14 trials evaluated the effect of calcitonin (injectable and intranasal) on vertebral and nonvertebral fracture risk (Kanis JA, 1999). The majority of the trials used salmon calcitonin and had a duration of one to three years. The cumulative study population was relatively small: 1,300 men and women. In this analysis, calcitonin reduced the risk of any fracture by 57% compared with placebo; however, further analysis of patients with fractures rather than total numbers of fractures found that calcitonin reduced the risk of fracture for any one patient by 26%. The authors noted in their conclusion that this effect was relatively small compared with the effects of bisphosphonates and SERMs. The Prevent Recurrence of Osteoporotic Fractures (PROOF) trial was designed to provide more conclusive data on the effect of intranasal calcitonin on fracture rates. PROOF enrolled 1,255 postmenopausal women with osteoporosis who were randomized to receive 100, 200, or 400 IU intranasal calcitonin daily or placebo (Chesnut CH, 2000). At Þve years, only the 200 IU dose signiÞcantly reduced incident vertebral fracture—by 33% compared with placebo—and only the 400 IU dose was associated with a signiÞcant increase in spinal BMD. Combined with an inconsistent effect on biochemical markers of bone turnover and a 60% dropout rate over Þve years, the PROOF data cast doubt on the efÞcacy of calcitonin. Nonetheless, calcitonin continues to play a role in the prevention and treatment of osteoporosis. Patients who are unable to tolerate the GI side effects of bisphosphonates or suffer pain from fracture, for example, are likely candidates for calcitonin because the intranasal and injectable formulations bypass the GI tract altogether. However, intranasal calcitonin has been associated with rhinitis, nasal irritation, and headache, and injectable calcitonin may cause nausea, ßushing, and hypersensitivity in some patients. Synthetic Steroids Overview. Synthetic steroids are structurally similar to estrogen and can confer some of the effects of estrogen (including prevention of postmenopausal bone loss) without the negative effects on the cardiovascular system and breast tissue that are associated with estrogen use. Mechanism of Action. Tibolone is a synthetic steroid that has weak estrogenic, progesterogenic, and androgenic properties. Tibolone’s multiple effects are the result of the activity of its three metabolites. Its effects on bone are achieved by the two estrogenic metabolites, which stimulate estrogen receptors on the bone. In the endometrial tissue, only the third metabolite is active; it functions as a progestogen and therefore does not cause the endometrial hyperplasia that estrogen can trigger. Tibolone. Tibolone (Organon’s Livial) (Figure 6) is a synthetic steroid approved for treatment of perimenopausal symptoms and prevention of osteoporosis. The

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FIGURE 6. Structure of tibolone.

agent Þrst launched in Germany in 1999 and is available in all European markets. Azko Nobel (Organon’s holding company) is conducting additional Phase III clinical trials for osteoporosis in the United States to address the FDA’s request for additional information about the agent’s labeling for postmenopausal osteoporosis (Azko Nobel, annual report, February 9, 2004). Tibolone is also currently under investigation for treatment of sexual dysfunction in postmenopausal women. An eight-year study of tibolone used an open-label, nonrandomized design in which 110 recently menopausal patients were chosen at baseline to be treated with 2.5 mg tibolone daily or placebo. After eight years, the average increase in BMD in the group receiving tibolone was 4.1% in the lumbar spine and 4.6% in the femoral neck; the placebo group demonstrated a 7.5% loss of BMD in the spine and 6.7% loss in the femoral neck (Rymer J, 2001). The calcium/creatinine urine ratio (a biochemical marker of bone resorption) decreased in the tibolone group but remained constant in the placebo group. Similar results were obtained in a two-year extension of this study. Of the 57 patients who remained in the study during the extension period, those who received tibolone experienced increased BMD of 4.8% in the lumbar spine and 3.7% in the femoral neck. Untreated patients exhibited BMD losses of 8.5% in the lumbar spine and 8.9% in the femoral neck (Rymer J, 2002). Three randomized, placebo-controlled trials of shorter duration (two years each) have demonstrated similar results. The combined results of two identical U.S. studies were published in 2001. The studies enrolled a total of 770 postmenopausal women. In the 519 women who completed the two-year assessment, tibolone increased lumbar spine BMD by 2.6% and hip BMD by 3.2%, compared with placebo (Gallagher J, 2001). A third study of 107 women found 7.2% and 2.6% increases in BMD in the lumbar spine and femoral neck, respectively, in the tibolone-treated group. In comparison, the placebo group demonstrated 0.9% and −1.6% changes in BMD in the spine and femoral neck, respectively (Pavlov P, 1999). It should be noted that none of the studies outlined in this section were designed with sufÞcient statistical power to assess tibolone’s effect on incident fracture. Unlike HRT and SERMs, tibolone does not appear to have a beneÞcial effect on serum lipids. A meta-analysis of trials found that tibolone reduces HDL cholesterol by approximately 34% and has no effect on LDL cholesterol (Modelska

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K, 2002). Common side effects of tibolone included vaginal bleeding in nearly one-third of treated patients. Synthetic Isoflavones Overview. Phytoestrogens extracted from soy products (isoßavones) have long been used as herbal remedies for perimenopausal symptoms because of their structural similarity to human estrogen. Synthetic isoßavones are chemically synthesized derivatives of plant isoßavones. Mechanism of Action. Synthetic isoßavones are structurally similar to human estrogen and are believed to exert the same mechanism of action on bone estrogen receptors as the HRT agents or the synthetic steroids. However, these agents are less potent than human estrogen because of subtle differences in their chemical structure. Ipriflavone. Iprißavone (Takeda’s Iprosten/Osten, generics)∗ is a synthetic derivative of an isoßavone (a family of phytoestrogens extracted from plants) isolated from soy that is approved for treatment of osteoporosis in Japan and Italy. Whereas some studies have shown iprißavone to be effective in slowing BMD loss in osteoporotic women, more-recent trial data have cast doubt on its efÞcacy. Iprißavone is marketed by Takeda as Iprosten in Italy and as Osten in Japan. Generic versions are also available. Two Italian studies found that iprißavone had a modestly beneÞcial effect on BMD. In one study, 453 postmenopausal women with low BMD were randomized to receive either 600 mg iprißavone daily or placebo for two years. At two years, vertebral and radial BMD increased 1.6% and 3.5%, respectively, in the group receiving iprißavone, while the placebo group demonstrated a consistent loss of BMD (Gennari C, 1997). A second study of 198 postmenopausal women utilized the same dosing and length of treatment and yielded similar results: vertebral BMD increased 1.4% after one year of iprißavone treatment and 1% at two years; the placebo group had a signiÞcant decrease in BMD after two years (Agnusdei D, 1997). In both studies, notably, increases in BMD were most pronounced after the Þrst year of treatment and were comparatively smaller by the second year. A more recent trial produced results that contradict those of the two Italian trials. Four hundred and seventy-four postmenopausal women with low BMD were randomized to receive 600 mg iprißavone daily or placebo for four years. After four years of treatment, no signiÞcant difference was seen in annual increases in vertebral BMD between the iprißavone (0.1%) and placebo (0.8%) groups; increases in hip and arm BMD were also similar (Alexandersen P, 2001). Additionally, iprißavone treatment was associated with the development of lymphocytopenia (a reduction in serum CD4 T-cell levels); 12.4% of the iprißavone group developed the condition, compared with 0.4% of the placebo group. On the whole, available data indicate that iprißavone has very modest efÞcacy in preventing or treating osteoporosis.

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EMERGING THERAPIES Most emerging agents for the treatment of osteoporosis have mechanisms of action similar to those of currently available therapies but offer improvements in either convenience or efÞcacy. Available data suggest that the full-length, recombinant human parathyroid hormone (PTH; NPS’ Preos/Preotact) is comparable to teriparatide in both safety and efÞcacy. The new bisphosphonates will continue the trend to improved convenience through less frequent dosing. Two novel agents are in the pipeline as well: strontium ranelate (Servier’s Protelos) and Amgen’s AMG-162. Although strontium ranelate’s bone-building effects have been documented, its mechanism of action remains unknown. AMG162 is a human monoclonal antibody (MAb) that mimics the action of osteoprotegerin, a soluble decoy receptor for the receptor activator of nuclear factor κB ligand (RANKL), which inhibits osteoclast activity. AMG-162 is the only agent in development that has an infrequent dosing schedule and beneÞcial effects that are long lasting without long-term persistence. In addition to the drugs discussed in this section, two other treatment approaches are in development: cathepsin K inhibitors and αv β3 integrin (vitronectin) antagonists. Cathepsin K is a cysteine protease secreted by osteoclasts that destroys the collagen matrix in bone but does not affect the demineralization that precedes this process. Novartis’ AAE-581 is a cathepsin K inhibitor currently in Phase IIb clinical investigation. Merck is developing MRL-123, an αv β3 integrin antagonist that inhibits osteoclast adhesion to bone matrix, thereby interfering with osteoclast activity. A one-year, placebo-controlled trial of 227 women with low lumbar or spine bone mineral density (BMD) found increases in BMD in both areas, but these increases were less than increases provided by most other osteoporosis agents. Also, total body BMD did not rise (Murphy M, 2004). These agents are not discussed further in this section. The most promising agents in development for the treatment of osteoporosis are listed in Table 6. RANKL Inhibitors Overview. RANKL inhibitors mimic the body’s own process for promoting bone growth. Members of this drug class have a short half-life, unlike the longacting bisphosphonates, so “frozen bone” (i.e., near-total suppression of bone turnover) is less likely to result in insufÞcient bone remodeling and prevent healing of minor fractures and other damage. Mechanism of Action. Osteoblasts secrete both osteoprotegerin and receptor activator of nuclear factor κB ligand (RANKL); their ratio determines whether bone formation or resorption predominates. RANKL promotes bone resorption by binding to RANK on osteoclasts, thereby activating them. Osteoprotegerin acts as a decoy receptor for RANKL, preventing it from binding to RANK. Without a signal from RANKL, osteoclasts fail to initiate resorption; the result is net bone formation (Coetzee M, 2004; Onyia JE, 2004). Additionally, RANKL causes

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TABLE 6. Emerging Therapies for the Treatment of Osteopenia and Osteoporosis Compound

Development Phase

RANKL inhibitors AMG-162 United States Europe Japan Parathyroid hormone analogues Preos United States Europe Japan Bisphosphonates Ibandronate, once-monthly oral United States Europe Japan Ibandronate, IV United States Europe Japan Zoledronate United States Europe Japan Minodronate United States Europe Japan Selective estrogen receptor modulators Lasofoxifene United States Europe Japan Bazedoxifene United States Europe Japan Strontium salts Strontium ranelate United States Europe Japan Vitamin D analogues ED-71 United States Europe Japan PR = Preregistered; R = Registered.

Marketing Company

III — —

Amgen — —

III III —

NPS Pharmaceuticals Nycomed —

PR PR I

Roche/GlaxoSmithKline Roche/GlaxoSmithKline Chugai

PR III II

Roche/GlaxoSmithKline Roche/GlaxoSmithKline Chugai

III III —

Novartis Novartis —

— — III

— — Ono

PR III III

Pfizer Pfizer Pfizer

III III —

Wyeth Wyeth —

— R II

— Servier Fujisawa

— — III

— — Chugai

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pre-osteoclasts to develop into osteoclasts (Viereck V, 2003). By inhibiting this pathway, RANKL inhibitors may function by a second mechanism of action. The only late-stage agent in this class, AMG-162, is a fully humanized monoclonal antibody of RANKL that behaves much like osteoprotegerin: it binds to RANKL, thereby preventing RANKL from activating RANK on osteoclasts. AMG-162. Amgen is developing recombinant human osteoprotegerin, an endogenous inhibitor of osteoclast development and activation, for treatment of osteoporosis. Amgen initiated two Phase III clinical trials of AMG-162 in August 2004 (Amgen, press release, August 10, 2004). Limited data from a one-year Phase II trial indicate that AMG-162 may be effective against osteoporosis. The trial enrolled 411 postmenopausal women who were randomized to one of several groups: one injection of AMG-162 (6, 14, or 30 mg) every three months; semiannual injections of AMG-162 (14, 60, 100, or 210 mg); weekly alendronate (70 mg); or placebo. The authors reported that, after one year, lumbar BMD increased by 7% in the group receiving 60 mg AMG-162 semiannually; the alendronate group improved by 5%. Two patients taking AMG-162 experienced vertebral fractures during the study. Because fracture data were not provided for placebo users, AMG-162’s effect on this outcome cannot be compared with the effects of other agents at this time. Adverse events occurred most frequently in the alendronate group: 20% of those patients suffered dyspepsia, whereas 4% and 5% of patients in the placebo and AMG-162 groups, respectively, experienced this side effect (McClung MR, 2004). Parathyroid Hormone Analogues Overview. In addition to NPS’s Preos/Preotact, an injectable form of PTH 184, several companies are developing PTH agents with more favorable modes of administration than teriparatide (Eli Lilly’s Forteo/Forsteo), which requires daily subcutaneous injections. Two oral formulations of PTH are in early development: one by Emisphere Technologies and Eli Lilly, and the other by GlaxoSmithKline and Unigene Laboratories. Preclinical results with Emisphere/Lilly’s oral PTH 1-34 show that serum PTH 1-34 concentrations similar to those that result from subcutaneous injection can be achieved, despite the low bioavailability of the orally administered PTH (Leone-Bay A, 2001). Alza is developing a transdermal patch to deliver PTH 1-34. A Phase I trial of this product found that a patch containing 30 µg of the agent delivered serum levels of PTH 1-34 equivalent to levels delivered by a 40 µg subcutaneous injection (Gopalakrishnan V, 2004). Roche and Suntory are developing SUN-E-3001, an intranasal formulation of PTH 1-34. In a three-month trial of 92 osteoporotic patients, lumbar BMD increased after three months (Matsumoto T, 2004). These alternative formulations would greatly increase patient compliance and encourage the acceptance of PTH for treatment of osteoporosis. However, very few data have been published about these formulations, which are all in the earliest stages of development.

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Mechanism of Action. Parathyroid hormone analogues increase BMD by stimulating osteoblast activity rather than preventing bone resorption by osteoclasts. Daily administration of PTH, which results in the intermittent rise and fall of serum PTH levels, can stimulate osteoblast activity, thereby increasing bone mass. This stimulation stems in part from PTH’s direct inhibition of apoptosis in mature osteoblasts, which extends their life spans (Jilka RL, 1999). Full-Length Recombinant Parathyroid Hormone. NPS Pharmaceuticals’ Preos/Preotact (formerly ALX1-11), an injectable, recombinant, full-length human PTH, will compete directly with Eli Lilly’s teriparatide. A long-term agreement between AstraZeneca and Allelix Biopharmaceuticals to develop and commercialize Preos was dissolved in 1998, when AstraZeneca discontinued all clinical development in Sweden, Canada, and the United States (Allelix Biopharmaceuticals, press release, September 24, 1998). NPS Pharmaceuticals acquired Allelix in 1999 and is developing Preos in the United States. In 2004, Nycomed acquired the rights to develop and market the agent in Europe, where it will be sold under the brand name Preotact. Preos and teriparatide are different forms of the PTH molecule: Preos is the full-length molecule (PTH 1-84), whereas teriparatide (PTH 1-34) encompasses only the Þrst 34 amino acids. However, it is this amino-terminal portion of the molecule that has been shown to be active in bone formation. In addition, the full-length molecule is processed in vivo to the shortened molecule (PTH 1-34). Therefore, Preos is expected to have efÞcacy and compound-attribute proÞles similar to those of teriparatide. Some late-breaking data were presented at the 2004 meeting of the American College of Rheumatology in San Antonio, Texas, and in a company press release (NPS Pharmaceuticals, press release, March 30, 2004). The Treatment of Osteoporosis with Parathyroid Hormone (TOP) study was a multicenter, double-blind trial that enrolled approximately 2,600 women who were randomized to receive daily injections of placebo or 100 µg of Preos for 18 months. Both groups also received daily supplements of vitamin D and calcium. At enrollment, patients’ mean T-scores were −3.0 (spine), −1.9 (hip), and −2.2 (femoral neck), and 19% of patients had a vertebral fracture. After 18 months, patients who took at least 75% of their prescribed doses (n = 1, 870) reduced their risk of vertebral fracture by 66% more than the placebo group: new fracture incidence was 1.1% in the Preos group and 3.3% in the placebo group. Fracture risk reduction for the entire intent-to-treat group was 59%. Only 2.6% of Preos users suffered another vertebral fracture during the study period, compared with 8.4% of patients receiving placebo. Among patients who did not have an initial vertebral fracture, a new fracture occurred during the study in 0.8% of patients in the Preos arm and in 2.2% of patients in the placebo arm (NPS Pharmaceuticals, press release, October 26, 2004). The results from the TOP study are similar to those from the pivotal trial for teriparatide, which demonstrated a 66% reduction in the risk of vertebral fracture (compared with 65% for Preos) in patients with a preexisting vertebral fracture.

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It should be noted that the teriparatide trial enrolled a far higher proportion of patients with baseline vertebral fractures than the Preos trials (90% versus 19%). The efÞcacy of different regimens of Preos and alendronate was assessed in the PTH/alendronate (PaTH) study. Results demonstrated that Preos monotherapy achieved better results than combination Preos/alendronate therapy or treatment with alendronate alone (Black DM, 2003). In the Þrst year of this two-year study, 238 postmenopausal women with low BMD at the spine or hip were randomized to receive 100 µg Preos, 10 mg alendronate, or both agents daily. Patients also received daily calcium and multivitamin supplements. Primary end points were change in bone density, biochemical markers of bone turnover, and fracture incidence. Preos monotherapy effected the greatest increase in volumetric density of trabecular bone (26% increase over baseline, compared with 11% and 13% for alendronate monotherapy and combination therapy, respectively). There were no signiÞcant differences in BMD metrics in other areas, which included hip and femoral neck. A total of eight fractures occurred during the Þrst year of the trial; incidence was similar in all three groups. Likewise, adverse event rates did not differ among the regimens. Side effects seen with Preos therapy were similar to those observed with teriparatide treatment: injection site complications, nausea, fatigue, headache, dizziness, or limb pain. The study authors concluded that the combination regimen provided no more beneÞts than Preos monotherapy. Markers of bone formation and resorption were also examined in the PaTH study. Preos monotherapy showed positive results in measurements of serum N-propeptide and C-terminal telopeptide of type I collagen (CTx), markers of bone formation and resorption respectively. Although bone resorption markers increased in the Preos monotherapy group, their increase was greatly exceeded by the rise in markers of bone formation (150%). This increase was greater than that of the other two groups and suggests net bone formation. In the combination therapy group, levels of the bone formation marker fell below baseline after three months, while levels of the bone resorption marker fell by 50% after one month and thereafter remained stable. Patients treated with alendronate alone exhibited a decrease in markers of both bone resorption and formation. In the second year of the PaTH study, most patients switched regimens. Those who were treated with Preos monotherapy in the Þrst year were randomized to receive either placebo or alendronate during the second year of the study. Patients treated with combination Preos/alendronate therapy or with alendronate monotherapy during the Þrst year of the study were switched to alendronate monotherapy during the second year. The study found that in the Preos-toalendronate group, BMD increased by 12% in the spine, 30% in the trabecular spine, and 5% in the total hip. In the alendronate-to-alendronate group, BMD increased 8% in the spine, 7% in the trabecular spine, and 3% in the total hip. The press release detailing these results did not provide data on patients who were switched from the combination Preos/alendronate regimen to alendronate monotherapy or on patients who were switched from Preos to placebo (NPS Pharmaceuticals, press release, October 4, 2004).

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Thus far, all PTH products available or in development are being scrutinized for the risk of osteosarcoma. An increased risk of osteosarcoma was observed in rats treated with high doses of teriparatide (prescribing information), resulting in a black-box warning for that product. A two-year carcinogenicity study found a similar effect with Preos treatment: rats receiving up to 4.6 times the human dose (10 µg/day) of Preos did not have increased risk of osteosarcoma, but the disease was observed in correlation with higher doses (NPS Pharmaceuticals, press release, October 4, 2004). Teriparatide’s black-box warning states that doses ranging from 3 to 20 times the recommended human dose led to increased risk of osteosarcoma in rats. Unlike the bisphosphonates, which have demonstrated differences between individual agents in either efÞcacy or tolerability, Preos and teriparatide appear to exhibit no substantive differences. European health care systems consider teriparatide too expensive for widespread use, leaving a space in the market for similar, less expensive agents. Bisphosphonates Overview. Bisphosphonates reduce the high bone turnover observed in osteoporosis and osteopenia, an effect that raises BMD and lowers fracture risk. Bisphosphonates are stable pyrophosphate analogues that bind avidly to bone and suppress the activity of bone-resorbing osteoclasts. Their use in the treatment of various bone mineralization disorders (such as Paget’s disease and multiple myeloma of bone) has been well described, and bisphosphonates have become Þrst-line therapy for patients experiencing signiÞcant bone turnover and bone loss. Given the chronic nature of osteoporosis and the relatively poor patient compliance with treatment, one of the greatest remaining unmet needs is for a therapy that requires less frequent administration. Because bisphosphonates have an extremely long half-life once bound to bone, they are promising candidates for intermittent therapy. With alendronate and risedronate already available in once-weekly formulations, the need for new, more potent oral bisphosphonates is debatable. Currently, the greatest need is for a bisphosphonate that would require less frequent administration intervals. However, the long half-lives of these agents have prompted mild unease about the longer-acting formulations in development—even if a patient were to cease therapy, the agent would persist in the body for several months. Intravenous (IV) administration of bisphosphonates is currently limited to off-label use of clodronate, pamidronate, and zoledronate, which must be administered in a slow infusion that can last for up to two hours. However, fracture data are insufÞcient to support these agents’ use for osteoporosis. Nevertheless, some physicians prescribe IV bisphosphonates to patients (especially the elderly) who complain of esophageal irritation with available bisphosphonates or refuse to comply with therapy because of the required fasting before and after administration.

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Mechanism of Action. The bisphosphonates examined in this sectuib (and alendronate and risedronate) are known as nitrogen-containing bisphosphonates. Their activity is somewhat different from and more potent than that of Þrstgeneration bisphosphonates such as etidronate and clodronate (Rogers MJ, 2003). Nitrogen-containing bisphosphonates are analogues of isoprenoid diphosphate lipids, which act within osteoclasts to inhibit the enzyme farnesyl pyrophosphate (FPP) synthase. FPP synthase is a required enzyme in the cholesterol biosynthesis pathway; inhibition of this enzyme induces apoptosis of osteoclasts and inhibits the formation of new osteoclasts. Ibandronate. Ibandronate (Roche/GlaxoSmithKline’s Bonviva/Boniva)∗ has been approved in several European countries for treatment of hypercalcemia associated with malignancy. A once-daily oral formulation was approved in the United States and Europe in 2003 for the prevention and treatment of osteoporosis (IDdb3, accessed February 2004). Ibandronate launched in early 2005. In December 2001, Roche and GlaxoSmithKline announced that they had entered a codevelopment and copromotion agreement for ibandronate in all world markets except Japan, where Chugai (Roche’s local subsidiary) is conducting Phase II trials for IV ibandronate and Phase I trials for once-monthly oral ibandronate for treatment of osteoporosis. A one-year, double-blind trial (the Dosing Intravenous Administration [DIVA] study) involving 1,395 postmenopausal women with osteoporosis found that three regimens of ibandronate (2 mg injected every two months [q2m], 3 mg injected every three months [q3m], and 2.5 mg orally daily) with calcium and vitamin D supplements effected similar increases in BMD. The oral regimen achieved a 3.8% increase in lumbar BMD over baseline. Fracture data were not provided, but BMD results for the IV formulations were even better than for the oral regimen; in the 3 mg q3m arm, lumbar BMD rose by 4.8%, and 2 mg q2m increased lumbar BMD by 5.1% (Recker R, 2004). These results suggest that the IV formulations may also provide superior protection against lumbar vertebral fracture, compared with daily oral ibandronate. In an earlier three-year trial, the daily oral regimen was found to reduce risk of vertebral fracture by 62% (Chesnut III CH, 2004). No evidence of renal toxicity and low incidence of other side effects was reported in the DIVA study. A 629-patient, placebo-controlled study found a similar trend. Patients received one injection of ibandronate (2 mg, 1 mg, or 0.5 mg) every three months (q3m) or placebo. All patients also received calcium supplements. In the 2 mg q3m group, BMD rose by 2.5%, versus a 2.4% decrease for placebo. BMD increased by 1.8% and 1.0% respectively in the 1 mg and 0.5 mg groups. All regimens appeared to be well tolerated (Stakkestad JA, 2003). A small (14-patient) openlabel study in men found that two years of ibandronate treatment (2 mg q3m) increased lumbar spine BMD by 3.2% (Lamy O, 2003). Preliminary one-year results have been released from the Monthly Oral Ibandronate in Ladies (MOBILE) study, which examined monthly (q1m, 100 mg or 150 mg) and daily (2.5 mg) oral ibandronate in 1,609 postmenopausal women.

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269

The authors found that one year of treatment yielded results similar to the results achieved by the q2m and q3m IV formulations in the DIVA study: lumbar spine BMD increased by 4.9% in the 150 mg q1m arm; the 100 mg q1m dose had less efÞcacy. Furthermore, the authors state that the 150 mg q1m results are superior to those effected by the 2.5 mg daily regimen, but details were not provided (Miller PD, 2004). Side effects and adverse events were not discussed. Zoledronate. Zoledronate (Novartis’s Zometa)∗ is marketed for tumor-induced hypercalcemia in Europe and the United States. In 2002, it received FDA approval for treatment of bone metastases. It is also in Phase III development in the United States and Europe for osteoporosis treatment. A Phase IIa trial for a transdermal formulation of zoledronate is under way; further data on this formulation are not available. Positive Phase II results with a once-yearly formulation of zoledronate have been demonstrated. A one-year, randomized, placebo-controlled clinical trial enrolled 351 postmenopausal women with osteoporosis (BMD more than 2.5 standard deviations (SD) below the normal young adult mean [i.e., T-score < −2.5]). Researchers observed signiÞcant increases in spine BMD (ranging from 4.3% to 5.1%) and hip BMD (ranging from 3.1% to 3.6%) in patients who were given zoledronate intravenously every three months (0.25, 0.5, and 1.0 mg), twice yearly (2.0 mg), or once yearly (4.0 mg) (Reid IR, 2001). In addition, bone turnover markers decreased by 40–50%—an indication that zoledronate markedly inhibits bone turnover. These results are similar to those achieved by other bisphosphonates administered daily or weekly that have proved effective in preventing fractures. Treatment-related adverse events (musculoskeletal pain, nausea, fever), although mild, were signiÞcantly higher in patients receiving zoledronate (47–67%) than in patients receiving placebo (27%). Most side effects were associated with the Þrst treatment dose. Novartis is undertaking a series of trials known as the Health Outcomes and Reduced Incidence with Zoledronic Acid Once Yearly (HORIZON) studies. More than 10,000 patients are expected to participate worldwide. The study is currently enrolling patients who have either osteoporosis or Paget’s disease. The threeyear study will examine the performance of an annual dose of IV zoledronate; primary end points include spine and hip fracture incidence. One study within HORIZON, the HORIZON Recurrent Fracture Trial, will test the zoledronate annual-dose regimen in men and women who have had recent hip fractures. This trial will be conducted in the United States and will enroll approximately 2,500 patients (Novartis, press release, November 14, 2004). Minodronate. In January 1999, Ono Pharmaceutical acquired from Yamanouchi the rights to develop and market minodronate∗ in Japan for osteoporosis, hypercalcemia, and bone metastases stemming from breast cancer. As of March 2002, minodronate was reported to be in Phase III development; however, no trial data have been released.

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Minodronate is not expected to have efÞcacy superior to that of alendronate or risedronate, agents that are already marketed in Japan. Additionally, minodronate is not being developed outside of Japan. Selective Estrogen Receptor Modulators Overview. Raloxifene is the only selective estrogen receptor modulator (SERM) currently on the market for osteoporosis. Its tendency to increase the risk of thromboembolism and induce hot ßashes is considered a distinct disadvantage, however. For this reason, its use is generally limited to postmenopausal women who have low bone mass and no vasomotor symptoms and are concerned about breast cancer risk. Therefore, raloxifene has beneÞted little from the results of the Women’s Health Initiative (WHI) study, even though many women abandoned HRT in its wake. Whether the new SERMs under development will possess characteristics that make them a more suitable substitute for HRT remains to be seen. In addition to the agents discussed in the following subsections, two other drugs are in Phase II development for osteoporosis: Eli Lilly’s arzoxifene (also in Phase III for breast cancer) and Hormos Medical’s ospemifene. No data are available on the effects of either drug on fracture incidence or BMD. In a trial of 159 healthy postmenopausal women, ospemifene was shown to reduce bone turnover (Komi J, 2004). However, the implications of this result for fracture incidence are unclear. Mechanism of Action. The estrogen receptor is a nuclear transcription factor that activates genes containing estrogen response elements. SERMS used to treat osteoporosis appear to function by stimulating subtypes of the estrogen receptor that promote production of osteoprotegerin, a soluble decoy receptor for RANKL. Occlusion of RANKL inhibits the activation of RANK, thereby preventing the development of osteoclasts from precursor cells (Viereck V, 2003). RANKL also binds to osteoclasts, causing them to initiate resorption. As SERMs promote osteoprotegerin production, they also block this effect of RANKL (Coetzee M, 2004; Onyia JE, 2004). SERMs have been shown to have little or no effect on reproductive tissues. The apparent speciÞcity of the SERMs may be due to the wide variety of estrogen receptors, some of which may bind only weakly to SERMs (Stepan JJ, 2003). Lasofoxifene. In 1991, PÞzer and Ligand Pharmaceuticals formed a strategic alliance to use intracellular receptor technology to identify SERMs for the prevention and treatment of osteoporosis. The research collaboration led to the development of lasofoxifene (CP-336156), a potent SERM analogue of droloxifene and raloxifene. In December 1999, PÞzer discontinued droloxifene’s development in favor of lasofoxifene. Lasofoxifene is preregistered in the United States and is in Phase III clinical trials in Europe and Japan. In August 2004, PÞzer Þled with the FDA for approval of lasofoxifene for prevention of osteoporosis (Ligand Pharmaceuticals, press release, September 16, 2004), triggering a milestone payment

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to Ligand. Ligand has also signed an agreement with Royalty Pharma in which the latter has purchased rights to a share of Ligand’s future royalty payments from late-stage SERM products, including lasofoxifene. A two-year Phase II study that enrolled 410 postmenopausal women compared lasofoxifene (0.25 mg or 1.0 mg per day) with raloxifene (60 mg per day) and placebo. Lumbar spine BMD increased by 1.8% and 2.2% in the lasofoxifene groups (0.25 and 1.0 mg, respectively). Meanwhile, lumbar spine BMD declined by 0.1% in raloxifene patients and by 1.7% in placebo patients. Total hip BMD metrics were similar in the treatment groups: lasofoxifene (1.0 mg and 0.5 mg) and raloxifene (60 mg) increased hip BMD by 1.3%, 1.9%, and 1.5%, respectively. In the placebo group, total hip BMD fell by 0.1% (McClung M, 2004). Although it is unclear whether lasofoxifene offers greater fracture protection than raloxifene, lasofoxifene appears to have other beneÞcial effects. In a twoyear Phase II study, lasofoxifene decreased low-density lipoprotein (LDL) cholesterol levels in the blood: patients who received 0.5 mg daily had a 21.1% reduction in LDL levels, versus a 6.1% reduction for patients who received placebo. No meaningful changes in the endometrium were observed during the study, and no cases of breast cancer were reported (Moffett AH, 2004). In another Phase II study, lasofoxifene was as effective as conjugated estrogens/medroxyprogesterone and signiÞcantly more effective than raloxifene in increasing BMD. Lasofoxifene also caused a greater reduction in LDL cholesterol (25%) than raloxifene (16%) (“PÞzer expects . . . ,” The Pink Sheet, November 22, 1999). Additional clinical studies found lasofoxifene comparable to tamoxifen in preventing breast cancer. Available data indicate that lasofoxifene is more effective than raloxifene in increasing BMD at the lumbar spine. However, direct comparisons of efÞcacy cannot be made until fracture data become available. Bazedoxifene. Bazedoxifene (TSE-424)∗ is a tissue-selective SERM being developed for treatment of osteoporosis and osteopenia through a collaborative agreement between Wyeth and Ligand Pharmaceuticals. Under the terms of the agreement, Wyeth will pay Ligand milestone fees and royalties on the commercialized product. This agent is in Phase III development in the United States and Europe, but it is not currently under development in Japan. Various Phase III trials involving bazedoxifene are under way: a comparison of bazedoxifene with raloxifene for osteoporosis prevention (1,700 patients); a trial for osteoporosis treatment (7,600 patients); and a comparison of bazedoxifene plus Premarin (Wyeth’s conjugated equine estrogen) with raloxifene for osteoporosis and uterine safety (3,500 patients). Results are expected in 2006 and 2007. Phase II trial results were presented at a company meeting in January 2001 (IDdb3, accessed February 22, 2005). Positive changes in bone markers were noted, with no estrogenic effect on breast or endometrial tissue. Similar data were presented at a 2004 company meeting. No information is available on this drug’s effect on BMD or fractures.

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Of the SERMs in late-stage development, bazedoxifene may come closest to offering an ideal HRT proÞle. In preclinical studies, bazedoxifene was found to be a more potent antiresorptive agent than raloxifene, exhibiting bone-strengthening effects without stimulating uterine tissues (Komm B, 2002). Furthermore, 107 perimenopausal women (mean age 55 years) enrolled in a Phase I clinical trial of bazedoxifene exhibited no vasomotor symptoms after 30 days of treatment (Ermer JC, 2000). Strontium Salts Overview. Strontium ranelate (Servier’s Protelos) is the sole member of the strontium salt class. Elemental strontium was Þrst used as a therapy for osteoporosis more than 50 years ago. Development efforts were abandoned because the drug led to mineralization defects in patients with impaired renal function (D’Haese PC, 2004). Such effects have not been observed with strontium ranelate in patients with normal renal function (Meunier PJ, 2004). Mechanism of Action. Strontium ranelate is the Þrst compound reported to uncouple the processes of bone resorption and formation: it inhibits bone resorption and stimulates bone formation (Buehler J, 2001; Marie PJ, 1993). Other emerging osteoporosis therapies demonstrate only one of the two effects. Beyond this, strontium ranelate’s mechanism of action is completely unknown. Strontium Ranelate. Servier launched this agent in the United Kingdom and Germany at the end of 2004. Strontium ranelate∗ is in Phase II development in Japan, where, in May 2001, Fujisawa licensed the rights to develop, manufacture, and market this agent. No agreements appear to have been concluded for development or marketing in the United States. Of the 1,649 postmenopausal women participating in the Phase III Spinal Osteoporosis Therapeutic Intervention (SOTI) trial, 87.5% had at least one prevalent vertebral fracture at the time of enrollment; these patients became the intentto-treat population. Over the three-year study period, the relative risk of new vertebral fracture was 41% lower in patients who received 2 g oral suspension of strontium ranelate daily than in patients who received placebo. At one year, the risk reduction was 49% in the strontium ranelate group (Meunier PJ, 2004). The incidence of fracture at three years was 21% in the strontium ranelate group and 33% in the placebo group, a reduction in fracture risk of 41% in patients treated with strontium ranelate; at one year, fracture incidence was 6% and 12% respectively, a reduction of 49%. However, this agent showed relatively little effect against nonvertebral fractures: they occurred in 15% of strontium ranelate patients and 17% of placebo patients. BMD in patients taking strontium ranelate in the SOTI trial increased over baseline by 13% at the lumbar spine, 7% at the femoral neck, and 9% at the total hip. In contrast, BMD scores in all three areas fell by 1% in placebo users. However, because strontium is a heavy element, the increase in BMD observed with

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strontium ranelate treatment overstates the percentage increase in BMD owing to increased mineralization or bone formation. Adjusting for this effect, the authors state that the increase in lumbar spine BMD was only 6.8% after three years. Adverse events occurred at statistically similar rates in both groups. Diarrhea was the most frequently observed side effect, occurring in 6% of strontium ranelate patients and 4% of patients taking placebo (Meunier PJ, 2004). Although strontium ranelate may offer a more convenient mode of administration than injectable PTH, it is less effective; similar efÞcacy can be obtained with other agents, such as the once-weekly bisphosphonates. The Treatment of Peripheral Osteoporosis study (TROPOS), which enrolled 5,091 postmenopausal women (average age 76), investigated the effects of strontium ranelate treatment on nonvertebral fracture risk. Patients were given daily supplements of vitamin D and calcium and randomized to receive either 2 g/day strontium ranelate or placebo over a period of three years. In this study, strontium ranelate effected a 41% greater reduction in hip fracture than placebo in compliant patients and a 16% greater reduction in peripheral fracture risk than placebo. In comparison, the SOTI study found an 11% reduction in total nonvertebral fracture risk (incidence in 15% of strontium ranelate patients and 17% of placebo users). However, because fracture risk was measured only in compliant patients in the TROPOS study, the results may be overstated. Although a better result than in the SOTI study, the 16% reduction in peripheral fracture risk in the TROPOS study does not outperform bisphosphonates. Among women older than 74 years who had very low bone density (n = 1, 977), strontium ranelate was able to reduce the risk of hip fracture by 36% more than placebo. This study did not assess the drug’s effect on vertebral fractures (Spector TD, 2004). Two randomized, placebo-controlled, dose-Þnding studies were conducted to examine strontium ranelate’s ability to increase lumbar BMD: one study (Strontium Administration for Treatment of Osteoporosis Study [STRATOS]) was conducted in postmenopausal women who had at least one vertebral fracture; the other study (Prevention of Osteoporosis Study [PREVOS]) was conducted in early postmenopausal women. In the STRATOS study, 353 women received either placebo or strontium ranelate (0.5, 1, or 2 g) daily for two years (Meunier PJ, 2002[b]). At the end of the study period, the increase in lumbar BMD was dosedependent—from 1.4% for the 0.5 g dose to 3.0% for the 2 g dose. However, these results are not adjusted for the effect that strontium (a heavy element) has on BMD measurements. Similar increases in lumbar BMD were reported from the PREVOS study, which involved 160 postmenopausal women who were randomized to receive placebo or strontium ranelate (0.125 , 0.5, or 1 g) daily (Reginster JY, 2002). After two years, adjusted lumbar BMD increased 2.4% in patients taking 1 g strontium ranelate compared with patients taking placebo. Lower doses of strontium ranelate did not signiÞcantly increase BMD when compared with placebo. These increases in BMD are similar to those reported for raloxifene but lower than those reported for either alendronate or teriparatide. Patients in these studies tolerated treatment with strontium ranelate well.

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Vitamin D Analogues Overview. Vitamin D is generated endogenously when the skin is exposed to sunlight. It is then metabolized by the liver and kidneys to its active analogue, calcitriol. Calcitriol, marketed for osteoporosis treatment in Japan, reduces bone resorption, but under certain circumstances it may also suppress bone formation by raising intestinal calcium absorption. Drug development in this class has focused on creating bone-speciÞc vitamin D analogues; to increase bone strength, an agent would have to act on osteoclasts without promoting excessive intestinal calcium resorption—a goal not yet met by any agent. Mechanism of Action. Vitamin D analogues act by binding to the nuclear vitamin D receptor (VDR) in osteoblasts (the receptor is also present in other cells). Studies in ovariectomized rats suggest that these agents derive their effect by suppressing PTH, which in turn reduces bone resorption by osteoclasts. PTH suppression appears to result from higher circulating levels of calcium due to increased intestinal absorption, the primary action of vitamin D analogues. However, the reduction in resorption is accompanied by a general decrease in bone turnover. If bone formation decreases to a great extent, vitamin D analogues may actually bring about a net decrease in BMD (Erben RG, 2001). Furthermore, the high levels of circulating calcium required to suppress bone resorption can lead to hypercalcemia and hypercalciuria (excessive calcium in the blood and urine) if vitamin D levels are too high. Conversely, if levels are too low, osteoclasts are stimulated, leading to bone loss. Thus far, all agents in this class have offered a narrow therapeutic window of effective doses (Uchiyama Y, 2002). This drug class may also have anabolic effects. Through an unknown mechanism, these agents act directly on the bone to promote osteogenesis by osteoblasts. If a tissue-speciÞc vitamin D analogue targeted bone without affecting intestinal calcium absorption, bone formation would presumably occur and improve osteoporosis without the risk of calcium-related side effects (Erben RG, 2001). However, no agent in development appears to act with this level of speciÞcity. ED-71. Chugai initiated Phase III trials of its vitamin D3 agonist, ED-71∗ , in October 2004. Currently, development is limited to the Japanese market. In a Phase II trial, 108 patients (101 women and 7 men) with osteoporosis of unspeciÞed etiology were randomized to receive ED-71 orally in doses ranging from 0.25 µg to 1.0 µg per day. The authors state that after six months, BMD rose dose-dependently in all groups; the study did not provide baseline BMD. However, the authors state that BMD rose by 3% or more in 46% of patients taking the highest dose. Of the patients taking the lowest dose, 22% experienced a 3% or greater increase in BMD. Notably, the 0.75 µg group had the highest proportion of patients (54.2%) with a 3% or more increase in BMD. Bone turnover markers suggest that this agent promotes bone formation while suppressing bone resorption. No hypercalcemia was observed, but the authors do not provide speciÞc data about serum levels (Matsumoto T, 2004).

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Type 1 Diabetes

ETIOLOGY AND PATHOPHYSIOLOGY Introduction Type 1 diabetes is a metabolic disorder in which the insulin-secreting beta cells of the pancreas are destroyed, leading to complete insulin deÞciency. Insulin—an endocrine hormone that is secreted by the beta cells of the pancreatic islets of Langerhans in response to elevated blood levels of glucose, fatty acids, ketone bodies, and amino acids—has two key functions in maintaining health: it signals the body that it has been fed, and it promotes efÞcient storage and consumption of energy sources and nutrients. It carries out the latter function by controlling the transport of glucose, amino acids, and fatty acids across cellular membranes. Insulin also promotes the synthesis of several important substances, including glycogen (the form in which glucose is stored in the liver), proteins, and lipids. In patients with type 1 diabetes, the body, robbed of its ability to produce insulin, feeds off stores of fats and proteins. By-products of this destructive metabolism by the liver of fatty acids and carbohydrates include organic compounds known as ketone bodies. These substances (beta-hydroxybutyric acid, acetoacetic acid, and acetone) accumulate in ßuids and tissues, eventually leading to a state of absolute or partial insulin deÞciency known as ketoacidosis, one of the complications of type 1 diabetes that make this disease a serious lifelong condition requiring close monitoring and chronic treatment. Etiology The etiology of type 1 diabetes remains largely unknown. Most recent data support the notion that the T-cell-mediated autoimmune response observed in type 1 Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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diabetes is caused by an interplay between numerous susceptibility genes and environmental modiÞers. Genetic Factors. Epidemiological evidence shows that the offspring and siblings of type 1 patients are at increased risk for type 1 diabetes; approximately 5–10% of Þrst-degree relatives of type 1 patients will develop disease (Genuth S, 2001). Genetic research demonstrates that type 1 diabetes is a heterogeneous and polygenic disorder, linking several sites—ranging from 11 to more than 18 different genes and at least 20 chromosomal regions—to increased risk of type 1 diabetes (Atkinson MA, 2001). The primary genes associated with type 1 diabetes are located in the major histocompatibility complex (MHC) locus on chromosome 6 (known as IDDM1 ), where the genes for human leukocyte antigens (HLA) reside. Individuals with these polymorphisms carry a 50% risk of developing type 1 diabetes (Genuth S, 2001). Among whites in the United States, type 1 diabetes is 20 times more prevalent in heterozygous individuals with HLA types DR3 and DR4 than in the general population (Genuth S, 2001). The genes encoded in this region are involved in the immunological role of antigenic presentation, but their speciÞc role in type 1 diabetes pathogenesis remains unclear. A second gene for type 1 diabetes has been isolated to chromosome 11 (known as IDDM2 ) in the region of genes that encode for the production of insulin and insulin-like growth factor. The IDDM2 locus contributes 10% toward genetic susceptibility (Atkinson MA, 2001). Another locus, IDDM12, has been identiÞed on chromosome 2. This region encodes a gene that acts on cytotoxic T-lymphocyte-associated protein 4 (CTLA-4), a mediator of immune signaling (Atkinson MA, 2001). Environmental Factors. The observation that only 30–50% of identical twin siblings of type 1 diabetic patients eventually develop diabetes suggests that environmental factors, in concert with genetic predisposition, play an important role in the development of type 1 diabetes (Genuth S, 2001). In the traditional view, environmental factors trigger the disease in genetically susceptible individuals. The modern hypothesis postulates that multiple exposures to environmental factors inßuence the immune dysregulation observed in type 1 diabetics (Atkinson MA, 2001). Putative environmental factors that induce the development of disease include viruses, early infant diet, and exposure to toxins. The viral infection most associated with type 1 diabetes is rubella. A high prevalence of diabetes exists among children whose mothers were infected with rubella during pregnancy. Coxsackievirus B has also been implicated in the development of type 1 diabetes; antibodies to this virus have been detected in pregnant mothers whose offspring developed type 1 diabetes (Dahlquist G, 1998). One highly controversial hypothesis holds that early exposure to certain dietary proteins—such as those found in infant formulas based on cow’s milk—can provoke an autoimmune reaction in children who begin ingesting these proteins before their gastrointestinal systems have fully matured. An epidemiological

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study conducted in Scandinavia concluded that the incidence of type 1 diabetes was lower among children who were born when breast-feeding was a common practice than among children who were born when formula-feeding was popular (Dahlquist G, 1998). Another dietary factor implicated in type 1 diabetes pathogenesis is nitrosamine; nitrosamine and nitrites are found (at low levels) in consumables such as cured meats, tobacco, beer, and preservatives. Animal studies have shown that nitrosamine compounds are toxic to pancreatic islet beta cells. Administration of streptozotocin, a nitrosourea compound, in low, multiple doses induces insulitis (inßammation of the pancreas) in mice. Several studies have demonstrated that the intake of food containing nitrosamine and nitrites is associated with increased risk of type 1 diabetes (Dahlquist G, 1998). Pathophysiology The course of type 1 diabetes is highly variable, often occurring rapidly in infants and children (evolving within only a few months) but slowly in adults. In some people, primarily adults, the autoimmune process that leads to clearly evident disease may take seven years from the initial production of antibodies to the decimation of the beta-cell population. Figure 1 depicts the clinical progression to type 1 diabetes. Symptoms of type 1 diabetes, such as excessive thirst, frequent urination, and extreme fatigue, appear late in the disease process, after approximately 90% of the pancreatic beta cells have been destroyed. The primary symptoms of type 1 diabetes are hyperglycemia (abnormally increased levels of blood glucose) and ketoacidosis (a potentially fatal imbalance of the body’s acid-base mechanisms). Ketoacidosis may be the Þrst manifestation of the disease in some children and adolescents; other juvenile patients exhibit a small to moderate degree of fasting

FIGURE 1. Clinical progression to type 1 diabetes.

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hyperglycemia that may convert rapidly to ketoacidosis and/or hyperglycemia following an immune challenge, such as an infection. Clinical onset of type 1 diabetes occurs most commonly before the age of 25, but it has been seen in patients as young as nine months and in those in their eighth or ninth decades of life, according to the American Diabetes Association (ADA). Eventually, all patients with type 1 diabetes require exogenous insulin. Chronic hyperglycemia is primarily responsible for the complications of diabetes, which include renal disease, cardiovascular abnormalities, retinal changes that can lead to blindness, and peripheral neuropathy. Early Immune Dysregulation. In people who go on to develop type 1 diabetes, development of anti-islet autoantibodies most often occurs between nine months and three years of age. Most type 1 diabetic patients will exhibit multiple autoantibodies: insulin autoantibodies (IAAs) are usually the Þrst autoantibodies to appear in infants, followed by glutamic acid decarboxylase autoantibodies (GADAs). Patients can also demonstrate autoantibodies against carboxypeptidase H, IA-2 (a protein tyrosine phosphatase) and against the islet cell itself. Although they are detected in diabetic patients, the role of autoantibodies in the pathogenesis of type 1 diabetes remains unclear (Atkinson MA, 2001). Some in vitro data indicate that autoantibodies can inhibit insulin secretion and lyse beta cells (Genuth S, 2001). Insulitis and Destruction of Pancreatic Beta Cells. The detection of autoantibodies (e.g., IAAs, GADAs, IA-2) suggests that some factor triggered the autoimmune response and the cascade leading to insulitis. Histology of the diabetic pancreas shows inÞltration by inßammatory immune cells, a condition known as insulitis. ScientiÞc evidence suggests that resident macrophages, also considered to be antigen-presenting cells (APCs), initiate insulitis and the subsequent destruction of pancreatic islet beta cells. Figure 2 illustrates the course of the autoimmune response leading to the destruction of pancreatic beta cells. Table 1 summarizes the various cytokines involved in the pathogenesis of type 1 diabetes. In the normal state, resident macrophages continuously scavenge for peptides within the islet microenvironment. An as-yet-unidentiÞed anomaly transforms the resident macrophages into inßammatory activators and mobilizes macrophages from the periphery into the islet (Bottino R, 2003). These activated cells release the cytokines interleukin-12 (IL-12) and IL-18. Experiments in non-obese diabetic (NOD) mice reveal that IL-12 accelerates the onset of type 1 diabetes and induces the differentiation of CD4+ T cells into TH 1 cells, which are effectors of the autoimmune response (Sharma A, 2003). High levels of IL-18 have been detected in high-risk individuals, and studies suggest that IL-18 may also activate T cells (Sharma A, 2003). The resident macrophages migrate out of the islets and into the pancreatic lymph node, where they present beta-cell autoantigens to autoreactive CD4+ T cells (those that escaped thymic deletion during fetal development). The activated CD4+ T cells release the cytokines IFN-gamma and IL-2. In vivo data show that

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FIGURE 2. Autoimmunity in type 1 diabetes.

IFN-gamma causes severe lymphocytic inÞltration of islets, promoting extensive beta-cell injury. Both of these cytokines activate CD8+ cytotoxic T cells—the Þnal effectors of beta-cell destruction (Sharma A, 2003). In addition to effecting beta-cell toxicity, IFN-gamma induces the activation of macrophages, thus triggering the release of IL-1-beta, a cytokine that

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TABLE 1. Selected Cytokines Involved in the Pathogenesis of Type 1 Diabetes Cytokine Interleukin-12 Interleukin-18

Interferon (IFN)-gamma

Interleukin-2

Interleukin-1/ interleukin-1-beta Tumor necrosis factor (TNF)-alpha

Role in Pathogenesis Secreted by antigen-presenting macrophages. Thought to be an important mediator of beta cell destruction. Secreted by antigen-presenting macrophages. High levels of IL-18 have been detected in individuals at high risk for developing type 1 diabetes. Thought to be a mediator of beta cell destruction in the absence of IL-12. Secreted by CD4+ T cells, CD8+ T cells, and activated macrophages. Activates cytotoxic CD8+ T cells and macrophages, which are putative final effectors of beta cell destruction. Upon activation, these two types of immune cells become major sources of IFN-gamma. Secreted by CD4+ T cells. Its role in the pathogenesis of type 1 diabetes is unclear. Blocking IL-2 in nonobese diabetic (NOD) mice suppresses insulitis and incidence of diabetes. However, other data show that exogenous administration of IL-2 in NOD mice decreases the incidence of diabetes. Secreted by activated macrophages. Damages islet cell DNA and inhibits mitochondrial energy production in pancreatic islet cells. Secreted by activated macrophages. Some data exist that support a role for TNF-alpha; it may potentiate the effect of IL-1.

Source: Based on Sharma A, et al. Autoimmune perspective of insulin-dependent diabetes mellitus: cytokines as therapeutic targets. Drugs of the Future. 2003;28(1):31–42.

inhibits mitochondrial energy production in islet cells. IL-1-beta has been shown to accelerate the beta-cell destructive process in rat models (Dahlquist G, 1998). Activated macrophages also secrete TNF-alpha, which appears in elevated levels in prediabetic NOD mice compared with controls (Sharma A, 2003). The cytokines IFN-gamma, TNF-alpha, and IL-1-beta impair beta cells by inducing nitric oxide (NO)-mediated cytotoxicity. NO damages nuclear DNA and reduces oxidative metabolism and adenosine triphosphate (ATP) formation in the islet-cell mitochondria of rodent models. Another mechanism by which cytokines induce cell death involves the activation of the Fas receptor on islet beta cells. The Fas ligand has been detected on islet-inÞltrating macrophages, suggesting that these immune cells trigger Fasmediated apoptosis in inßamed islets (Atkinson MA, 2001; Devendra D, 2003; Sharma A, 2003). Insulin Deficiency and Clinical Manifestation. Destruction of islet beta cells leads to the loss of insulin secretion, which in turn impairs glucose uptake by target tissues (e.g., muscle, adipose) and suppresses hepatic glucose output. Hyperglycemia ensues, and, ultimately, clinical symptoms of type 1 diabetes

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FIGURE 3. Clinical symptoms resulting from insulin deficiency.

manifest. Figure 3 depicts the various clinical symptoms resulting from the deÞciency of insulin. Increased plasma glucose results in the elevation of the glucose load into the renal tubules, causing osmotic diuresis and presenting as polyuria (i.e., passage of large urine volume within a given period of time) and compensatory polydipsia (chronic excessive thirst). Insulin deÞciency also leads to increased lipolysis (breakdown of lipids in adipose tissue) that results in high plasma lipid levels, elevated hepatic uptake, elevated metabolism of free fatty acids, and ketosis (metabolism of free fatty acids and formation of ketone bodies). Excessive accumulation of the ketone bodies produced during ketosis causes an imbalance in metabolism, and the resulting diabetic ketoacidosis can lead to coma. Insulin deÞciency also causes increased degradation of muscle tissue that clinically presents as weight loss and fatigue. Insulin Resistance. Insulin resistance, a condition in which the body does not respond effectively to insulin, is considered to be the central factor in the development of type 2 diabetes. However, the contribution of insulin resistance to the development of type 1 diabetes is a concept that has yet to be studied extensively. To date, only indirect evidence suggests that insulin resistance plays a role in type 1 diabetes. Some researchers speculate that insulin resistance explains

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why only a portion of islet-antibody-positive individuals develop overt disease. In addition, investigators note that the incidence of type 1 diabetes is highest during puberty—a time during which insulin sensitivity declines (Greenbaum CJ, 2002). Hyperglycemia and Diabetic Complications. Elevated blood glucose levels are the cause of several microvascular diabetic complications, as depicted in Figure 4: retinopathy (which may lead to blindness), nephropathy (which may lead to end-stage renal disease [ESRD]), and neuropathy (which may lead to the need for limb amputation). The deposition and accumulation of advanced glycosylated end products (AGEs) in tissues are a biochemical result of hyperglycemia that, in turn, leads to diabetic complications. Under hyperglycemic conditions, glycosylation of proteins becomes irreversible and makes the proteins resist proteolytic degradation. AGE toxicity impairs multiple biological processes, including neurotransmission,

FIGURE 4. Pathways of hyperglycemia-induced damage.

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wound healing, and blood ßow, thus giving rise to diabetic complications (Friedman E, 1999). In tissues with insulin-independent glucose transport (e.g., nerves, lenses, kidneys, blood vessels), hyperglycemia disturbs the polyol pathway, resulting in various diabetic complications. Aldose reductase converts intracellular glucose into sorbitol and, subsequently, fructose. Elevated glucose levels lead to the accumulation of sorbitol and fructose, resulting in greater intracellular osmolarity, which leads to an inßux of water and osmotic cell injury. Retinopathy. Diabetic retinopathy is best described as capillary microangiopathy that involves the loss of pericytes and endothelial cells with changes in basement membrane thickness. This condition results in microinfarctions, exudates, retinal edema, and neovascularization—all of which can progress to vitreous hemorrhage, traction, and retinal detachment, causing severe loss of vision. Diabetic retinopathy can be classiÞed by severity into a Þrst stage of nonproliferative retinopathy and a later, progressive stage of proliferative retinopathy. Uncontrolled proliferative retinopathy can progress to the point where complications lead to severe visual acuity loss and/or blindness. Nephropathy. Diabetic nephropathy is a multistage condition that often becomes clinically overt several years after diabetes onset. There are Þve stages: • • • • •

Glomerular hyperÞltration. Microalbuminuria/subclinical nephropathy (albumin excretion rate of 20–200 mg/minute). Macroalbuminuria/overt nephropathy (albumin excretion rate greater than 200 mg/minute, accompanied by hypertension). Advanced nephropathy (glomerular Þltration rate less than 75 mL/minute and symptoms of uremia). End-stage renal disease (ESRD) requiring dialysis or renal transplantation.

Neuropathy. Diabetic neuropathy, one of the most common long-term complications of diabetes, is classiÞed into three major categories: • • •

Peripheral, which causes tingling, numbness, and often intense pain in the legs and feet. Focal, which causes third-nerve palsy and blurred vision. Autonomic, which can cause severe gastroparesis (stomach-emptying problems), orthostatic hypotension, and impotence.

The numbness caused by peripheral neuropathy has several serious consequences. Autonomic neuropathy can lead to sudden death from an unfelt, “silent” heart attack. A patient with peripheral neuropathy is liable to develop neuropathic foot ulcers caused by small-Þber sensory dysfunction. Because these chronic ulcers heal slowly and are often untreated, they may lead to infection and ultimately to amputation.

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Cardiovascular Disease. Diabetic patients face increased risk of atherosclerosis and cardiovascular complications compared with nondiabetics, although the underlying pathology of this relationship is not well understood. Cardiovascular disease that eventually produces cardiac ischemia is the primary cause of death among people with type 1 diabetes. Most individuals with type 1 diabetes will develop some form of cardiovascular illness during their lives. Indeed, more than half of all people who have been diagnosed with diabetes have hypertension. People with diabetes have a signiÞcantly elevated risk of developing atherosclerosis, and the disease is more likely to progress rapidly (Biondi-Zoccai GG, 2003). In addition, researchers recently discovered that glucose increases the expression of the macrophage scavenger receptor that increases oxidation of low-density lipoproteins (LDLs), in turn fostering formation of foam cells (macrophages that have taken up LDL), a hallmark of atherosclerosis (GrifÞn E, 2001). This mechanism may explain why diabetics develop more aggressive atherosclerotic disease than do people without diabetes (Liao D, 2002). CURRENT THERAPIES Pharmaceutical companies have followed two complementary strategies for improving glycemic control: improving the pharmacokinetic proÞles of insulins and creating more-convenient devices to administer insulin. Currently, type 1 diabetics use insulin for maintaining basal insulin levels between meals and achieving bolus insulin levels at meals. The purpose of the basal pattern (or steady release of low levels of insulin) is to maintain normal blood glucose levels between meals and overnight, whereas bolus insulin administration controls glucose levels during and immediately after meals. Products designed for strictly basal use are divided into two release proÞles: intermediate-acting and longacting. To administer a pre-meal bolus of insulin, patients use only short-acting insulins. A summary of selected insulins used in type 1 diabetes management is summarized in Table 2. Insulin can be administered through three types of devices: syringes, pens, and pumps. A pen is a type of syringe that is easier to use; both syringes and pens require users to inject themselves with bolus insulin before meals or basal insulin once or twice per day. Pumps administer short-acting insulin in response to meals (bolus), and they also deliver the same type of insulin in small amounts throughout the day to achieve a basal effect. The device remains connected to a patient by a subcutaneous insertion tube and therefore does not require a needle insertion for each administration of insulin. Currently, the most relevant division among basal and bolus insulins is human and analogue. (Animal insulins are rarely used and are not discussed here.) Genetically modiÞed bacteria produce human insulin; the Þnal product may be modiÞed by the addition of certain agents to attain the desired activity proÞle (short-, intermediate-, or long-acting). Insulin analogues, also synthetically produced by bacteria, achieve speciÞc release proÞles through the substitution of particular amino acids. Thus far, the panoply of marketed analogues includes

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TABLE 2. Current Therapies Used for Type 1 Diabetes Agent

Company/Brand

Short-acting insulins Regular human insulin (RHI)

Eli Lilly’s Humulin R; Novo Nordisk’s Novolin R/Actrapid Insulin lispro Eli Lilly’s Humalog Insulin aspart Novo Nordisk’s NovoLog Intermediate-acting insulins Neutral protamine Eli Lilly’s Humulin N; Novo Hagedorn (NPH) Nordisk’s Novolin N insulin Lente Eli Lilly’s Humulin L; Novo Nordisk’s Novolin L Combination insulins Human insulin Eli Lilly’s Humulin 70/30, combinations Humulin 50/50; Novo Nordisk’s Novolin 70/30, Mixtard 70/30; others Analogue insulin Eli Lilly’s Humalog Mix combinations 75/25; Novo Nordisk’s NovoLog Mix 70/30 Long-acting insulins Ultralente Eli Lilly’s Humulin U; Novo Nordisk’s Ultratard Insulin glargine Aventis’ Lantus U = units. J = Japan.

Daily Dose

Availability

25–50 U

US, F, G, I, S, UK, J

25–50 U 25–50 U

US, F, G, I, S, UK, J US, F, G, I, S, UK, J

25–50 U

US, F, G, I, S, UK, J

25–50 U

US, F, G, I, S, UK, J

50–100 U

US, F, G, I, S, UK, J

50–100 U

US, F, G, I, S, UK, J

25–50 U

US, F, G, I, S, UK, J

25–50 U

US, F, G, I, S, UK, J

US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom;

only short- and long-acting insulins. The two largest manufacturers of insulin, Eli Lilly and Novo Nordisk, also offer products that combine short- and intermediateacting insulin in a single premixed pen. These formulations reduce the total number of daily injections for some patients. Short-Acting Insulin Overview. Short-acting insulin products best mimic the form of insulin secreted by the body. The proper use of these products depends on patients’ advance estimates of their insulin needs and administration of the product at correct intervals to avoid hyper- and hypoglycemia. These agents are used as bolus insulin in syringes, pens, and pumps. Short-acting insulin is also used in pumps to maintain basal levels by supplying a small, steady ßow of insulin. Since the early 1980s, regular human insulin, consisting of dissolved zincinsulin crystals puriÞed from the recombinant proteins, has been available. Patients must inject it 30–60 minutes before a meal to ensure that their insulin levels are elevated enough to metabolize glucose from the meal. Eli Lilly and Novo Nordisk have launched insulin analogues that differ slightly from regular human insulin. In products from both companies, the amino acids are modiÞed—a process that allows the body to absorb analogues more rapidly by preventing

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insulin molecules from forming dimers or hexamers (larger molecules consisting of two or six identical insulin molecules, respectively) that are absorbed more slowly than individual molecules. These rapid-acting analogues allow patients to inject themselves immediately before eating, giving them more ßexibility in timing their meals. Mechanism of Action. The mechanism of action for all insulin products is relatively simple: the agents replace the insulin that the pancreas of the type 1 diabetic patient is no longer capable of producing. The manufactured insulin products mimic the physiological actions of endogenous insulin. Because they must be introduced into the body artiÞcially, however, they are not able to mimic perfectly the responsiveness of a healthy pancreas to variations in glucose levels. For a full discussion of the role of insulin in the metabolism of glucose, see the “Etiology and Pathophysiology” section. Short-acting insulin products do not contain any substrate such as neutral protamine Hagedorn (NPH) to delay the body’s absorption of insulin. Because all the insulin is immediately available, patients can metabolize a large inßux of glucose that occurs after a meal. Formulation. Patients can administer short-acting insulins through traditional syringes, pens, or pumps. Patients who use syringes obtain insulin in vials. The vials must be refrigerated for long periods of storage but can be left at room temperature for up to four weeks. Pens need not be refrigerated after their Þrst use, provided that all the contents are used within 10–28 days, depending on the particular product. Because they have a dial that allows patients to determine their dose accurately, pens simplify the injection process. Insulin pumps offer a radically different paradigm for insulin administration. Traditional insulin injections require the patient to maintain his or her basal insulin level by using an intermediate- or long-acting insulin. However, these products have unpredictable pharmacokinetic proÞles. They can be affected by factors as diverse as the temperature of the product and the patient’s level of activity. With an electronic insulin pump, patients can use a predictable shortacting insulin to achieve basal levels. The pumps are roughly the size of a pager and can be worn on a belt or kept in a purse. A ßexible tube connects the pump’s insulin reservoir to an infusion site on the patient’s body, usually the abdomen. Patients program the pump to release a bolus of insulin immediately before a meal and to continuously release lower levels of insulin between meals to maintain basal levels. In this way, short-acting insulin products can be used to replace less-predictable intermediate- or long-acting insulin and achieve tighter glycemic control. Because the Diabetes Control and Complications Trial (DCCT) demonstrated the beneÞts of precise glycemic control (as achieved through ßexible insulin therapy, or FIT), the insulin pump is an attractive option for patients who are motivated enough to deal with the complexities of the device. Indeed, a secondary endpoint of the DCCT showed that patients using insulin pumps exhibited lower HbA1c values than those who used multiple daily injections (DCCT

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Research Group, 1995). A complete discussion of the various insulin pumps is beyond the scope of the discussion here, but it should be noted that patients have access to a wide variety of products of differing levels of sophistication, and most patients can obtain reimbursement for them. Regular Human Insulin. Eli Lilly launched its Humulin R in 1983, and Novo Nordisk launched Novolin R (also called Actrapid) in 1987. They do not require prescriptions, but in most cases patients must have a prescription to obtain reimbursement and/or to purchase syringes or pens. These human insulins supplanted pork- and beef-derived insulin. Regular human insulin is produced by recombinant DNA bacteria. It is identical to the insulin naturally produced by a healthy pancreas, but because it is stored in a container rather than secreted directly into the bloodstream, the insulin molecules may cluster into dimers or hexamers. The body absorbs these aggregates more slowly than single insulin molecules, resulting in a delayed onset of action. Patients must administer regular human insulin 30–60 minutes before a meal for sufÞcient levels of insulin to be available when glucose from the meal reaches the bloodstream. In a six-month, randomized trial of 1,070 type 1 diabetic patients, regular human insulin was less effective than insulin aspart (Novo Nordisk’s NovoLog) in all but one of the main outcomes. Notably, regular human insulin was inferior on the measures of HbA1c levels, postprandial glucose levels, patient satisfaction, and risk of experiencing a hypoglycemic episode. The trial was open-label; nearly all insulin trials must be open-label because the different insulin products have different dosing regimens and appearances. Patients in both groups used NPH insulin to maintain basal levels. For bolus control, aspart was administered immediately before meals in one treatment arm, while regular human insulin was injected 30 minutes preprandially in the other arm. HbA1c levels did not change from baseline (7.98%) in patients treated with regular human insulin, while HbA1c dropped from 7.96% to 7.86% (a 1.3% improvement) in aspart-treated patients. Further, the regular human insulin group averaged a WHO Diabetes Treatment Satisfaction Questionnaire (DTSQ) quality-of-life score of 29.7–7.2% lower than that observed in the aspart group. Patients on regular human insulin were 20% more likely to experience a major hypoglycemic event than aspart users. On the other hand, regular human insulin reduced preprandial glucose levels before breakfast and dinner to a greater extent than aspart. However, given that HbA1c levels improved in aspart users, the improvement in preprandial glucose levels observed with regular human insulin does not necessarily indicate that regular human insulin has a clinical beneÞt over aspart (Home PD, 2000). Another study compared the efÞcacy of regular human insulin with that of another insulin analogue, lispro. An open-label, eight-month crossover trial of 463 adolescents (aged 9–18) comparing regular human insulin to insulin lispro (Eli Lilly’s Humalog) found that treatment with the regular human insulin resulted in poorer two-hour postprandial glucose levels after breakfast and dinner (patients treated with regular human insulin attained average glucose levels that were 9.7% higher for the two meals), though HbA1c levels were equal (8.70% for regular

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human insulin, 8.69% for lispro). Regular human insulin-treated patients experienced 8.7% more hypoglycemic episodes per day and 70% more hypoglycemic episodes in the midnight to 6 a.m. period (Holcombe JH, 2002). In light of the DCCT’s positive results for tight glycemic control regimens, insulin pumps are becoming more popular. An open-label, four-month, 146-patient trial examined regular human insulin, insulin aspart, and insulin lispro in pump users. After a four-week run-in with regular human insulin, the investigators noted no signiÞcant change in HbA1c levels from baseline at the end of the trial. Patients treated with regular human insulin experienced slightly more hypoglycemic episodes than aspart or lispro users (4.8 per month versus 3.7 and 4.4, respectively). All three products were compatible with insulin pumps, though a greater percentage (78%) of subjects in the regular human insulin group had at most three clogs compared with those subjects using aspart or lispro (75% and 64%, respectively) (Bode BW, 2002). Insulin Lispro. Eli Lilly launched insulin lispro (Humalog) in 1996. Its name indicates that the positions of two amino acids, lysine at B28 and proline at B29, have been switched from their positions in regular human insulin. This modiÞcation prevents the insulin molecules from forming dimers and hexamers without changing insulin’s pharmacological properties. In an open-label, crossover study of 1,008 type 1 diabetic patients comparing insulin lispro to regular human insulin, patients treated with lispro demonstrated signiÞcantly lower postprandial glucose levels. At one hour after meals, glucose levels in the lispro arm were 7.2% lower than those in the regular human insulin; levels were 13% lower two hours after eating. For both groups, HbA1c levels were equal (8.2%) after three months of treatment, as stated in the Humalog prescribing information. The labeling also indicates that in a 12-month parallel study, Humalog resulted in an overall rate of hypoglycemic episodes equal to that of regular human insulin. Nonetheless, Humalog users experienced fewer episodes in the midnight to 6 a.m. period. Further details, such as study size and rate of episodes, were not given. A smaller but more detailed study showed that insulin lispro offers some beneÞt over regular human insulin. An open-label, eight-month crossover trial of 463 adolescents (aged 9–18) comparing regular human insulin with insulin lispro (Eli Lilly’s Humalog) revealed that treatment with lispro resulted in better twohour postprandial glucose levels after breakfast and dinner (insulin lispro users’ glucose levels were an average of 9.0% lower for the two meals), though HbA1c levels were equal (8.69% for lispro, 8.70% for regular human insulin). Lisprotreated patients experienced 8.0% fewer hypoglycemic episodes per day and 41% fewer hypoglycemic episodes in the midnight to 6 a.m. period (Holcombe JH, 2002). Two studies directly compared the two short-acting insulin analogues. An open-label, four-month, 146-patient trial examined insulin lispro, insulin aspart, and regular human insulin in pump users. After a four-week run-in with regular human insulin, the investigators noted no signiÞcant change in HbA1c levels from baseline at the end of the trial. Lispro users experienced more hypoglycemic

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episodes than aspart users but fewer episodes than those on regular human insulin (4.4 per month versus 3.7 and 4.8, respectively). All three products were compatible with insulin pumps, although a greater percentage (78%) of subjects in the regular human insulin group had at most three clogs compared with those subjects using aspart or lispro (75% and 64%, respectively) (Bode BW, 2002). The other trial that involved both insulin aspart and insulin lispro examined only 24 patients. In this double-blind crossover study, patients received a postprandial dose of analogue insulin equal to the number of units they typically use before a meal, as determined by an insulin usage diary. On the second visit, patients received the same meal and injected the same number of units of the other insulin. For both visits, investigators ensured a consistent basal insulin level with an insulin clamp infusion before the meal. Both insulin analogues resulted in equivalent blood glucose levels at four and six hours postprandially. No difference in pharmacokinetic properties was noted. Because the study incorporated only two assessments within 14 days of each other, the effect of the study medications on HbA1c was not examined (Plank J, 2002). Insulin Aspart. Novo Nordisk launched insulin aspart (NovoLog) in 2001, several years after the launch of Eli Lilly’s analogue product. Unlike insulin lispro, in which the order of two amino acids is switched, insulin aspart prevents the problem of dimer and hexamer formation by replacing the amino acid proline at position B28 with aspartic acid. Two six-month, open-label studies described in insulin aspart’s labeling (one study of 1,040 patients and another study of 845 patients) found that insulin aspart is equivalent to regular human insulin in terms of HbA1c reduction and hypoglycemic events. In a six-month, open-label, randomized trial of 1,070 type 1 diabetic patients, insulin aspart performed better than regular human insulin in all but one of the main outcomes. Notably, insulin aspart was superior on the measures of HbA1c levels, postprandial glucose levels, patient satisfaction, and risk of experiencing a hypoglycemic episode. Patients in both groups used NPH insulin to maintain basal levels. For bolus control, aspart was administered immediately before meals in one treatment arm, while regular human insulin was injected 30 minutes preprandially in the other arm. HbA1c dropped from 7.96% to 7.86% (a 1.3% improvement) in insulin aspart-treated patients. HbA1c levels did not change from baseline (7.98%) in subjects treated with regular human insulin. Further, the insulin aspart group averaged a WHO DTSQ quality-of-life score of 29.7—7.7% higher than that observed in the regular human insulin arm. Patients using insulin aspart were 17% less likely to experience a major hypoglycemic event than patients treated with regular human insulin. However, insulin aspart did not reduce preprandial glucose levels to the same extent as regular human insulin. Given that HbA1c levels improved in aspart users, the improvement in preprandial glucose levels observed with regular human insulin does not necessarily indicate that regular human insulin has a clinical beneÞt over aspart (Home PD, 2000). An open-label, three-month trial of 423 patients showed that insulin aspart reduces HbA1c levels to a greater degree than regular human insulin. After

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treatment, HbA1c levels improved 4% over baseline (from 8.36% to 8.02%) for the insulin aspart group, whereas HbA1c levels of regular human insulin group improved by only 1.3%. (The improvement for regular human insulin may be attributed to dose-optimization at the beginning of the study.) The investigators reported no difference in frequency of hypoglycemic events. However, patients using insulin aspart found it to be more ßexible than their previous, pretrial regimen of regular human insulin plus NPH insulin. The study did not provide enough data to quantify the improvement in ßexibility, though overall treatment satisfaction was similar. Two studies directly compared the two short-acting insulin analogues. An open-label, four-month, 146-patient trial examined insulin lispro, insulin aspart, and regular human insulin in pump users. After a four-week run-in with regular human insulin, the investigators noted no signiÞcant change in HbA1c levels from baseline at the end of the trial. Aspart users experienced fewer hypoglycemic episodes than patients using insulin lispro or regular human insulin (3.7 per month versus 4.4 and 4.8, respectively). All three products were compatible with insulin pumps, though more aspart-treated patients (75%) experienced at most three clogs than those using lispro (64%) (Bode BW, 2002). The other trial that involved both insulin aspart and insulin lispro examined only 24 patients. In this double-blind crossover study, patients received a postprandial dose of analogue insulin equal to the number of units they typically use before a meal, as determined by an insulin usage diary. On the second visit, patients received the same meal and injected the same number of units of the other insulin. For both visits, investigators ensured a consistent basal insulin level with an insulin clamp infusion before the meal. Both insulin analogues resulted in equivalent blood glucose levels at four and six hours postprandially. No difference in pharmacokinetic properties was noted. Because the study incorporated only two assessments within 14 days of each other, the effect of the study medications on HbA1c was not examined (Plank J, 2002). Intermediate-Acting Insulin Overview. Intermediate-acting insulins provide basal levels of insulin. They are not used in pumps because their slow release proÞle makes them ill-suited to the pumps’ ability to respond quickly to changes in glucose levels. Type 1 diabetics who use intermediate-acting insulin typically inject themselves twice daily to maintain basal levels. No intermediate-acting insulin analogue is marketed as a separate product, though Eli Lilly and Novo Nordisk describe their combination analogue products as containing both short- and intermediate-acting forms of insulin analogue. Fundamentally, the intermediate-acting release proÞle is a stopgap measure; nearly all type 1 diabetics would beneÞt more from a long-acting insulin—but only one with a predictable release proÞle. However, because intermediate-acting insulins are more predictable than long-acting human insulin (with the exception of the analogue insulin glargine), intermediate-acting insulins remain the product of choice among human insulins for maintaining basal levels.

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Intermediate-acting insulins may contain either NPH or zinc to give the products the desired release proÞle. Though the zinc-based lente product (Novo Nordisk’s Novolin L, Eli Lilly’s Humulin L) once dominated the intermediateacting insulin market because of its better release proÞle and small number of allergic reactions, it has lost favor because of its incompatibility with regular human insulin—a major problem because the latter is a key element in the maintenance of tight glycemic control. Therefore, lente is not covered in detail here. Mechanism of Action. The mechanism of action of intermediate-acting insulins is identical to that of the short-acting products: these agents simply replace the endogenous insulin that the pancreas of the type 1 diabetic cannot secrete. Intermediate-acting insulin includes a delaying agent, typically NPH or zinc, giving it a slower onset of action than regular human insulin. Formulation. As stated in the overview, intermediate-acting agents are used in syringes and pens but not in insulin pumps. NPH Insulin. Eli Lilly launched its NPH insulin (Humulin N) in 1983. Novo Nordisk brought its product (Novolin N) to market in 1985. Although many trials use NPH insulin to maintain basal levels, nearly all do so to compare the bolus activity of different kinds of short-acting insulins—the performance of NPH insulin is not one of the trials’ end points. One study that did compare NPH insulin’s basal function with that of another basal insulin was a 619-patient, open-label, four-month trial against insulin glargine (an insulin analogue, Aventis’s Lantus). Patients used insulin lispro (Eli Lilly’s Humalog) for bolus insulin. Patients received NPH insulin once daily at bedtime, twice daily at bedtime and before breakfast, or insulin glargine once daily at bedtime. NPH users were assigned to a treatment group based on their pretrial NPH use (once daily or more than once daily). Reductions in HbA1c levels in both groups were comparable, indicating that the products have similar effects. All patients had similar incidences of nocturnal and severe hypoglycemia. Notably, NPH insulin users reported mild injection-site pain less frequently than insulin glargline users (0.3% versus 6.1%). With the exception of injection-site pain, NPH insulin performs similarly to the more-convenient insulin glargine. The investigators noted that insulin glargine resulted in improved fasting glucose levels over NPH; they speculate that if the patients’ bolus insulin (i.e., the insulin lispro) had been better titrated, the study would have yielded concurrent improvements in HbA1c levels and in incidences of hypoglycemia in insulin glargine users (Raskin P, 2000). A smaller study of 51 patients compared intensive NPH insulin therapy (four times daily) with glargine at dinner or bedtime. In this open-label, three-month trial, HbA1c levels worsened slightly in the NPH group (rising from 6.9% to 7.0%) while improving in both glargine groups (dropping from 6.8% to 6.4% for the dinnertime group and from 7.0% to 6.6% for the bedtime group). No episodes

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of severe hypoglycemia occurred during the study period, but mild hypoglycemia occurred more often in the NPH group (3.6 episodes versus 1.7 for glargine before bed and 2.0 for glargine at dinner). Furthermore, both glargine regimens were more effective than NPH insulin at maintaining targeted pre- and postprandial blood glucose levels (Rossetti P, 2003). Fixed Combination Insulin Overview. Fixed combination insulins eliminate the need for some patients to inject themselves with separate basal and bolus doses: a single pre-meal injection can supply both the bolus insulin necessary to metabolize the meal and the basal dose needed to maintain proper glucose levels until the next meal. Although other compositions may be available in other countries, the most common are premixed products that contain short-acting insulin (25%, 30%, or 50%) and intermediate-acting insulin (75%, 70%, or 50%). Eli Lilly and Novo Nordisk offer human insulin versions (regular human insulin plus NPH insulin) as well as analogue versions (short-acting analogue plus intermediate-acting analogue; intermediate-acting analogues are not available as separate products). These agents offer convenience to patients whose lifestyles are compatible with the premixed dosages. They are signiÞcantly less popular among type 1 diabetics than type 2 diabetics: because the former patients lack any pancreatic function, they typically require extremely precise amounts of insulin, a therapeutic need that is less likely to coincide with the available combination dosages. Furthermore, type 1 diabetics may obtain better glycemic control by injecting the bolus insulin at a different time from the basal insulin. For such patients, combination products are not an option. Mechanism of Action. Fixed combination insulins combine short-acting (bolus) and intermediate-acting (basal) insulins. Short-acting insulin products do not contain any substrate such as neutral protamine Hagedorn (NPH) to delay the body’s absorption of insulin. Because all the insulin is immediately available, patients can metabolize a large inßux of glucose that occurs after a meal. The mechanism of action of intermediate-acting insulins is identical to that of the short-acting products: these agents simply replace the endogenous insulin that the pancreas of the type 1 diabetic cannot secrete. Intermediate-acting insulin includes a delaying agent, typically NPH or zinc, giving it a slower onset of action than regular human insulin. Formulation. Human insulin combination products use NPH insulin as their intermediate-acting component. Both analogue products (Eli Lilly’s Humalog Mix75/25 and Novo Nordisk’s NovoLog Mix) use a crystal suspension of protamine and the short-acting analogue to achieve an intermediate-acting component. Like intermediate- and long-acting insulins (the former being a component of combination products), this class of agents should not be used in insulin pumps. However, they can be used in both syringes and pens. Pens are particularly

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appropriate for combination insulins; the purpose of these products is to improve convenience. Human Insulin Combinations. Eli Lilly launched Humulin 70/30 in 1989 and Humulin 50/50 in 1992. Novo Nordisk launched Novolin 70/30 in 1986. Novo Nordisk also offers other combinations of concentrations outside the United States under the Mixtard brand name. Because these agents are simply combinations of already available nonprescription products, clinical trial data were not collected for approval purposes. No trial appears to examine any Humulin product in type 1 diabetics. The labeling for Humalog Mix75/25 includes several graphs that show a slower absorption rate and less basal stability with Humulin 70/30 than with Humalog Mix75/25 in 30 nondiabetic patients. According to the prescribing information, “identical results were found in patients with type 1 diabetes.” A study comparing Novolin 70/30 to NovoLog Mix 70/30 found similar results. This open-label study followed 104 type 1 patients and 190 type 2 patients for three months (Boehm BO, 2002). The authors did not report separate results for the two groups, but they did state that “the analyses performed for type 1 and type 2 diabetes separately are all in agreement with the results of the overall analyses.” They report a drop in HbA1c of 0.01% and signiÞcantly lower postprandial blood-glucose levels for NovoLog 70/30 users. A trial described in the prescribing information of NovoLog Mix 70/30 involving 92 type 1 diabetes patients found similar changes in HbA1c among Novolin 70/30 users and NovoLog Mix 70/30 users after three months. However, NovoLog Mix 70/30 controlled postprandial glucose better than Novolin 70/30. Analogue Insulin Combinations. Eli Lilly launched Humalog Mix75/25 (lispro protamine suspension plus lispro) in 2000, while Novo Nordisk’s NovoLog Mix 70/30 (aspart protamine suspension plus aspart) did not launch until 2002. No detailed studies in type 1 diabetics have been published about either product. Because they simply contain new formulations of already approved insulin analogue products, major trials were not required. The prescribing information for Humalog Mix75/25 provides no clinical trial data in type 1 diabetic subjects. Nearly all studies cited in the prescribing information give some details about the product’s performance in nondiabetics and note that “similar results were found in patients with type 1 diabetes.” The data cited relating to nondiabetic patients show that the body absorbs Humalog Mix75/25 more rapidly than Humulin 70/30. Novo Nordisk has published more-detailed information in the prescribing information for NovoLog Mix 70/30. A three-month open-label trial involving 92 type 1 diabetes patients found similar changes in HbA1c levels and better control of postprandial glucose levels compared to Novolin 70/30. An additional study involving 104 type 1 patients and 190 type 2 patients supports these results. HbA1c levels fell 0.01% in NovoLog Mix 70/30 users compared with patients using Novolin 70/30 (a difference of questionable clinical signiÞcance), and postprandial blood glucose levels were signiÞcantly lower in the NovoLog Mix 70/30 group.

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Long-Acting Insulins Overview. Novo Nordisk launched the Þrst long-acting insulin, beef-derived Ultralente, in 1953. Today, Novo Nordisk offers the human insulin Ultratard, a product available in all regions under study except the United States. Lilly launched its Humulin U, a similar product, in 1987. Some patients lament the phasing-out of beef-derived long-acting products, which occurred in 1993 (Eli Lilly) and 1994 (Novo Nordisk), on the grounds that they have a steadier release proÞle and a longer duration of action than long-acting human insulins. Most experts consider the long-acting insulin analogue glargine (Aventis’s Lantus) superior to any of the previously available long-acting products. Unfortunately, clinical trials compare insulin glargine only to NPH insulin (an intermediateacting agent), not to long-acting human or animal-derived insulin. Mechanism of Action. The mechanism of action for all insulin products is relatively simple: the agents replace the insulin that the pancreas of the type 1 diabetic patient is no longer capable of producing. The manufactured insulin products mimic the physiological actions of endogenous insulin. Because they must be introduced into the body artiÞcially, however, they are not able to mimic perfectly the responsiveness of a healthy pancreas to variations in glucose levels. For a full discussion of the role of insulin in the metabolism of glucose, see the “Etiology and Pathophysiology” section. Formulation. As stated in the class overview, patients do not use long-acting agents in insulin pumps. However, they are compatible with syringes and pens. Ultralente. A double-blind study comparing once-daily ultralente insulin (Eli Lilly’s Humulin U) with once-daily NPH insulin (along with pre-meal insulin lispro for both groups) in 178 type 1 diabetics found no major difference in the two regimens. In both groups, patients had the option to switch to twice-daily injections of their medication (while remaining blinded) if once-daily injections did not adequately control glucose levels; 21% of NPH insulin patients did so, as did 24% of ultralente users. Levels of HbA1c and rates of severe hypoglycemia were similar for all patients. Patients using NPH insulin recorded lower blood glucose levels than ultralente users between the after supper and before lunch measurements, but the clinical signiÞcance of this observation is not known (Zinman B, 1999). Conventional wisdom holds that ultralente is less predictable than NPH insulin or insulin glargine. Although this may well have a negative effect on episodes of hyper- and hypoglycemia, no studies have shown that ultralente poorly regulates HbA1c levels or inadequately prevents diabetic complications compared to NPH insulin or insulin glargine. Insulin Glargine. Aventis launched insulin glargine (Lantus) in 2001. It is produced by replacing the aspargine amino acid at position A21 with glycine and by adding two arginines to the C-terminus of the B-chain. Insulin glargine

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cannot be mixed with other insulin products, so patients must inject it separately from their bolus insulin. Insulin glargine distinguishes itself from other long- and intermediate-acting insulins by its 24-hour action and ßat absorption and activity curves. These characteristics make it the most predictable injectable choice for maintaining basal insulin levels. The labeling for insulin glargine describes three trials (open-label, with 534, 585, and 619 patients; the Þrst two lasted seven months and the third for four months) that compared the product with NPH insulin in type 1 diabetic patients. The results for the two types of insulin were similar, though glargine users experienced small increases in HbA1c levels versus NPH insulin users. However, patients treated with insulin glargine reported lower average fasting blood glucose levels than those using NPH insulin. The prescribing information did not detail the differences between once-daily and twice-daily NPH insulin use. A seven-month study of pediatric type 1 diabetics (aged 6–15) also found slightly worse HbA1c changes with insulin glargine and improved average fasting blood glucose levels. A 619-patient, open-label, four-month trial of insulin glargine against NPH insulin showed that both regimens had similar results, but in this trial, insulin glargine yielded slightly better results than NPH insulin on HbA1c (Raskin P, 2000). Patients received either NPH insulin once a day at bedtime, twice daily at bedtime and before breakfast, or insulin glargine once daily at bedtime. NPH users were assigned to a treatment group based on their pretrial NPH use (once a day or more than once a day). In both groups, HbA1c levels fell by 0.1%; because baseline was 7.6% for the insulin glargine group and 7.7% for the NPH users, the drop was a greater improvement for the glargine group. All patients had similar incidences of nocturnal and severe hypoglycemia. Notably, insulin glargine users reported mild injection-site pain more frequently than NPH insulin users (6.1% versus 0.3%); this difference may stem from the fact that insulin glargine is more acidic than other types of insulin. With the exception of injection-site pain, insulin glargine, a more convenient agent, performed similarly to NPH insulin. Additionally, the investigators noted that insulin glargine resulted in better fasting glucose levels than NPH; they speculate that if the patients’ bolus insulin (i.e., the insulin lispro) had been better titrated, they would have observed concurrent improvements in insulin glargine users’ HbA1c levels and incidences of hypoglycemia. A smaller study of 51 patients compared glargine at dinner or bedtime with intensive NPH insulin therapy (four times daily) (Rossetti P, 2003). In this openlabel three-month trial, HbA1c levels in both glargine groups improved (dropping from 6.8% to 6.4% for the dinnertime group and from 7.0% to 6.6% for the bedtime group), but levels worsened slightly in the NPH group (rising from 6.9% to 7.0%). No episodes of severe hypoglycemia occurred during the study period, but mild hypoglycemia occurred more often in the NPH group (3.6 episodes versus 1.7 for glargine before bed and 2.0 for glargine at dinner). Furthermore, both glargine regimens were more effective than NPH insulin at maintaining targeted pre- and postprandial blood glucose levels.

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EMERGING THERAPIES The development of new therapies for type 1 diabetes focuses on three strategies. First, many companies are seeking to improve the properties of recombinant insulin to more accurately mimic the physiology of endogenous insulin. Second, efforts are under way to generate more-convenient modes of delivery, such as inhaled or oral formulations; selected emerging therapies are presented in Table 3. Finally, although in early-stage development, innovative approaches are aimed at modulating the immune response and thus preventing type 1 diabetes or reversing the pancreatic damage caused by this disease. An emerging nonpharmacological treatment approach to type 1 diabetes that warrants careful attention is the successful development of pancreatic islet-cell transplantation, which can revolutionize type 1 diabetes therapy by potentially eliminating the need for exogenous insulin. The endogenously produced insulin from the islet cell is physiologically more likely to meet needs for glucose control than do insulin injections and will not lead to peaks commonly seen with the latter. However, the effective transplantation of a pancreas or pancreatic islet cell faces many technical and medical hurdles, including isolating intact and active cells, preserving tissue, preventing transplant rejection, and the need for repeat procedures (the implanted pancreatic cells do not replicate). Despite these obstacles, recent studies indicate that progress is being made in this area and that diabetes patients can be helped by transplantation of pancreatic cells. Too many hurdles prevail in this Þeld to attract the interest of venture capitalists or corporate investors to Þnancially back this research, however, and as of this writing it is primarily academic physicians and scientists who are pursuing pancreatic isletcell transplantation. Short-Acting Insulin Analogues Overview. In vitro studies have revealed that insulin glargine has increased growth-stimulating potency compared with human insulin (Kurtzhals P, 2000). However, the safety implications of this Þnding are unclear. Properties of the analogues insulin aspart and lispro resembled those of regular human insulin, whereas insulin detemir exhibited lower metabolic and mitogenic properties than regular human insulin. Neither renal nor hepatic impairment appears to affect the pharmacokinetics of insulin detemir in a clinically signiÞcant manner (Jacobsen LV, 2002). Mechanism of Action. The mechanism of action for all insulin products is relatively simple: the agents replace the insulin that the pancreas of the type 1 diabetic patient is no longer capable of producing. The manufactured insulin products mimic the physiological actions of endogenous insulin. However, because they must be introduced into the body artiÞcially, they are not able to mimic perfectly the responsiveness of a healthy pancreas to variations in glucose levels. For a full discussion of the role of insulin in the metabolism of glucose, see the “Etiology and Pathophysiology” section.

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TABLE 3. Emerging Therapies in Development for Type 1 Diabetes Compound

Development Phase

Short-acting insulin analogues Insulin glulisine United States R Europe PR Japan — Long-acting insulin analogues Insulin detemir United States PR Europe PR Japan III Inhaled insulins Exubera United States III Europe PR Japan — AERx Insulin Diabetes Management System (NN-1998) United States III Europe III Japan I Advanced Inhaled Research (AIR) inhaled insulin United States II Europe — Japan — Oral insulins NIN-058 United States II Europe II Japan — Oralin United States II Europe II Japan — Oral insulin, Emisphere United States I Europe I Japan — Immunomodulators Anti-CD3 antibody United States II Europe — Japan — Diamyd United States — Europe II (Sweden) Japan — NBI-6024 United States II Europe II Japan I

Marketing Company

Aventis Aventis —

Novo Nordisk Novo Nordisk Novo Nordisk

Pfizer/Aventis Pfizer/Aventis — Novo Nordisk Novo Nordisk Novo Nordisk Eli Lilly — —

Nobex Nobex — Generex Generex — Emisphere Emisphere —

Columbia University — — — Diamyd Therapeutics — Neurocrine Biosciences Neurocrine Biosciences Taisho

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TABLE 3. (continued) Compound

Development Phase

Insulin sensitizers Somatokine United States Europe Japan T-1095 United States Europe Japan Amylin agonists Pramlintide United States Europe Japan

Marketing Company

II — —

Insmed — —

II II II

Johnson & Johnson Johnson & Johnson Tanabe

PR III —

Amylin Pharmaceuticals Amylin Pharmaceuticals —

R = Registered; PR = Preregistered.

Short-acting insulin products do not contain any substrate such as neutral protamine Hagedorn (NPH) to delay the body’s absorption of insulin. Because all the insulin is immediately available, patients can metabolize a large inßux of glucose that occurs after a meal. Insulin analogues contain one or two amino acid modiÞcations that confer pharmacokinetic advantages over regular human insulin. This chemical alteration prevents the insulin molecules from forming dimers and hexamers that slow absorption without changing insulin’s pharmacological properties. Insulin Glulisine. To complete its portfolio of insulin products, Aventis is developing a new rapid-acting human insulin analogue, insulin glulisine. In April 2004, the FDA approved insulin glulisine for the treatment of type 1 and type 2 diabetes. The drug launched in 2004 and is marketed under the brand name Apidra. Aventis is also planning a clinical development program for insulin glulisine in Japan. Preliminary pharmacokinetic data show that this agent has a proÞle similar to that of insulin lispro (Becker RHA, 2003[a]). Long-Acting Insulin Analogues Overview. In vitro studies have revealed that insulin glargine has increased growth-stimulating potency compared with human insulin (Kurtzhals P, 2000). However, the safety implications of this Þnding are unclear. Properties of the analogues insulin aspart and lispro resembled those of regular human insulin, whereas insulin detemir exhibited lower metabolic and mitogenic properties than regular human insulin. Neither renal nor hepatic impairment appears to affect the pharmacokinetics of insulin detemir in a clinically signiÞcant manner (Jacobsen LV, 2002). Mechanism of Action. The mechanism of action for all insulin products is relatively simple: the agents replace the insulin that the pancreas of the type 1

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diabetic patient is no longer capable of producing. The manufactured insulin products mimic the physiological actions of endogenous insulin. However, because they must be introduced into the body artiÞcially, they are not able to mimic perfectly the responsiveness of a healthy pancreas to variations in glucose levels. For a full discussion of the role of insulin in the metabolism of glucose, see the “Etiology and Pathophysiology” section. Insulin analogues contain one or two amino acid modiÞcations that confer pharmacokinetic advantages over regular human insulin. This chemical alteration prevents the insulin molecules from forming dimers and hexamers that slow absorption without changing insulin’s pharmacological properties. Insulin Detemir. Novo Nordisk is developing the long-acting insulin analogue insulin detemir (NN-304) to complement its marketed short-acting insulin analogue aspart. Insulin detemir is preregistered in the United States and Europe and in Phase III trials in Japan. In an October 2003 company press release, Novo Nordisk announced that, in an approvable letter for insulin detemir, the FDA requested additional clinical information. In February 2003, the company stated that it expected U.S. and European marketing approvals in mid 2004. The drug launched in 2004 and is marketed under the brand name Levemir. The most recent clinical trial data in type 1 diabetics were presented as abstracts at the 63rd ScientiÞc Sessions of the American Diabetes Association (ADA) in June 2003. In a study investigating the pharmacodynamics of insulin detemir in 54 type 1 diabetics, insulin detemir exhibited less variability in glycemic control than NPH and insulin glargine (Heise TC, 2003). The multicenter, 16-week, open, parallel trial randomized 408 type 1 diabetics (baseline HbA1c = 8.6%) into three groups: insulin detemir once daily at 12-hour intervals, insulin detemir twice daily at morning and bedtime, and the comparator NPH twice daily at morning and bedtime. Data showed that administration of insulin detemir is ßexible—both regimens resulted in a level of glycemic control (HbA1c) similar to that of NPH (HbA1c 7.75% for the once-daily regimen and 7.78% for the twice-daily regimen versus 7.94% for NPH). Moreover, fasting plasma glucose (FPG) and variability in FPG were lower in detemir-treated patients than in NPH subjects, and detemir-treated patients gained less weight than NPH-treated patients (0.02 and 0.24 kg for detemir versus 0.86 kg for NPH) (Home PD, 2003). A six-month, multinational, open trial randomized 747 type 1 diabetics to oncedaily bolus regimens of NPH or insulin detemir. Although HbA1c reductions were not statistically signiÞcant, investigators observed a trend toward a greater HbA1c reduction in insulin-detemir-treated patients. Insulin detemir also induced statistically signiÞcant reductions in FPG and less variability in FPG, including glucose ßuctuations, than NPH, and it reduced the risk of nocturnal hypoglycemia by 26% compared with NPH. In addition, weight gain was less in patients treated with insulin detemir compared with NPH (0.2 kg versus 0.4 kg, p = 0.003) (Russel-Jones D, 2003). Investigators compared the efÞcacy and safety proÞles of insulin detemir and NPH in an open-label, randomized, crossover Phase II trial involving 59 type

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1 diabetics. Patients received NPH insulin as their basal insulin for a two-week run-in period, followed by two six-week periods when patients received NPH or insulin detemir therapy at bedtime. All patients were also administered subcutaneous injections of short-acting insulin 30 minutes before each meal. Results indicated that 24-hour serum glucose concentrations were comparable between treatment groups. FPG levels, which were assessed during the last four days of the treatment period, were similar between insulin detemir and NPH (8.32 [± 3.58] mmol/L versus 8.75 [± 4.16] mmol/L, respectively). However, researchers found that patients on insulin detemir exhibited less variability in FPG than did those on NPH. Six-week data showed that the number of hypoglycemic occurrences in the insulin detemir group was similar to that in the NPH treatment groups. Yet, during the last week of treatment (when patients were asked not to adjust their insulin doses), insulin detemir-treated patients demonstrated fewer hypoglycemic episodes than did those on NPH (60% and 77%, respectively). The insulin dosing required for patients on insulin detemir was 2.35 times higher than for patients on NPH (Hermansen K, 2001). In September 2001, Novo Nordisk presented trial data for insulin detemir at the annual meeting of the European Association for the Study of Diabetes. In a study of 447 patients with type 1 diabetes, patients received either insulin detemir or NPH insulin as their basal insulin for six months. Patients on insulin detemir demonstrated a signiÞcantly reduced FPG level compared with those on NPH. Patients on insulin detemir also had a 20% lower relative risk of having a hypoglycemic episode than did those on NPH insulin. Insulin detemir will likely be a once-daily subcutaneous injection (at bedtime) or a twice-daily subcutaneous injection (bedtime and morning dose) in patients with morning fasting hyperglycemia. Inhaled Insulin Overview. The three major producers of insulin—Eli Lilly, Novo Nordisk, and Aventis (with PÞzer)—are partnering with small companies to develop inhaled formulations. Three inhalers are in late-stage development: PÞzer/Aventis/Nektar’s Exubera, Aradigm/Novo Nordisk’s AERx, and Alkermes/Eli Lilly’s AIR. Other companies developing inhaled formulations of insulin include Bristol-Myers Squibb (in collaboration with QDose), MannKind (formerly Pharmaceutical Discovery), Kos Pharmaceuticals, BioSante Pharmaceuticals, and Coremed. Despite a generally positive outlook for inhaled insulin therapy because of its convenient mode of administration, there are several reasons for caution in predicting how widely it will be adopted: •

•

Although the cost of inhaled insulin therapy is not yet known, it is likely to be signiÞcantly more expensive than injectable insulin. This mode of delivery is less efÞcient than direct injection, so it requires more insulin. Because insulin is a growth factor for many cell types, some physicians are concerned that long-term use might increase the risk of pulmonary Þbrosis.

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• •

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In the clinic, physicians will need to scrutinize the reproducibility of each patient’s breathing and the precise amount of insulin inhaled to optimize dosing. Although the convenience and noninvasiveness of inhaled insulin are generally regarded as signiÞcant advantages, the inhalers now in development are bulkier than asthma inhalers and often less convenient than an insulin pen.

Mechanism of Action. The mechanism of action for all insulin products is relatively simple: the agents replace the insulin that the pancreas of the type 1 diabetic patient is no longer capable of producing. The manufactured insulin products mimic the physiological actions of endogenous insulin. However, because they must be introduced into the body artiÞcially, they are not able to mimic perfectly the responsiveness of a healthy pancreas to variations in glucose levels. For a full discussion of the role of insulin in the metabolism of glucose, see the “Etiology and Pathophysiology” section. The mechanism of inhaler devices for insulin is similar to that of asthma inhalers. The insulin is propelled into the lungs upon inspiration and is absorbed into the bloodstream through the alveoli. Exubera. Nektar (formerly Inhale Therapeutic Systems), in collaboration with PÞzer and Aventis, is developing a short-acting dry-powder insulin formulation called Exubera that is in Phase III trials in the United States and is preregistered in Europe. A pegylated, long-acting form of Exubera is in preclinical development. Unlike conventional dry-powder inhalers, Nektar’s Inhance dry-powder inhaler technology is designed to deliver the correct dosage independent of the patient’s inspiratory ßow rate. The three companies are codeveloping Exubera, and PÞzer and Aventis will commercialize the product worldwide. In June 1999, Phase III trials involving type 1 and 2 diabetics began at 117 sites in the United States and Germany; they were completed in July 2001. In December 2001, the developers decided to postpone the NDA Þling to collect additional long-term pulmonary safety data. In October 2002, the collaborators announced that additional long-term clinical trials in type 1 and 2 diabetic patients were under way and that discussions with regulatory authorities regarding the timing of their submissions were ongoing (Aventis, press release, October 11, 2002). At the Credit Suisse First Boston (CSFB) European Pharmaceutical Conference, Aventis conÞrmed in November 2002 that the companies had not yet decided on a date for submission because of issues surrounding subject enrollment, data collection and analysis, and FDA requirements. Studies of type 1 diabetes patients reveal that Nektar’s Exubera reduces HbA1c to the same extent as injected insulin. In an open-label, proof-of-concept, parallelgroup, randomized trial, 73 type 1 diabetics were randomized to subcutaneous insulin therapy (preprandial short-acting insulin injections plus twice-daily NPH) or Exubera (preprandial doses plus ultralente bedtime injectable insulin) for 12 weeks. At the end of the treatment period, Exubera reduced HbA1c (from 8.5% at baseline to 7.9%) at a level comparable to that of subcutaneous insulin therapy (from 8.5% at baseline to 7.7%). Fasting and postprandial glucose proÞles and the number of hypoglycemic episodes were similar in both treatment groups (Skyler JS, 2001).

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A six-month, multicenter Phase III trial involving 328 type 1 diabetics (aged 12–65 years) compared the glycemic control effected by an intensive inhaledinsulin regimen with the control effected by injected insulin (Skyler JS, 2002). Patients were randomized into one of two groups: inhaled insulin before meals (1–2 inhalations, corresponding to 1 and 3 mg) plus morning and bedtime NPH; and injected insulin before meals plus morning and bedtime NPH. FPG fell by 35 mg/dL in the inhaled-insulin group but increased by 4 mg/dL in the injectedinsulin group. Postprandial glucose (PPG) fell in the inhaled- and injected-insulin groups by 21 mg/dL and 14 mg/dL, respectively. HbA1c declined by 0.3% and 0.1% from baseline in the inhaled- and injected-insulin groups, respectively. Of the patients randomized to inhaled insulin, 64% achieved an HbA1c less than 8% and 23% achieved an HbA1c less than 7%. In the comparator, 60% achieved an HbA1c less than 8% and 22% achieved an HbA1c less than 7%. In the aforementioned study, severe hypoglycemic rates were greater in the inhaled-insulin group, compared with the injected-insulin group (6.5 events/100 subject-months versus 3.3 events). Coughs, all mild to moderate, were more common with inhaled insulin than with injected insulin (25% versus 7%). Studies presented at the 63rd ScientiÞc Sessions of the ADA in June 2003 addressed concerns about pharmacodynamics. One assessment demonstrated that smoking alters the absorption of Exubera in healthy nondiabetic subjects and concluded that patients should abstain before and during treatment (Becker RHA, 2003[b]). In another study involving elderly, obese type 2 diabetics, investigators showed that the absorption proÞle of Exubera was comparable to that of subcutaneous insulin (Henry R, 2003). Exubera’s efÞcacy is promising, but its long-term side effects have yet to be established. At the May 2001 meeting of the American Thoracic Society in San Francisco, Exubera researchers reported that the product was found to be safe after two years’ administration in a trial involving 140 patients: no evidence of lung function impairment was observed in the trial. In the same month, however, Aventis and PÞzer disclosed one case of pulmonary Þbrosis in the 1,000 patients who participated in various trials lasting more than six months—although a direct link to Exubera has not been established. If more cases of Þbrosis emerge, the FDA may require warnings on the product label discouraging physicians from using Exubera as a Þrst-line therapy. AERx Insulin Diabetes Management System (NN-1998). Aradigm, in collaboration with Novo Nordisk, is developing the AERx Insulin Diabetes Management System (iDMS) (NN-1998) for pulmonary delivery of insulin. Novo Nordisk has exclusive rights for worldwide marketing of any products resulting from the development program. It is in Phase III development for type 1 diabetes in the United States and Europe and in Phase I in Japan. In a September 2002 press release, Novo Nordisk announced the initiation of a two-year, multicenter, open-label Phase III trial that will investigate the long-term safety and efÞcacy of AERx iDMS compared with subcutaneous injections of insulin aspart in 300 type 1 diabetics. However, an investment research report published in

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August 2003 indicated that additional pivotal Phase III trials have been delayed until 2005 because of a cosmetic manufacturing defect in the AERx insulin dosage packets. Interim data from a 12-month Phase III trial in type 1 diabetics show no changes in pulmonary function but reveal unexpected glucose control. According to researchers, postprandial plasma glucose levels were higher and nighttime plasma glucose levels were lower in the AERx-treated group compared with the subcutaneous insulin-treated group. Aradigm and Novo Nordisk will continue to study the implications of these results (Aradigm, press release, April 30, 2004). NN-1998 produces a Þne aerosol mist of insulin, deposited deep into the lung for absorption into the bloodstream. It is a rapid-acting insulin that can be taken immediately before meals. The AERx system has approximately the same dimensions and weight as a small video camera. It releases a Þne, controlled aerosol mist of insulin while a handheld electronic inhaler with computerized sensors guides the patient’s breathing to automatically deliver the drug at the optimal point during inhalation. Aradigm has been issued a U.S. patent relating to the regulation of patient breathing for effective and reproducible pulmonary delivery of insulin. AERx is by far the most technologically advanced of the insulin inhalers in development. Because of the importance of dosage precision and reproducibility in insulin-treated patients, particularly type 1 diabetics, this product may provide an advantage over Nektar’s pulmonary delivery system. Additionally, the electronic compliance monitoring feature will provide valuable information to physicians and represents a competitive advantage over other inhaler technologies (Luery D, 2003). At this time, published clinical data on the efÞcacy of AERx iDMS in type 1 diabetics is sparse. In one published study—an open-label, crossover Phase I trial—18 type 1 diabetics were randomized to preprandial subcutaneous injections of regular insulin (0.12 U/kg) or AERx iDMS (0.3, 0.6, 1.2, or 1.8 U/kg) (Brunner GA, 2001). The pharmacokinetic and pharmacodynamic data show that AERx iDMS increases serum insulin and glucose infusion rates in a dosedependent manner. Additionally, the inhaled insulin demonstrated a faster onset of action than regular insulin. There are a few studies that demonstrate that the system is safe and well tolerated, but it requires specialized dosing in diabetics with asthma and those who smoke. In one trial, researchers examined a group of 45 nondiabetic, nonsmoking subjects, 17 of whom were diagnosed with mild-to-moderate asthma (Henry RR, 2003). Results indicated that patients with asthma consistently exhibited less insulin absorption and, consequently, lower reduction of blood glucose levels compared with the control group. Researchers did not observe signiÞcant changes in lung function measurements after administration of AERx iDMS. Although the pulmonary delivery of insulin was well tolerated in this trial, the data suggest that type 1 diabetic patients with asthma—because of changes in their airway caliber, blood ßow, and airway secretions—may require a higher dose of insulin to control their blood glucose.

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Another trial investigated the pharmacokinetics of this system in 27 nondiabetic smokers and 16 nonsmokers (Himmelmann A, 2003). Because a signiÞcant proportion of the diabetic population consists of smokers, the researchers sought to evaluate the effect of smoking on the absorption of inhaled insulin. Results indicated that approximately 60% more insulin was absorbed in smokers versus nonsmokers. Advanced Inhaled Research Inhaled Insulin. In April 2001, Lilly entered into a partnership with Alkermes to develop short- and long-acting inhaled formulations of insulin utilizing Alkermes’ Advanced Inhalation Research (AIR) delivery system. Under the terms of the agreement, Eli Lilly holds exclusive worldwide rights to products resulting from this collaboration and has responsibility for conducting clinical trials, securing regulatory approvals, overseeing large-scale manufacturing, and marketing the drug. Two months later, Lilly ended ´ its collaboration with Elan (formerly Dura Pharmaceuticals), with which it had ´ planned to develop an inhaled insulin formulation using Elan’s Spiros pulmonary delivery technology. Alkermes announced in a January 2003 presentation that a formulation of the AIR delivery system had entered Phase II trials in the United States for type 1 diabetes. Although the formulation that has advanced to Phase II is unclear, the available clinical data suggest that it is likely to be short-acting insulin. As a small inhaler that can deliver small peptides and complex macromolecules, the AIR system has the potential for sustained-release delivery of drugs such as long-acting insulin, representing an advantage over other inhaled insulin formulations. No efÞcacy data are available for AIR inhaled insulin. Alkermes announced positive preliminary Phase I results of AIR insulin but did not provide speciÞc data (Alkermes, press release, November 20, 2000). No efÞcacy data in type 1 diabetics are currently available, but Phase I data presented at the June 2001 ScientiÞc Sessions of the American Diabetes Association show that, in a study of 12 healthy subjects, AIR inhaled insulin has a faster onset of action than insulin lispro or regular insulin (Osborn C, 2001). Oral Insulins Overview. While inhaled insulin is currently garnering the attention of the medical community, several companies are developing oral formulations of insulin. Three companies are leading the development of oral insulin. The most clinically advanced agents are Nobex’s NIN-058 and Generex Biotechnology’s Oralin, both of which are in Phase II trials and have recently dissolved partnerships with pharmaceutical companies. Emisphere’s oral insulin is in an earlier stage of development. In addition to its convenient administration, oral insulin offers numerous physiological beneÞts. It mimics the natural secretion pathway of endogenous insulin in that oral insulin enters the liver and is released into the periphery via the hepatic portal vein. In addition, it has the potential to provide better hepatic glucose control, which may improve fasting plasma glucose levels. However, the

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development of orally administered insulin presents many challenges: the technology must offer protection from the hostile environment of the digestive tract; the patient’s eating and drinking behavior will greatly inßuence the effective doses that are absorbed into the bloodstream; and long-term safety still needs to be determined. Mechanism of Action. The mechanism of action for all insulin products is relatively simple: the agents replace the insulin that the pancreas of the type 1 diabetic patient is no longer capable of producing. The manufactured insulin products mimic the physiological actions of endogenous insulin. However, because they must be introduced into the body artiÞcially, they are not able to mimic perfectly the responsiveness of a healthy pancreas to variations in glucose levels. For a full discussion of the role of insulin in the metabolism of glucose, see the “Etiology and Pathophysiology” section. NIN-058. Nobex (formerly Protein Delivery) is developing an oral formulation of insulin using its proprietary polymer conjugation technology. In May 2002, Nobex entered into a strategic alliance with GlaxoSmithKline, but GlaxoSmithKline dissolved the collaboration in November 2003, citing internal business reasons. Nobex is seeking another partner. NIN-058 is in Phase II development in the United States and Europe. An open-label, dose-escalation Phase I/II trial examined the safety and tolerability of oral insulin when added to basal insulin therapy in 16 type 1 diabetics. The study revealed that 15 patients experienced 58 adverse events, all of which were classiÞed as mild or moderate. There were no discontinuations. The most frequently reported events were asthenia (n = 4) and headache (n = 4). Other adverse events that were reported in one or two patients included thirst, nausea, anemia, dry mouth, sleepiness, ataxia, dizziness, nervousness, paresthesia, urinary urgency, and back pain. These results suggest that oral insulin is well tolerated. Although it was not the primary endpoint of the study, researchers observed that oral insulin improved glucose excursions after a meal (Clement S, 2004). In a dose-escalation trial of oral insulin, researchers studied 18 fasting, type 1 diabetes patients (Clement S, 2001). Thirty minutes after the cessation of an intravenous insulin infusion, patients took capsules of oral insulin (0.6 mg/kg, 0.8 mg/kg, or 1.0 mg/kg). They received a second dose after 120 minutes. One week later, patients took identical doses to assess the reproducibility of the results. Blood glucose levels declined within 20 minutes of administration of oral insulin. After more than 240 minutes, 13 of 18 patients had blood glucose levels that remained below 150% of their baseline levels. No episodes of hypoglycemia were observed. The Nobex formulation maintained two-hour PPG at pretreatment levels (between 80 and 250 mg/dL) in a small, double-blind study of 31 type 1 diabetics (Clement S, 2002). Results were similar to those of insulin lispro. Investigators noted an increased incidence of gastrointestinal side effects with the Nobex formulation compared with that of insulin lispro.

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The results of another trial of oral insulin involving 15 type 1 diabetes patients (Paresh D, 2001) were presented at the American Diabetes Association 61st Annual ScientiÞc Session in Philadelphia (June 2001). In this study, patients took oral insulin after an eight-hour fast on day one. On day two, patients did not receive any oral insulin but ate a “standard” meal. On day three, patients received oral insulin followed by a standard meal 30 minutes later. Patients also received continuous subcutaneous insulin infusion therapy throughout the study. On average, oral insulin dosing resulted in a 25% reduction in postmeal glucose levels compared with basal insulin alone. Some patients in the fasting and fed states experienced hypoglycemic responses, although no signiÞcant adverse events were reported. Nobex claims that its oral delivery of insulin may be safer than injectable or other peripheral modes of delivery because it is absorbed into the portal vein that enters the liver, which plays a critical role in blood glucose control. In contrast, insulin injections may boost insulin levels in the blood, increasing the risk of hypoglycemia (Nobex, press release, February 16, 2001; Surguladze D, 2002). Oralin. Until May 2003, Generex and Lilly were codeveloping an oral insulin formulation utilizing RapidMist, a Generex proprietary buccal drug delivery technology. After Lilly’s withdrawal from the partnership, Generex announced its intention to continue developing the product, named Oralin, with other partners (Eli Lilly and Generex Biotechnology, press release, May 23, 2003). The RapidMist drug delivery system allows patients to spray a rapid-acting insulin into the buccal cavity; this in-mouth spray has been shown to have a faster absorption and elimination proÞle than injected short-acting insulin (Modi P, 2002). The compound is in Phase II trials for type 1 diabetes in the United States and Europe. Preliminary results from an open-label, active-comparator trial involving 20 type 1 diabetics were recently released. Subjects were randomized into the following groups: (1) once-daily Oralin before a meal, (2) one dose preprandially and a second dose postprandially, or (3) regular human insulin administered 15 minutes before the meal. Although speciÞc results were not revealed in the press release, the data suggest that Oralin has a faster onset than regular insulin (Generex, press release, May 4, 2004). Pharmacodynamic studies in healthy individuals and type 1 diabetics show that Oralin’s onset of action is faster than that of subcutaneous regular insulin and its time to peak onset is similar to that of insulin lispro (Guevara-Aguirre J, 2003[a]; Raz I, 2003). In type 1 diabetics, researchers observed no differences in absorption on three different occasions (Guevara-Aguirre J, 2003[b]). Preliminary efÞcacy studies in type 1 diabetic patients demonstrate that Oralin controls glucose levels in a manner comparable to that of subcutaneous insulin (Modi P, 2003). Among type 2 diabetics, data suggest that Oralin can be used effectively in combination with currently available oral antidiabetic agents to effect glycemic control (Guevara-Aguirre J, 2002; Schwartz S, 2001). Data presented at the 62nd ScientiÞc Sessions of the ADA in June 2002 conÞrm intriguing early reports

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that Generex’s oral insulin can be used effectively in combination with currently available oral antidiabetics to effect glycemic control. No human trial data are available regarding the safety of this compound, but Generex announced the results of a two-year safety and toxicology animal study, which indicated that chronic administration of Oralin does not change the epithelial cells of the mouth or induce toxic injury (Generex, press release, August 15, 2002). The company previously announced results of studies that suggested oral insulin poses no risk to the lungs because its droplets are too large to enter deep lung regions (Generex, press release, November 8, 2001). Emisphere. Emisphere is developing enteric-coated capsule and tablet formulations of insulin using its proprietary Eligen technology that allows for the oral delivery of insulin. The company is actively seeking potential partners. It is currently in Phase I trials in the United States and Europe. Emisphere’s Eligen technology utilizes a carrier that forms a stable complex with insulin. This stable molecular complex slows down the degradation of insulin by gastrointestinal enzymes and increases the insulin’s absorption rate into the bloodstream. Although it appears that Emisphere is developing oral insulin for type 1 and type 2 diabetes, preliminary results from studies in type 2 diabetic patients and normal subjects are the only data available at this time. In a single-dose, proofof-concept study, 20 type 2 diabetics were administered 300 units of oral insulin at bedtime (Arbit E, 2003). Results collected the following morning indicated that oral insulin decreases fasting plasma glucose, insulin, and C-peptide levels. In another single-dose study in 18 male type 2 diabetics, oral insulin demonstrated a pronounced reduction in postprandial glucose levels compared with a subcutaneous injection of short-acting insulin or placebo (Arbit E, 2003). A small trial in 29 normal volunteers demonstrated rapid absorption of Emisphere’s oral insulin formulation; peak serum insulin levels were achieved within 25 minutes. C-peptide levels (a marker for endogenous insulin) were suppressed maximally at one hour (Abbas R, 2002). Other than hypoglycemia, no signiÞcant side effects were observed in one study of 29 normal volunteers. A similar single-dose study demonstrated no adverse effects 12 weeks after dosing (Kidron M, 2002). Immunomodulators Overview. Prior to the discovery of insulin in 1921, there was little help for people who were afßicted with type 1 diabetes, so the development of insulin therapy was a breakthrough in the treatment of this disease. The next revolutionary therapies are likely to arise from a group of disease-modifying agents that will modulate the autoimmune response that causes type 1 diabetes and potentially present a cure for the disease. Mechanism of Action. Although the pathogenesis of type 1 diabetes has yet to be fully understood, it is clear that the disease is caused by autoimmunity. Immunomodulators, through various mechanisms, block the T cell-mediated autoimmune response that results in pancreatic beta-cell destruction.

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Anti-CD3 Antibody. A nonbinding Fc receptor, the anti-CD3 monoclonal antibody (hOKT3gl [Ala-Ala]) is currently in Phase II development by Columbia University. This antibody suppresses the autoimmune response by blocking T-cell signaling via CD3 and leading to immunological tolerance. A randomized, placebo-controlled Phase II trial assessing the efÞcacy of anti-CD3 antibody (diabetes) found that this humanized monoclonal antibody maintains or improves insulin production in patients with type 1 diabetes (Herold KC, 2002[a]). Twelve of 24 patients received a 14-course daily treatment with the antibody. Researchers measured C-peptide, glutamic acid decarboxylase (GAD), and HbA1c levels before treatment and at 6 and 12 months following treatment. At six months, patients treated with anti-CD3, as compared with those on placebo, exhibited a marked reduction in the decline of the C-peptide response following a meal, an indication that disease progression had slowed signiÞcantly. Researchers made the same observation at 12 and 18 months. Prior to treatment, levels of HbA1c were nonstatistically lower in the placebo group. At both 6 and 12 months, researchers found HbA1c levels were lower in the drug-treated group, an indication of tighter glycemic control among patients treated with the monoclonal antibody. Supporting this discovery was the observation that this patient group used less insulin than the placebo group did during the study period. However, a majority of patients taking anti-CD3 antibody experienced mild side effects. These effects included rashes, mild to moderate fever during the period of antibody administration, nausea, vomiting, headache, and arthralgia. Several patients expressed anti-idiotype antibodies, but after 12 months these levels were undetectable. Additional clinical results from a small trial involving 31 patients (17 drugtreated, 14 untreated control patients) treated with anti-CD3 monoclonal antibody for 10 days were recently reported (Herold, KC, 2002[b]). The drug-treated cohort exhibited a signiÞcant reduction in disease progression as assessed by the area under the curve of C-peptide response during a four hour mixed meal tolerance test, HbA1C levels and CD8:CD4 T-cell ratio. Researchers concluded that antiCD3 treatment induces energy of TH1 cells. They also reported that insulin usage among drug-treated patients was signiÞcantly reduced and that HbA1C levels improved. Diamyd. Diamyd is a diabetes vaccine in development by Diamyd Therapeutics, currently in Phase II trials in Europe for the prevention and treatment of type 1 diabetes. Advancement to a two-year, multicenter Phase III study in 350 type 1 diabetics is dependent upon Phase II results and potential outlicensing deals that the company is currently seeking. The Diamyd vaccine consists of an alum adjuvant and a 65 kD portion of the enzyme glutamic acid decarboxylase (GAD), a major autoantigen in type 1 diabetes. The vaccine promotes immunological tolerance such that T cells will no longer attack pancreatic beta islets. SpeciÞcally, regulatory T cells, which are a small subpopulation of CD4+ T cells, are upregulated, leading to the suppression of types of effector T cells that are responsible for pancreatic beta-cell destruction.

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Because human clinical trial data are currently available only in type 2 diabetes, this section presents data that are relevant to this population. A six-month, placebo-controlled, double-blind Phase II study was conducted in 47 GADpositive type 2 patients who are at high risk of developing either type 1 diabetes or latent autoimmune diabetes in adults (LADA) (Lernmark A, 2003). Participants were randomized into four treatment groups: 4, 20, 100, and 500 mcg. Within each dosing arm, patients were administered either the vaccine or a placebo at week 1 and week 4. After six months, results demonstrated that the optimal dose is 20 mcg—patients in this group exhibited improved fasting and postprandial C-peptide levels compared with placebo, suggesting that pancreatic beta-cell function is preserved. Positive results have been reported in preclinical experiments with a GAD vaccine (Tisch R, 2001). T-helper 2 (TH2) cells, speciÞc for the GAD antigen, are present in young, nonobese diabetic (NOD) mice. TH cell clones speciÞc for the GAD antigen were established and used for adoptive transfer experiments. When these TH2 cells were transferred to female NOD mice that were either 2 or 12 weeks old, the animals were protected from developing type 1 diabetes. NBI-6024. NBI-6024 is an altered peptide ligand based on the beta chain of the insulin molecule. In January 2000, Neurocrine and Taisho entered into an agreement in which Taisho had the option to obtain rights to the worldwide development and commercialization of NBI-6024. By October 2002, the agreement had been restructured, granting development and commercialization rights to Neurocrine in regions outside of Japan. NBI-6024 is currently in Phase II development in the United States and Europe and in Phase I trials in Japan. NBI-6024 has been shown to downregulate the immune response and reduce the incidence of diabetes in nonobese diabetic mice (Neurocrine Bioscience, Web site, March 2002). Immune cells harvested from diabetic patients react strongly to NBI-6024, whereas cells taken from nondiabetic patients show no reactivity (Neurocrine Bioscience, press release, January 2000). In a placebo-controlled Phase I/II study, 35 adolescent type 1 diabetics (aged 12–17 years) were randomized into two cohorts to determine the safety proÞle of NBI-6024 (Philotheou A, 2002). The Þrst cohort received treatment with placebo or escalating doses of NBI-6024 (0.1 mg, 1 mg, or 5 mg) once every two weeks over a period of eight weeks. The second cohort received treatment with placebo or NBI-6024 (1 mg or 5 mg) once every 2 weeks for 4 weeks, then once monthly for two months, for a total study period of 12 weeks. The most frequently reported adverse events in the treatment groups were upper respiratory infections and headaches. Three patients experienced serious adverse events: gastritis, hypoglycemic coma, and vomiting; however, none of these adverse events were attributed to the drug. In summary, the safety proÞle of NBI-6024 was not statistically different from placebo. Insulin Sensitizers Overview. Skeletal muscle is responsible for approximately 75% of total glucose uptake. Subjects with diminished sensitivity to insulin (or insulin resistance) are

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less able to absorb glucose into skeletal muscle, possibly due to a malfunction in the glucose transport mechanisms. While this pathology is associated with the etiology of type 2 diabetes, there is still debate as to whether insulin resistance plays a role in type 1 diabetes. Nevertheless, insulin sensitizing agents may potentially allow for a reduction in the dose of insulin used in the treatment of type 1 diabetes by improving patient responsiveness to insulin. Mechanism of Action. Although the role of insulin resistance in type 1 diabetes pathophysiology has yet to be clearly understood, many researchers speculate that it may contribute, albeit to a small extent, to disease pathology. The insulin sensitizers in development for type 1 diabetes act in various ways. For example, one investigational drug mimics the actions of insulin, while another acts on glucose transporters. These different approaches lead to the same result—the uptake of glucose by the main reservoirs of the body (muscles, liver, and adipose tissue). Somatokine. Insmed is developing Somatokine, a recombinant protein complex consisting of insulin-like growth factor-1 (IGF-1) and binding protein-3, as an adjunct therapy to insulin treatment in type 1 diabetics. It is formulated as a once-daily injection. In October 2001, the compound was licensed to WelÞde (now Mitsubishi Chemical) in Japan. In August 2002, Insmed entered into a manufacturing agreement with Avecia, a company based in the United Kingdom. Somatokine is currently in Phase II trials in the United States. IGF-1 (also known as somatomedin) is a trophic factor that originates mainly in the liver, but numerous other tissues synthesize it. It circulates at high levels in the bloodstream, eliciting many of the same physiological effects of insulin. Somatokine’s large structure will restrict it to the circulatory system, where it will serve as a reservoir of IGF-1. Insmed believes that insulin resistance is associated with IGF-1 deÞciency in type 1 diabetics, and that Somatokine can replenish the reserves. In a double-blind, placebo-controlled study in six type 1 diabetic adolescents, the efÞcacy of Somatokine as an insulin sensitizer was examined. In combination with their current insulin regimens, subjects were administered Somatokine via subcutaneous injections once daily for two days at various daily doses (placebo, 0.1 mg, 0.4 mg, 0.2 mg, or 0.8 mg) (Saukkonen T, 2003). To measure the level of insulin sensitivity induced by Somatokine, fasting insulin was measured by fasted variable rate insulin infusion clamp and a two-step hyperinsulineamic euglycemic clamp. Researchers observed dose-dependent increases in circulating IGF-1 levels, restoring IGF-1 levels to normal levels, along with reductions in insulin requirements. Insulin sensitivity was improved in the highest dosing group but not in the lower dosing groups. Safety and side-effect data were not described. T-1095. T-1095, an inhibitor of the sodium glucose cotransporter (SGLT), is being codeveloped by Johnson & Johnson and Tanabe. The licensing agreement grants worldwide rights to the development and marketing of T-1095 to Johnson

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& Johnson, excluding Japan and Asian countries where the rights are held by Tanabe. The compound is currently in Phase II trials in the United States, Europe, and Japan. Preclinical studies involving streptozocin (STZ)-induced diabetic rats and Zucker diabetic fatty (ZDF) rats suggest that T-1095 suppresses hyperglycemia, reduces fasting blood glucose levels, and improves insulin sensitivity. The most recent preclinical data in db/db mice treated with T-1095 for 11 days showed reductions in prandial and fasting plasma glucose levels with a concomitant decrease in urinary glucose excretion (Conway BR, 2003). No changes in body weight were observed. In another study, STZ-induced diabetic rats received one oral dose of T-1095. This treatment substantially reduced blood glucose levels as well as increased insulin-mediated glucose utilization. Increased glucose utilization, by the body as a whole as well as by the skeletal muscles, suggests that T-1095 improves insulin sensitivity. The researchers also observed a decrease in the rate of hepatic glucose production (Oku A, 2000[a]). In another preclinical study, db/db mice were given T-1095 with their food for 12 weeks (Arakawa K, 2001). The researchers reported decreases in hemoglobin A1c (HbA1c) and blood glucose. T-1095 also prevented the typical age-related decrease in plasma insulin levels. Only a minor effect on blood glucose levels was observed in drug-treated db/+ mice. This Þnding suggests that T-1095 will not reduce blood glucose to abnormally low levels when it is already within the normal range. Further, T-1095 suppressed the progression of diabetic neuropathy. In contrast to other insulin sensitizers, particularly the glitazones, T-1095 will not lead to weight gain (Oku A, 2000[a]). Given that an increase in insulin secretion is not needed for T-1095 to be effective, this agent is associated with only a low risk of hypoglycemia (Oku A, 2000[b]). Instead of allowing elevated levels of glucose to circulate through the blood, T-1095 inhibits the sodiumglucose transporter in the kidneys, resulting in glucose excretion through the urine (Oku A, 1999). This mechanism of action may increase urine volume, requiring the patient to consume more water. T-1095 will not be an effective therapy for patients who have lost kidney function. Amylin Agonists Overview. In 1901, Eugene L. Opie Þrst discovered islet amyloids (originally known as proteinacious deposits) in the pancreas of hyperglycemic patients. In 1986, these deposits were renamed islet amyloid polypeptides (IAPP), or amylin. Amylin, a 37-amino-acid peptide secreted (along with insulin) by beta cells, congregates to form Þbrils that are impervious to proteolytic digestion. Some researchers believe that amylin works in cooperation with insulin to maintain glucose homeostasis. Animal and human studies have demonstrated that amylin regulates glucose inßux into the circulation by reducing the rate of gastric emptying and by postprandially suppressing hepatic glucose production. Amylin agonists are seen as a way to treat type 1 diabetes in concert with insulin therapy.

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Mechanism of Action. Amyloid deposits—or amylin (a naturally occurring substance secreted by pancreatic beta cells)—in the diabetic pancreas have been implicated in the pathogenesis of type 2 diabetes (Epstein F, 2000). Amylin’s role in type 1 diabetes has not yet been elucidated. However, scientiÞc data suggest that amylin complements the actions of insulin and that its deÞciency plays a role in the onset of hyperglycemia. Hence, amylin agonists may improve the efÞciency of insulin therapy in the management of type 1 diabetes. Pramlintide. Pramlintide (Amylin Pharmaceuticals’ Symlin) is an amylin analogue that is being developed on the premise that amylin, like insulin, is deÞcient in type 2 diabetic patients and, therefore, introducing exogenous amylin (with insulin) will provide therapeutic value. Pramlintide is currently preregistered in the United States and in Phase III trials in Europe. In October 2001, the FDA’s Endocrinologic and Metabolic Drug Advisory Committee requested additional clinical and pharmacokinetic data before granting approval to pramlintide. The committee was concerned about cases of severe hypoglycemia reported during the Þrst month of therapy, which include two deaths of type 2 diabetes patients that may have resulted from hypoglycemia; the cases were presented by the FDA’s safety analyst during the committee meeting in July 2001. Currently, a dose-titration safety trial in type 1 diabetes patients and four pharmacokinetic studies are under way to address these issues. In December 2003, Amylin received a second approvable letter. The FDA required additional data to identify the patient population and pramlintide’s method of use in patients in whom there is either no increased risk of severe hypoglycemia or an added clinical beneÞt that outweighs the risks. In Europe, after discussions with the EMEA in 2002, it was clear that the FDA would require additional clinical data. But because the European system does not allow for amending the MAA, Amylin decided to withdraw the application. When subcutaneously injected with exogenous insulin, pramlintide slows the rate of stomach emptying, thereby slowing the rate at which glucose enters the bloodstream after a meal. It also reduces postprandial glucagon secretion from the pancreas and subsequently the release of glucose from stored liver glycogen. After a meal, these mechanisms help to minimize glucose levels in the blood (Weyer C, 2001[a]). A multicenter, randomized, placebo-controlled Phase III trial demonstrated improvements in glucose control after the addition of pramlintide to existing insulin therapeutic regimens in type 1 diabetics over one year (Amylin, press release, November 8, 1999). Patients were randomized to three treatment arms: (1) 60 mcg three times daily; (2) 60 mcg four times daily; or (3) or placebo (insulin only). The number of patients was not noted in the press release. After one year of therapy, patients in both treatment arms achieved HbA1c reductions of 0.4% compared with insulin. Approximately 40% of pramlintide-treated patients responded to therapy (that is, these patients achieved HbA1c reductions of 0.5% or more at four weeks) compared with 20% in the placebo arm. Additionally, responders in the Þrst group achieved HbA1c reductions of 0.7% after one year.

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Researchers observed that pramlintide-treated patients used lower insulin doses, while patients in the placebo arm required greater doses of insulin to maintain glycemic control. Further, patients treated with pramlintide improved weight control after one year, losing 3.5 pounds compared with a gain of 3.5 pounds in the placebo group. Nausea was the most frequently observed adverse event, but researchers noted that the incidence decreased over time. In a double-blind, randomized, placebo-controlled, open-extension study in 480 type 1 diabetics, results support the Þndings of the previous Phase III study. In this trial, pramlintide was added four times daily (120 or 240 mcg/day) to current insulin treatment regimens (Kolterman OG, 1999). Among those patients who responded to therapy (that is, patients who achieved HbA1c reductions of 0.5% or more at four weeks), the addition of pramlintide reduced HbA1c by 0.9–1.2% from baseline after two years of treatment. Among nonresponders, HbA1c reduction was 0.2%. Approximately 45% of pramlintide-treated patients experienced nausea compared with 17% of patients in the placebo group. In addition, 17% of pramlintide-treated patients experienced anorexia compared with 2% of patients in the placebo group. In another multicenter, double-blind study, 586 type 1 diabetics were randomized into four treatment regimens in addition to current insulin treatments: (1) 90 mcg twice daily; (2) 60 mcg three times daily; (3) 90 mcg three times daily; or (4) placebo (insulin alone) (Fineman M, 1999). At the end of 26 weeks, HbA1c in the treated groups were reduced by 0.4% to 0.6%. In total, 41–45% of the pramlintide-treated groups responded to therapy compared with 25% in the placebo arm (insulin alone). Insulin requirements of patients treated with a total daily dose of 180 mcg pramlintide decreased by 2.2–2.6% compared with baseline. Anorexia, nausea, vomiting, and hypoglycemia were higher in pramlintide-treated patients compared with placebo—20% versus 2%, 59% versus 15%, 18% versus 5%, and 27% versus 14%, respectively. In a multicenter, double-blind Phase III trial in 24 type 1 diabetics on continuous subcutaneous insulin infusion, results indicated that pramlintide reduced ßuctuations in glucose levels (Levetan C, 2002). In another study involving 296 type 1 patients receiving multiple daily injections of insulin or insulin pumps, adding pramlintide therapy reduced the amount of insulin by 12% (Amylin Pharmaceuticals, press release, May 19, 2003). It also reduced postprandial glycemia—an effect that lasted throughout the seven-month study. Titration of doses reduced the impact of nausea that has been observed upon initiation of therapy. REFERENCES Abbas R, et al. Oral insulin: pharmacokinetics and pharmacodynamics of human insulin following oral administration of an insulin/delivery agent capsule in healthy volunteers. 62nd ScientiÞc Sessions of the American Diabetes Association Abstracts. 2002;51(suppl 2):Abstract 197-OR. Akazawa Y. Prevalence and incidence of diabetes mellitus by WHO criteria. Diabetes Research and Clinical Practice. 1994;24:S23–S27.

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Amos A, et al. The rising global burden of diabetes and its complications: Estimates and projections to the year 2010. Diabetic Medicine. 1997;14:S7–S85. Amschler DH. The alarming increase of type 2 diabetes in children. Journal of School Health. 2002;72:39–41. Arakawa K, et al. Improved diabetic syndrome in C57BL/KsJ-db/db mice by oral administration of the Na(+)-glucose cotransporter inhibitor T-1095. British Journal of Pharmacology. 2001;132:578–586. Arbit E. Oral insulin. Diabetes Technology Meeting at Hyatt Regency; November 8, 2003; San Francisco, CA. www.emisphere.com. Accessed on January 9, 2004. Aschner P. Current concepts of diabetes mellitus. International Ophthalmology Clinics. 1998;38:1–10. Atkinson MA, et al. Type 1 diabetes: new perspectives on disease pathogenesis and treatment. Lancet. 2001;358:221–229. Becker RHA, et al. Pharmacodynamics and pharmacokinetics of a new, rapid-acting insulin analogue, insulin glulisine. 63rd ScientiÞc Sessions of the American Diabetes Association Abstracts. 2003;52(suppl 1):Abstract 471-P. [a] Becker RHA, et al. The effect of smoking cessation and subsequent resumption on absorption of inhaled insulin (Exubera). 63rd ScientiÞc Sessions of the American Diabetes Association Abstracts. 2003;52(suppl 1):Abstract 160-OR. [b] Biondi-Zoccai GG, et al. Atherosclerosis, inßammation and diabetes. Journal of American College of Cardiology. 2003;41(7):1071–1077. Bode B, et al. Comparison of insulin aspart with buffered regular insulin and insulin lispro in continuous subcutaneous insulin infusion: a randomized study in type 1 diabetes. Diabetes Care. 2002;25(3):439–444. Bode BW, et al. EfÞcacy, safety, and pump compatibility of insulin aspart used in continuous subcutaneous insulin infusion therapy in patients with type 1 diabetes. Diabetes Care. 2001;24:69–72. Boehm BO, et al. Premixed insulin aspart 30 vs. premixed human insulin 30/70 twice daily: a randomized trial in type 1 and type 2 diabetic patients. Diabetic Medicine. 2002;19:393–399. Bottino R, et al. Gene- and cell-based therapeutics for type 1 diabetes mellitus. Gene Therapy. 2003;10:875–889. Brunner GA, et al. Dose-response relation of liquid aerosol inhaled insulin in type 1 diabetic patients. Diabetologia. 2001;44:305–308. Clement S, et al. Presentation at the 61st ScientiÞc Sessions of the American Diabetes Association Abstracts. 2001;50(suppl 2):Abstract 109. Clement S, et al. Effects of multiple doses of orally administered hexyl insulin M2 (HIM2) on postprandial blood glucose (PPG) concentrations in type 1 diabetic (T1) patients. 62nd ScientiÞc Sessions of the American Diabetes Association Abstracts. 2002; 51(suppl 2):Abstract 11-LB. Clement S, et al. Oral modiÞed insulin (HIM2) in patients with type 1 diabetes mellitus: results from a Phase I/II clinical trial. Metabolism. 2004;53:54–58. Centers for Disease Control and Prevention. Diabetes Projects: Children and Diabetes. 2000. www.cdc.gov/diabetes/pubs/factsheets/search.htm. Accessed April 7, 2003. Cherubini V, et al. Incidence of IDDM in the Marche Region, Italy. Diabetes Care. 1994;17:432–435.

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Cherubini V, et al. Large incidence variation of type I diabetes in central-southern Italy 1990–1995: lower risk in rural areas. Diabetologia. 1999;42:789–792. Chueca M, et al. Incidence of type I diabetes mellitus in Navarre, Spain (1975–91). Acta Paediatrica. 1997;86:632–637. Conway BR, et al. Combination of a sodium-glucose cotransporter inhibitor and a PPAR gamma agonist on metabolic parameters and body weight in female db/db mice. 63rd ScientiÞc Sessions of the American Diabetes Association Abstracts. 2003;52(suppl 1):Abstract 493-P. Dabelea D, et al. Type 2 diabetes mellitus in minority children and adolescents: an emerging problem. Pediatric Endocrinology. 1999;28:709–729. Dahlquist G. The aetiology of type 1 diabetes: an epidemiological perspective. Acta Pediatric Supplement. 1998; 425:5–10. Devendra D, et al. Immunologic endocrine disorders. Journal of Allergy and Clinical Immunology. 2003;111:S624–S636. Diabetes Control and Complications Trial (DCCT) Research Group. Implementation of treatment protocols in the Diabetes Control and Complications Trial. Diabetes Care. 1995;18:361–376. Dokheel TM. An epidemic of childhood diabetes in the United States? Evidence from Allegheny County, Pennsylvania, Pittsburgh Diabetes Epidemiology Research Group. Diabetes Care. 1993;16:1606–1611. Doutreix J, Levy-Marchal C. Presentation on insulin-dependent diabetes mellitus in children at diagnosis: the experience of the French incidence registry. Revue d Epidemiologie et de Sante Publique. 1996;44(suppl 1):90–96. Elias D, et al. Peptide therapy for diabetes in NOD mice. Lancet. 1994;343:704–706. Epstein F. Islet amyloid and type 2 diabetes mellitus. New England Journal of Medicine. 2000;343:411–419. EURODIAB ACE Study Group. Variation and trends in incidence of childhood diabetes in Europe. Lancet. 2000;355:873–876. Expert Committee on the Diagnosis and ClassiÞcation of Diabetes Mellitus. Report of the Expert Committee on the diagnosis and classiÞcation of diabetes mellitus. Diabetes Care. 2003;26:S5–S20. Fabietti PG, et al. Prevalence of known diabetes mellitus in a central region of Italy. Diabetes, Nutrition and Metabolism—Clinical and Experimental. 1996;9:3–8. Fagot-Campagna A. Type 2 diabetes among North American children and adolescents: an epidemiologic review and public health perspective. Journal of Pediatrics. 2000;136:664–672. Fineman M, et al. Effects of six months administration of pramlintide as an adjunct to insulin therapy on metabolic control in people with type 1 diabetes. 59th ScientiÞc Sessions of the American Diabetes Association Abstracts. 1999;48(suppl1):113–114. Friedman E. Advanced glycosylated end products and hyperglycemia in the pathogenesis of diabetic complications. Diabetes Care. 1999;22(suppl 2):65B–71B. Gan D, ed. Diabetes Atlas, 2000. Brussels, Belgium: International Diabetes Federation; 2001. Garancini MP, et al. Epidemiology of known diabetes in Lombardy, North Italy. Clinical characteristics and methodological aspects. Acta Diabetologica. 1995;32:268–272.

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Genuth S. Metabolism VI: Diabetes Mellitus; August 2001. www.acpmedicine.com. Accessed September 17, 2003. Goday A, et al. Incidence of type 1 (insulin-dependent) diabetes mellitus in Catalonia, Spain. The Catalan Epidemiology Diabetes Study Group. Diabetologia. 1992;35:267–271. Green A, et al. Epidemiological studies of diabetes mellitus in Denmark: 5. Mortality and causes of death among insulin-treated diabetic patients. Diabetologia. 1984;26:190–194. Greenbaum CJ. Insulin resistance in type 1 diabetes. Diabetes/Metabolism Research and Reviews. 2002;18:192–200. GrifÞn E, et al. A link between diabetes and atherosclerosis: glucose regulates expression of CD36 at the level of translation. Nature Medicine. 2001;7:840–846. Guevara-Aguirre J, et al. Oral insulin spray as an add-on therapy in combination with failing oral agents (OHAs) for treatment of type 2 diabetes. 62nd ScientiÞc Sessions of the American Diabetes Association Abstracts. 2002;51(suppl 2):Abstract 401-P. Guevara-Aguirre J, et al. Dose-ranging study of oralin in healthy subjects. 63rd ScientiÞc Sessions of the American Diabetes Association Abstracts. 2003;52(suppl 1):Abstract 445-P. [a] Guevara-Aguirre J, et al. Reproducibility of oralin absorption in type 1 diabetics on 3 different occasions. 63rd ScientiÞc Sessions of the American Diabetes Association Abstracts. 2003;52(suppl 1):Abstract 446-P. [b] Gujral JS, et al. Childhood-onset diabetes in the White and South Asian population in Leicestershire, UK. Diabetic Medicine. 1994;11:570–572. Hansen JR, et al. Type 2 diabetes in youth: a growing challenge. Clinical Diabetes. 2000;18:52–56. Harris MI, et al. Self-monitoring of blood glucose by adults with diabetes in the U.S. population. Diabetes Care. 1993;16:1116–1123. Harris MI, et al. Prevalence of adult-onset IDDM in the U.S. Diabetes Care. 1994;17:1337–1340. Harvey JN, et al. Estimation of the prevalence of diagnosed diabetes from primary care and secondary care source data: comparison of record linkage with capture-recapture analysis. Journal of Epidemiology and Community Health. 2002;56:18–23. Heise TC, et al. Lower within-subject variability of insulin detemir in comparison to NPH insulin and insulin glargine in subjects with type 1 diabetes. 63rd ScientiÞc Session of the American Diabetes Association Abstracts. 2003:52(suppl 1):Abstract 518-P. Henry R, et al. Within-subject variability of inhaled insulin (Exubera) versus subcutaneous regular insulin in elderly obese patients with type 2 diabetes mellitus. 63rd ScientiÞc Sessions of the American Diabetes Association Abstracts. 2003;52(suppl 1):Abstract 448-P. Henry RR, et al. Inhaled insulin using the AERx insulin diabetes management system in healthy and asthmatic patients. Diabetes Care. 2003;26:764–769. Hermansen K, et al. Comparison of the soluble basal insulin analog detemir with NPH insulin. Diabetes Care. 2001:24:296–301. Herold KC, et al. Anti-CD3 monoclonal antibody in new-onset type 1 diabetes mellitus. New England Journal of Medicine. 2002;346:1692–1698. [a]

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Herold, KC, et al. Treatment with anti-CD3 monoclonal antibody arrests the loss of insulin responses and results in improved metabolic control in patients with type 1 diabetes. 62nd ScientiÞc Session of the American Diabetes Association Abstracts. 2002:51(suppl 2):Abstract 183-OR. [b] Himmelmann A, et al. The impact of smoking on inhaled insulin. Diabetes Care. 2003;26:677–682. Home PD, et al. Improved glycemic control with insulin aspart. Diabetes Care. 1998;21:1904–1909. Home PD, et al. Comparative pharmacokinetics and pharmacodynamics of the novel rapid-acting insulin analogue, insulin aspart, in healthy volunteers. European Journal of Clinical Pharmacology. 1999; 55:199–203. Home PD, et al. Insulin aspart vs. human insulin in the management of long-term blood glucose control in type 1 diabetes mellitus: a randomized controlled trial. Diabetic Medicine. 2000;17(11):762–770. Home PD, et al. Insulin detemir offers improved glycemic control, less weight gain, and ßexible timing of administration compared to NPH insulin. 63rd ScientiÞc Session of the American Diabetes Association Abstracts. 2003:52(suppl 1):Abstract 522-P. Holcombe JH, et al. Comparison of insulin lispro with regular human insulin for the treatment of type 1 diabetes in adolescents. Clinical Therapeutics. 2002;24:629–638. Jacobsen LV, et al. Pharmacokinetics of insulin detemir in subjects with renal or hepatic impairment. 62nd ScientiÞc Sessions of the American Diabetes Association. 2002;51(suppl 2):Abstract 413-P. Karvonen M, et al. Type 1 diabetes: global epidemiology. In: Ekoe JM, et al, eds. The Epidemiology of Diabetes Mellitus: An International Perspective. Chichester, England: John Wiley & Sons Ltd; 2001. Kaufman FR. Type 2 diabetes mellitus in children and youth: a new epidemic. Journal of Pediatric Endocrinology and Metabolism. 2002;15:737–744. Kawasaki E, Nagataki S. Japanese type 1 diabetic syndrome. Diabetes Metabolism Reviews. 1996;12:175–194. Kidron M. A novel per-oral insulin formulation for human subjects. 62nd ScientiÞc Session of the American Diabetes Association Abstracts. 2002:51(suppl 2):Abstract 526-P. Kiess W, et al. Type 2 diabetes mellitus in children and adolescents: a review from a European perspective. Hormone Research. 2003;59(suppl 1):77–84. Kitagawa T, et al. Epidemiology of type 1 (insulin-dependent) and type 2 (non-insulindependent) diabetes mellitus in Japanese children. Diabetes Research and Clinical Practice. 1994;24(suppl):S7–S13. Kolterman OG, et al. Pramlintide as an adjunct to insulin therapy in patients with type 1 diabetes improved glycemic control over 2 years. 59th ScientiÞc Sessions of the American Diabetes Association Abstracts. 1999;48(suppl 1):104–105. Kurtzhals P, et al. Correlations of receptor binding and metabolic and mitogenic potencies of insulin analogues designed for clinical use. Diabetes. 2000;49:999–1005. Laakso M, et al. Age of onset and type of diabetes. Diabetes Care. 1985;8:114–117. LaPorte RE, et al. Prevalence and incidence of insulin-dependent diabetes. In Harris MI, et al., eds. Diabetes in America. 2nd ed. Washington, DC: National Institutes of Health (NIH pub. no. 95–1468). 1995;37–46.

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Lernmark A, et al. A Phase II clinical trial involving alum-formulated GAD65 vaccination of GAD65-autoantibody positive adult diabetes patients. 63rd ScientiÞc Session of the American Diabetes Association Abstracts. 2003:52(suppl 1):Abstract 32-LB. Leury D, et al. Physicians’ reaction to inhaled insulin, a new insulin delivery system. 63rd ScientiÞc Sessions of the American Diabetes Association Abstracts. 2003;52(suppl 1):Abstract 456-P. Levetan C, et al. Reduced glucose ßuctuations following 4 weeks of pramlintide treatment in patients with type 1 diabetes intensively treated with insulin pumps. 62nd ScientiÞc Sessions of the American Diabetes Association Abstracts. 2002;51(suppl 2):106. Liao D, et al. Lower heart rate variability is associated with the development of coronary heart disease in individuals with diabetes: the Atherosclerosis Risk in Communities (ARIC) study. Diabetes. 2002;51(12):3524–3531. MayÞeld J. Diagnosis and classiÞcation of diabetes mellitus: new criteria. American Family Physician. 1998;58:1355–1370. Mazzella M, et al. Incidence of type I diabetes in the Liguria Region, Italy. Results of a prospective study in a 0- to 14-year age-group. Diabetes Care. 1994;17:1193–1196. Melton LJ, et al. Incidence of type 1 diabetes mellitus by clinical type. Diabetes Care. 1983;6:75–86. Modi P, et al. Oral insulin spray as a meal insulin in treatment of type 2 diabetes. 62nd ScientiÞc Sessions of the American Diabetes Association Abstracts. 2002;51(suppl 2):Abstract 439-P. Modi P, et al. Comparison of oral insulin spray vs. s.c. bolus in patients with type 1 diabetes stabilized on insulin pump (I.V. drip). 63rd ScientiÞc Sessions of the American Diabetes Association Abstracts. 2003;52(suppl 1):Abstract 458-P. Molbak AG, et al. Incidence of insulin-dependent diabetes mellitus in age groups over 30 years in Denmark. Diabetic Medicine. 1994;11:650–655. Morales-Perez FM, et al. Incidence of type 1 diabetes among children and young adults (0–29 years) in the province of Badajoz, Spain, during 1992–1996. Acta Paediatrica. 2000;89:101–104. Muggeo M, et al. The Verona diabetes study: a population-based survey on known diabetes mellitus prevalence and 5-year all-cause mortality. Diabetologia. 1995;38:318–325. Nawano M, et al. Hyperglycemia contributes insulin resistance in hepatic and adipose tissue but not skeletal muscle of ZDF rats. American Journal of PhysiololgyEndocrinolology and Metabolism. 2000;278(3):E535–E543. Neu A, et al. Rising incidence of type 1 diabetes in Germany. Diabetes Care. 2001;24:785–787. Oku A, et al. T-1095, an inhibitor of renal Na+-glucose cotransporters, may provide a novel approach to treating diabetes. Diabetes. 1999;48:1794–1800. Oku A, et al. Antidiabetic effect of T-1095, an inhibitor of Na(+)-glucose cotransporter, in neonatally streptozotocin-treated rats. European Journal of Pharmacology. 2000;391:183–192. [a] Oku A, et al. Correction of hyperglycemia and insulin sensitivity by T-1095, an inhibitor of renal Na+-glucose cotransporters, in streptozotocin-induced diabetic rats. Japanese Journal of Pharmacology. 2000;84(3):351–354. [b]

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Russel-Jones D, et al. Once-daily dosing with insulin detemir offers advantages compared to NPH insulin in subjects with type 1 diabetes. 63rd ScientiÞc Sessions of the American Diabetes Association Abstracts. 2003;52(suppl 1):Abstract 565-P. Saukkonen T, et al. Combined recombinant human rhIGF-1/rhIGFBP-3 suppresses growth hormone secretion and improves insulin sensitivity in adolescents with type 1 diabetes. 63rd ScientiÞc Sessions of the American Diabetes Association Abstracts. 2003;50(suppl 2):Abstract 569-P. Schwartz S, et al. Replacement of failing sulfonylurea drugs with oral insulin, a long-term efÞcacy study in treatment of type 2 diabetes. 61st ScientiÞc Sessions of the American Diabetes Association Abstracts. 2001;50(suppl 2):Abstract 181-OR. Serrano-Rios M, et al. Incidence of type 1 (insulin-dependent) diabetes mellitus in subjects 0–14 years of age in the Comunidad of Madrid, Spain. Diabetologia. 1990;33:422–424. Sharma A, et al. Autoimmune perspective of insulin-dependent diabetes mellitus: cytokines as therapeutic targets. Drugs of the Future. 2003;28(1):31–42. Skyler JS, et al. EfÞcacy of inhaled human insulin in type 1 diabetes mellitus: a randomised proof-of-concept study. Lancet. 2001;357:331–335. Skyler JS, et al. EfÞcacy and safety of inhaled insulin (Exubera) compared to subcutaneous insulin therapy in an intensive insulin regimen in patients with type 1 diabetes: results of a 6-month, randomized, comparative trial. 62nd ScientiÞc Sessions of the American Diabetes Association Abstracts. 2002;51(suppl 1):Abstract 540-P. Staines A, et al. The epidemiology of diabetes mellitus in the United Kingdom: the Yorkshire Regional Childhood Diabetes Register. Diabetologia. 1993;36:1282–1287. Surguladze D, et al. Insulinization of the liver in normal mice by oral delivery of HIM2, a novel modiÞed insulin. 62nd ScientiÞc Sessions of the American Diabetes Association Abstracts. 2002;51(suppl 1):Abstract 542-P. Tajima N, et al. Lack of regional variation in IDDM risk in Japan. Diabetes Care. 1993;16:796–800. Tisch R, et al. A glutamic acid decarboxylase 65-speciÞc Th2 cell clone immunoregulates autoimmune diabetes in nonobese diabetic mice. Journal of Immunology. 2001;166:6925–6936. Weyer C. Amylin replacement with pramlintide as an adjunct to insulin therapy in type 1 and type 2 diabetes mellitus: a physiological approach toward improved metabolic control. Current Pharmaceutical Design. 2001;7(14):1353–1373. World Health Organization (WHO) Consultation. DeÞnition, diagnosis, and classiÞcation of diabetes mellitus and its complications. I. Diagnosis and classiÞcation of diabetes mellitus. Geneva, Switzerland: World Health Organization. 1999;99:2. Zinman B, et al. Effectiveness of human ultralente versus NPH insulin in providing basal insulin replacement for an insulin lispro multiple daily injection regimen. Diabetes Care. 1999;22:603–608.

Type 2 Diabetes

ETIOLOGY AND PATHOPHYSIOLOGY Introduction Type 2 diabetes mellitus stems from impaired insulin activity and failure of pancreatic beta cells to produce a sufÞcient compensatory insulin secretory response. (In contrast, type 1 diabetes mellitus is marked by the absolute inability of pancreatic beta cells to produce insulin.) These conditions result in the progressive worsening of insulin resistance (IR) in muscle and liver tissue and impaired glucose tolerance (IGT) as patients transition from a prediabetic state toward actual diabetes. During the progression to type 2 diabetes, the pancreatic beta cells no longer produce sufÞcient quantities of insulin to meet the demands of insulinresistant tissues, and hyperglycemia (elevated serum glucose) ensues. Glucose toxicity, in turn, is responsible for much of the morbidity and mortality associated with both type 1 and type 2 diabetes, which include microvascular complications such as peripheral neuropathic pain, retinopathy, and nephropathy as well as macrovascular complications that markedly increase the risk of cardiovascular disease (CVD). Etiology Type 2 diabetes is a complex metabolic disorder. Although researchers have identiÞed important genetic pathways and lifestyle factors that contribute to its development, the pathophysiologic mechanisms that lead to insulin resistance and beta-cell failure remain poorly understood. Table 1 lists known risk factors for type 2 diabetes used by physicians to identify susceptible individuals. Many Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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TABLE 1. Risk Factors for Type 2 Diabetes Risk Factor Family history of type 2 diabetes Ethnicity

History of gestational diabetes Metabolic syndrome

Potential Impact Individuals with a first-degree relative who is a type 2 diabetic have a 40% lifetime risk of developing type 2 diabetes. The prevalence of type 2 diabetes is higher in African, Native American, Asian, Pacific Islander, and Hispanic ethnic populations than in Caucasian populations. Insulin resistance can develop during the late stages of pregnancy as a result of underlying impairments in insulin secretory activity. These individuals are at greater risk of developing type 2 diabetes later in life. Individuals with obesity, elevated blood pressure, and dyslipidemia (low HDL and/or elevated triglycerides) are at great risk of developing insulin resistance and type 2 diabetes. These conditions cluster together in a high percentage of patients.

HDL = High-density lipoprotein cholesterol. Source: Adapted from the National Diabetes Information Clearinghouse, National Institutes of Diabetes and Digestive and Kidney Diseases; http://diabetes.niddk.nih.gov/dm/pubs/riskfortype2, accessed May 13, 2005.

of these risk factors are also strongly linked to obesity, a condition thought to precede the development of type 2 diabetes in the majority of patients because of the role of excess fat in triggering insulin resistance. Genetic Factors. Substantial evidence suggests that type 2 diabetes is a heritable disorder. First-degree relatives of a patient with type 2 diabetes have a 40% lifetime risk of developing the disease (National Diabetes Information Clearinghouse). Furthermore, scientists have identiÞed numerous candidate genes for type 2 diabetes from diabetic individuals, studies of genetically susceptible populations, and studies employing animal models of diabetes. The strongest genetic links to type 2 diabetes exist in individuals with maturity-onset diabetes of the young (MODY), a rare form of type 2 diabetes that stems from inheriting defective genes required for insulin secretion (Tsakiris D, 2004; Timsit J, 2005). Type 2 diabetics with these genetic defects typically have a lean diabetic phenotype (are not obese) and develop type 2 diabetes relatively early in life (before age 25). Table 2 lists some of the known single-gene defects that lead to MODY. However, researchers emphasize that there is no single genetic cause of type 2 diabetes for the majority of type 2 diabetic patients. In most cases, the etiology of type 2 diabetes involves numerous genes that act on multiple physiological pathways. Furthermore, genetics alone do not explain the rapid global rise in the prevalence of type 2 diabetes over the past 30 years (Zimmet P, 2001). Thus, scientists must also unravel the complex gene-environment interactions that contribute to the development of type 2 diabetes in genetically susceptible populations. These factors have made it difÞcult for companies to develop targeted treatments for the highly heterogeneous population of type 2 diabetic patients. Lifestyle Factors. In addition to genetic susceptibility, lifestyle factors contribute signiÞcantly to the development of type 2 diabetes. Increased adoption of

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TABLE 2. Single-Gene Defects That Cause Maturity-Onset Diabetes of the Young Mutation

Gene Name

Function in Type 2 Diabetes

MODY1 Hepatic nuclear Transcription factor required to activate the Kir6.2 gene, factor-4alpha (HNF-4α) which encodes an ATP-dependent potassium-channel subunit required for insulin secretion by pancreatic beta cells (Gupta RK, 2005). MODY2 Glucokinase Converts glucose to glucose-6-phosphate, which stimulates insulin secretion. Patients present with mild hyperglycemia from birth (Timsit J, 2005). MODY2 is often diagnosed in cases of gestational diabetes. MODY3 Hepatic nuclear Mutations in HNF-1α are one of the most common factor-1alpha (HNF-1α) causes of MODY. Patients exhibit severe hypoglycemia after puberty and are sometimes misdiagnosed as having type 1 diabetes mellitus (Timsit J, 2005). Patients retain sensitivity to sulfonylureas (insulin secretagogues; see Chapter 4, ‘‘Current Therapies’’). MODY4 Insulin promoter factor-1 Also called pancreatic-duodenal homeobox 1 (PDX-1), a (IPF-1) protein required for normal pancreatic development (Servitja JM, 2004). Mouse studies show a role for IPF-1 in modulating GLP-1-receptor function (Li Y, 2005). MODY5 Hepatic nuclear Form of MODY associated with developmental defects of factor-1beta (HNF-1β) the pancreas, liver, kidneys, and genital tract. Mice deficient in HNF-1β have defective insulin secretion due to defective glucose sensing (Wang L, 2004). MODY6 Neurogenic differentiation Transcription factor also known as BETA2, involved in factor-1 (NeuroD1) both neurological development and pancreatic beta-cell differentiation. NeuroD1 can regulate expression of the glucokinase gene (Moates JM, 2003). ATP = Adenosine triphosphate. MODY = Mature-onset diabetes of the young. GLP-1 = Glucagon-like peptide 1. Note: Full source citations appear in ‘‘References.’’

Western diets, the availability of fat- and calorie-rich foods, and reduced physical activity stemming from the rising use of automobiles and other modern conveniences have contributed to the sharp rise in the prevalence of obesity and type 2 diabetes over the past 50 years. Obesity Þgures prominently in both the etiology of type 2 diabetes and the etiology of metabolic syndrome (a clustering of cardiovascular risk factors that include obesity, insulin resistance, dyslipidemia, and hypertension) (Eckel RH, 2005; for more information, see the section on “Pathophysiology”). Obesity. Obesity is a metabolic disease characterized by an imbalance between energy intake and energy output. Excessive caloric intake (i.e., from overeating) combined with low energy expenditure (i.e., from lack of physical activity)

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result in a net gain of surplus calories that are metabolized into free fatty acids (FFA) and triglycerides (TG) and subsequently stored in adipocytes (fat cells). Obese individuals exhibit both increased adipocyte size and increased numbers of adipocytes, which form the basis of fat deposits in subcutaneous and visceral (abdominal) fat tissue. Adipocytes were originally thought to serve only as a storage depot for triglycerides. However, emerging research has revealed adipose tissue (especially visceral fat) to be a highly active source of hormones, cytokines, and other metabolic factors. Although low-level chronic inßammation has long been associated with obesity and type 2 diabetes, it is now becoming increasingly clear that these factors secreted by adipocytes, collectively termed adipokines, can trigger insulin resistance and impaired glucose tolerance (Wellen KE, 2005). Table 3 lists several key adipokines and the roles they play in the etiology and pathogenesis of type 2 diabetes. In addition to releasing adipokines into the circulation, adipocytes are responsible for increasing plasma levels of circulating FFAs. Insulin-resistant adipocytes exhibit elevated rates of lipolysis, which results in increased conversion of TGs to FFAs and glycerol. High levels of circulating FFAs have been implicated in the development of muscle and hepatic insulin resistance (Boden G, 1997; Machann J, 2004). Excess FFAs in skeletal muscle tissue inhibit insulin-mediated uptake of glucose by interfering with normal insulin signaling (McGarry JD, 2002; Schulman GI, 2000). Meanwhile, FFAs stimulate hepatic gluconeogenesis, an activity that is normally inhibited by the actions of insulin. Table 4 lists key genes involved in altered lipid metabolism in type 2 diabetics. Lack of Physical Activity. Sedentary lifestyles have been implicated in the rapid rise in the prevalence of type 2 diabetes over the past 50 years. Indeed, regular physical activity has been identiÞed as an important factor in maintaining insulin sensitivity in liver and muscle tissue. A prospective study of 487 Finnish patients with impaired glucose tolerance (IGT) who were followed for an average of four years demonstrated that individuals who participated in regular moderate exercise had 63–65% reduced risk of developing type 2 diabetes than those who did not exercise regularly (Laaksonen DE, 2005). Similar results were observed in the Da Qing Impaired Glucose Tolerance (IGT) and Diabetes Study, which followed 577 individuals diagnosed with IGT at baseline for a period of six years. Patients were randomized to diet, exercise, diet and exercise, or control groups and screened every two years for progression to type 2 diabetes. Individuals in the diet, exercise, and combined diet and exercise groups exhibited 31%, 46%, and 42%, respectively, reduced risk of developing type 2 diabetes compared with patients in the control groups (Pan XR, 1997). Other Factors. The majority of patients with type 2 diabetes develop the disease as a result of obesity, lack of physical activity, and/or genetic susceptibility to insulin resistance. However, type 2 diabetes can also develop as a secondary manifestation of another disease. Loss of pancreatic beta-cell mass and secretory

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TABLE 3. Key Proteins Secreted by Adipocytes with Roles in Type 2 Diabetes Molecule Leptin

Adiponectin

Resistin

TNF-alpha

Role in Type 2 Diabetes

Comment

Leptin signals levels of fat stores to Leptin was developed by Amgen the hypothalamus, where it as an antiobesity agent but functions upstream of several was later discontinued regulators of appetite in the brain. because of poor efficacy. Leptin receptors are also located on Several other companies have adipocytes and other tissues. leptin analogues and Mice lacking the gene for leptin leptin-receptor agonists in become obese, hyperinsulinemic, preclinical development. and insulin resistant. Mouse studies indicate that leptin has a role in neurobiological programming by serving as a link between fetal and neonatal nutritional status and hypothalamic development (Bouret SG, 2004; Pinto S, 2004). Regulates fat and glucose The protein is structurally similar metabolism by enhancing insulin to TNF-alpha and is sensitivity and stimulating uptake upregulated following of free fatty acids. Mechanism of treatment with PPAR-gamma action involves activation of agonists. AMP-activated protein kinase There is an inverse relationship (AMPK). between adiponectin levels Adiponectin receptors are found in and insulin resistance. Obese skeletal muscle tissue and the and insulin-resistant liver. individuals have low levels of Adiponectin may also act centrally to adiponectin; lean individuals regulate energy homeostasis and have high levels. body weight (Berg AH, 2002; Pharmaceutical companies are Bouskila M, 2005; Qi Y, 2004). evaluating increased adiponectin production as a secondary endpoint in clinical trials of weight-loss drugs and antidiabetic agents. The normal function of resistin in Resistin is found in high humans is unknown. circulating levels in both obese Promotes inflammation by and insulin-resistant up-regulating TNF-alpha and individuals. IL-6. Exogenous administration of Mouse studies suggest that resistin recombinant resistin in mice regulates fasting plasma glucose leads to insulin resistance in levels. liver and muscle tissue (Banerjee RR, 2004; Steppan CM, 2004). TNF-alpha is a strong inflammatory Chronic treatment with infliximab cytokine implicated in (Johnson & atherosclerosis. Johnson/Schering’s Remicade, TNF-alpha impairs insulin signaling an anti-TNF-alpha antibody) by inhibiting the function of may improve insulin sensitivity insulin-receptor substrate 1 (Yazdani-Biuki B, 2004). (IRS-1).

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TABLE 3. (continued) Molecule

Role in Type 2 Diabetes

Interleukin-6 (IL-6)

IL-6 is a potent proinflammatory cytokine that is associated with glucose intolerance and insulin resistance.

11betahydroxysteroid dehydrogenase (11beta-HSD)

11beta-HSD is an enzyme that converts cortisone to its active form, cortisol. Glucocorticoids stimulate liver gluconeogenesis, inhibit glucose uptake by muscle cells and adipocytes, and mobilize free fatty acids from adipocytes.

Visfatin

Visfatin binds to the insulin receptor and acts as an insulin mimetic but does not compete with insulin. Visfatin is believed to work together with adiponectin and insulin to improve insulin sensitivity and inhibit hepatic glucose production (Fukuhara A, 2005).

Comment IL-6 is also known as B-cell stimulatory factor-2 (BSF-2) and interferon-2 (IFN-2). May be responsible for the high levels of C-reactive protein (CRP) observed in obese individuals. CRP is a marker for generalized, systemic inflammation. Patients with Cushing’s disease (arising from cortisol excess) have metabolic profiles similar to those of obese patients. Several companies are developing inhibitors of 11beta-HSD as treatments for type 2 diabetes. All agents are in preclinical development. Visfatin is expressed more highly by visceral adipocytes than subcutaneous adipocytes. Also known as pre-B-cell colony-enhancing factor (PBEF). Mice deficient in visfatin develop both fasting and postprandial hyperglycemia but only a minor defect in processing oral glucose.

Note: Full source citations appear in ‘‘References.’’ Source: Adapted from Kershaw EE, 2004; and Bays H, 2004.

activity can occur as a result of pancreatic cancer, chronic pancreatitis, or cystic Þbrosis. Likewise, many disorders that disrupt normal endocrine function or regulation disrupt insulin signaling and beta-cell secretory activity. Table 5 lists possible nongenetic causes of type 2 diabetes. Pathophysiology The pathophysiology of type 2 diabetes remains an area of active research. Insulin resistance is widely recognized as one of the earliest steps in the progression of type 2 diabetes (Figure 1). As muscle and liver tissue become increasingly resistant to the effects of insulin, the body’s ability to metabolize glucose becomes compromised. In patients with prediabetes and early type 2 diabetes, the pancreatic beta cells are able to compensate for the insulin resistance by secreting

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TABLE 4. Key Genes Involved in Altered Lipid Metabolism in Type 2 Diabetes Gene Peroxisome proliferator-activated receptor (PPAR)-gamma

Beta-3-adrenergic receptor

Hormone-sensitive lipase (HSL)

Adipose triglyceride lipase (ATGL)/desnutrin/calciumindependent phospholipase A2 (iPLA2)

Potential Role in Type 2 Diabetes PPAR-gamma agonists are an important class of oral antidiabetic agents with effects on glucose homeostasis, lipid metabolism, and insulin sensitivity (see ‘‘Current Therapies’’). A meta-analysis demonstrated that individuals harboring the Pro12Ala polymorphism of the PPAR-gamma gene have a statistically significant increase in the relative risk (RR-1.25) of developing type 2 diabetes. The Pro12Ala polymorphism may be present in as much as 25% of the general population (Altschuler D, 2000). Beta-3-adrenergic receptors are expressed in brown adipose tissue, where they regulate thermogenesis and lipolysis. Several polymorphisms in this gene have been identified in families susceptible to type 2 diabetes (Mitchell B, 1998; Oizumi T, 2001; Parikh H, 2004). Several companies are developing novel therapeutics for type 2 diabetes and obesity that target these receptors. Beta-adrenergic receptors stimulate lipolysis by activating HSL enzymes in adipocytes. Hormone-sensitive lipase responds to signals transmitted by the beta-3-adrenergic receptors. Mice lacking HSL are insulin-resistant and fail to secrete insulin in response to IV-administered glucose (Roduit R, 2001). Individuals harboring the HSLi6 polymorphism exhibit reduced rates of adipocyte lipolysis (Hoffstedt J, 2001). ATGL is the second enzyme discovered that can hydrolyze triglycerides in mammalian adipose tissue (Zimmermann R, 2004). HSL was previously thought to be the only enzyme involved in the lipolytic pathway. ATGL is regulated by tumor necrosis factor-alpha and insulin in mouse adipocytes (Kralisch S, 2005).

Note: Full source citations appear in ‘‘References.’’

greater amounts of insulin to maintain normal glycemic control. This stage of disease is largely silent because there are no outward clinical signs of impaired fasting glucose or hyperinsulinemia. Patients move from a prediabetic state to a diabetic state as pancreatic beta cells gradually become exhausted and are no longer able to meet the demands of insulin-resistant tissues. Impaired pancreatic beta-cell function is marked by the reduced capacity of beta cells to mount a Þrst-phase insulin secretory response following ingestion of meals. As the patient’s disease worsens, pancreatic beta-cell function further deteriorates with the loss of the prolonged second-phase insulin secretory response. Loss of insulin activity results in chronic hyperglycemia stemming from the reduced ability to lower postprandial glucose and the dysregulation of hepatic glucose production. However, as is the case for prediabetes, type 2 diabetes and chronic hyperglycemia can be clinically silent, producing only vague,

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TABLE 5. Nongenetic Causes of Type 2 Diabetes Disease/Condition Pancreatic diseases

Endocrine dysfunction

Iatrogenic (drug- or chemical-induced)

Comments Full or partial pancreatectomy for treatment of chronic pancreatitis, pancreatic cancer, or insulinomas results in loss of beta cells and insulin secretory activity. The exocrine pancreas of patients with cystic fibrosis becomes damaged over time as a result of tissue degradation by pancreatic enzymes in blocked ducts, thereby reducing insulin secretory activity. Other rare diseases such as hemochromatosis and fibrocalculous pancreatopathy (tropical pancreatic diabetes) can damage the pancreas and inhibit insulin production and secretion. Chronically elevated levels of cortisol in patients with Cushing’s syndrome cause obesity, hypertension, and diabetes. Hormonal imbalances resulting from acromegaly (excess growth hormone) and hyperthyroidism (excess thyroid hormone) trigger insulin resistance and increase hepatic glucose production, respectively. Glandular carcinomas (e.g., glucagonomas, pheochromocytomas, somatostatinomas, aldosteromas) can produce high levels of hormones that disrupt insulin secretion and signaling. Many commonly prescribed pharmaceutical agents cause insulin resistance in susceptible individuals via various mechanisms. Hormonally active drugs (e.g., glucocorticoids, thyroid hormone, oral contraceptives) cause endocrine dysfunction. Antihypertensive agents (e.g., thiazide diuretics, beta-adrenergic agonists, diaxozide) have been linked to increased risk of type 2 diabetes in some epidemiological studies. Anti-infective agents (e.g., alpha-interferon) can trigger autoimmune reactions that damage pancreatic beta cells.

Source: Adapted from American Diabetes Association. Diagnosis and classification of diabetes mellitus. Diabetes Care. 2005;28:S37-S42.

nonspeciÞc symptoms such as polyuria, polyphagia, and/or polydipsia (increased urination, hunger, and thirst, respectively) and/or fatigue. Glucose toxicity from chronic hyperglycemia is responsible for much of the morbidity and mortality associated with type 2 diabetes, which includes such microvascular complications as diabetic neuropathy, retinopathy, and nephropathy as well as macrovascular complications (i.e., CVD). Indeed, type 2 diabetes is one of the most frequent causes of leg amputation, blindness, end-stage renal disease, heart attack, and stroke—all sources of tremendous disability and cost to health care systems. In 2002, an estimated 10% of total health care expenditure in the United States was attributed to direct and indirect costs of diabetes and diabetic complications (American Diabetes Association, www.diabetes.org/diabetes-statistics/costof-diabetes-in-us.jsp, accessed June 17, 2005). Therefore, tremendous need exists

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FIGURE 1. Clinical progression of type 2 diabetes.

for disease-modifying agents that can slow or halt the progression of type 2 diabetes, thereby preventing the development of diabetic complications. Insulin Secretion. In nondiabetic patients, serum glucose levels are maintained in a narrow physiological range via the actions of insulin and glucagon, two key hormones that regulate serum glucose levels. Insulin and glucagon are produced by pancreatic beta and alpha cells, respectively. These hormones have opposing effects that serve to maintain serum glucose levels during fasting and postprandial states. The beta and alpha cells reside in highly vascularized glandular tissue clusters in the pancreas known as the islets of Langerhans. The islet cells sense blood glucose levels and release glucagon or insulin as needed when hypoglycemia (low blood sugar) or hyperglycemia (high blood sugar) occurs, respectively. Glucagon. Glucagon is synthesized and released by pancreatic alpha cells during periods of fasting; it raises blood glucose concentrations to normal physiological levels by stimulating gluconeogenesis (glucose production) and glycogenolysis (breakdown of glycogen) in the liver. Both hepatic gluconeogenesis and glycogenolysis raise serum glucose levels during sleep and between meals, when blood glucose levels decline. The most widely used oral antidiabetic agent, metformin, works in part by inhibiting hepatic glucose production (see “Current Therapies” for more information). Patients with type 2 diabetes often have high circulating plasma glucagon levels stemming from resistance of pancreatic alpha cells to the effects of insulin, which suppresses glucagon secretion. The resultant dysfunction in the regulation of glucagon secretion causes elevated fasting glucose levels in insulin-resistant individuals. To address this mechanism of hyperglycemia, several companies have agents in preclinical development that function by inhibiting glucagon receptors. Excess plasma glucagon levels can also be caused by rare pancreatic alpha cell tumors (glucagonomas) that secrete large quantities of glucagon. Insulin. Pancreatic beta cells constantly secrete small quantities of insulin to maintain basal insulin levels. Following meals, glucose transporters on the beta cells sense the rise in plasma glucose levels and signal the release of pre-formed

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FIGURE 2. Glucose-dependent first-phase and second-phase insulin secretion from pancreatic beta cells.

insulin molecules stored in secretory granules (Figure 2). This initial release is known as Þrst-phase insulin secretion and results in a sharp peak in serum insulin levels following food intake. Second-phase insulin secretion occurs 20–30 minutes later as the glucose transporters induce synthesis of more insulin and stimulate release of the hormone over a prolonged period. Defects in any of the biochemical pathways responsible for regulating insulin production and secretion can lead to the development of type 2 diabetes. One class of currently marketed oral antidiabetic agents, the sulfonylureas, function as insulin secretagogues and help lower blood glucose levels by stimulating insulin production by pancreatic beta cells (see “Current Therapies” for more information about sulfonylureas).

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FIGURE 3. Regulation of insulin.

In addition to elevations in serum glucose levels, the passage of nutrients in the gastrointestinal (GI) tract stimulates insulin release (Figure 3). This mechanism of stimulating insulin secretion is known as the incretin effect because it is mediated by incretins (gut hormones) released by the intestines during digestion. The incretin effect is responsible for stimulating as much as 20–60% of postprandial insulin secretion, depending on the amounts and type of nutrients passing through the GI tract (Meier JJ, 2005). In addition to stimulating insulin secretion, the incretin effect lowers blood glucose levels by inhibiting the secretion of glucagon by pancreatic alpha cells. The two best-characterized incretins are glucagon-like peptide 1 (GLP-1) and glucose-dependent insulinotropic peptide (GIP, also called gastric inhibitory peptide). Table 6 lists the physiological functions of GLP-1 and GIP. Several emerging antidiabetic agents function by stimulating the incretin effect, either by activating GLP-1 receptors or by inhibiting dipeptidyl peptidase IV (DPP-IV), the enzyme responsible for converting

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TABLE 6. The Physiological Functions of the Incretins GLP-1 and GIP Incretin

Physiological Function

Glucagon-like peptide 1 (GLP-1)

Slows gastric emptying and reduces appetite. Suppresses glucagon secretion. Stimulates insulin secretion in a glucose-dependent manner.

Glucose-dependent insulinotropic polypeptide (GIP)

Stimulates insulin secretion in a glucose-dependent manner. Inhibits gastric acid production. Stimulates glucagon secretion under certain circumstances (Meier JJ, 2003).

Comments Produced by L cells in the ileum and colon. Derived from the same precursor molecule as glucagon. Increases beta-cell mass in animal models and in vitro studies. Also called gastric inhibitory polypeptide. Produced by K cells in the duodenum and jejunum. Insulinotropic effect of GIP is almost absent in type 2 diabetics.

Source: Adapted from Meier JJ, Nauck MA. Glucagon-like peptide 1 (GLP-1) in biology and pathology. Diabetes/Metabolism Research and Reviews. 2005;21:91–117.

active GIP and GLP-1 into their inactive forms. There is also evidence that GLP-1 has a trophic effect on beta cells, resulting in proliferation and maturation of existing and new beta cells, respectively (Holst JJ, 2002). (See “Current Therapies” and “Emerging Therapies” for more information about these novel therapeutic approaches.) Impaired Glucose Transport. Approximately 75% of serum glucose uptake occurs in skeletal muscle tissue. The remainder is metabolized by the liver, adipocytes, and other tissues. Individuals with insulin resistance are less able to absorb glucose into skeletal muscle and adipocytes, a result in part of defects in glucose transport mechanisms. Figure 4 depicts the basic signal transduction pathways that activate glucose transport in response to insulin signaling; disruption of any step in the pathway can result in insulin resistance. Table 7 lists some of the key genes associated with defective insulin signaling and glucose transport. Glucose Transporters. Glucose transporters (GLUT) play a crucial role in maintaining glycemic control by performing diverse functions such as glucose sensing and transport. Thirteen members of the GLUT family of proteins have been identiÞed so far (Wood IS, 2003). Different combinations of GLUT proteins are expressed on tissues such as muscle, liver, and adipocytes. Activation of GLUT proteins increases uptake of glucose by these tissues, thereby lowering circulating blood glucose levels and maintaining glycemic control. Several GLUT family members are activated by peroxisome proliferator-activated receptor (PPAR)-gamma, which is an important target of several currently marketed oral antidiabetic agents (Kim HI, 2004; see “Current Therapies” for more information).

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TABLE 7. Key Genes Involved in Impaired Glucose Transport and Insulin Signaling Gene INS (insulin)

IRS-1 (insulin receptor substrate-1)

Calpain 10/2 (within the NIDDM1 locus)

IR (insulin receptor)

PTP-1B (protein tyrosine phosphatase-1B)

Comments Genetic association study in 155 British parent-offspring demonstrated that the variable number tandem repeat (VNTR) allele of the insulin gene may influence susceptibility to type 2 diabetes (Huxtable SJ, 2000). IRS-1 is a signaling molecule involved in insulin signaling. Codon 972 of the IRS-1 gene has been implicated in type 2 diabetes susceptibility (Almind K, 1993). In another study, approximately 11% of the diabetic subjects, who were from South India and Finland, exhibited this mutation (Hitman GA, 1995). The calpains (calcium-activated neutral proteases) are an omnipresent family of nonlysosomal cysteine proteases that are implicated in various cellular functions, such as intracellular signaling, proliferation, adipocyte differentiation, and insulin-induced downregulation of IRS-1. Although the pathogenic role of the calpains in type 2 diabetes is not yet fully known (Horikawa Y, 2000), researchers believe that the finding that variation in the CAPN10 gene influences FFA levels and insulin resistance (which are known to predict type 2 diabetes) may explain how this gene increases susceptibility to type 2 diabetes (Orho-Melander M, 2002). Mutations of the insulin receptor have been identified in several studies involving patients with type 2 diabetes. The prevalence of these mutations is estimated to range from 0.4% to 7.8% in the type 2 diabetic patient population (Sesti G, 2001). PTP-1B is involved in insulin signal transduction. A P387L variant of this gene is associated with type 2 diabetes in Danish Caucasian diabetic patients (Echwald SM, 2002).

Note: Full source citations appear in ‘‘References.’’

Insulin Receptor-Mediated Signal Transduction. The role of protein kinase signaling mechanisms in the etiology of insulin resistance and type 2 diabetes remains poorly understood. Activation of insulin receptors by insulin or insulin-like growth factor (IGF) results in the phosphorylation of insulin-receptor substrate (IRS-1), which serves as a link between the insulin receptor and downstream signal transduction pathways (Figure 4). Researchers note that some type 2 diabetic patients with chronic myelogenous leukemia who were treated with imatinib mesylate (Novartis’s Gleevec), a tyrosine kinase inhibitor with high speciÞcity for the BCR/ABL tyrosine kinase, exhibited marked improvements in glucose control (Brecchia M, 2004; Veneri D, 2005). Their observations suggest that the pathogenesis of type 2 diabetes may involve the aberrant activation of protein kinase signaling mechanisms. However, the speciÞc signal transduction pathways inhibited by imatinib mesylate to improve glycemic control remain to be identiÞed. Several emerging therapies that modulate signal transduction

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FIGURE 4. Insulin-dependent glucose transporter activation and expression in skeletal muscle or adipocytes.

pathways (e.g., protein tyrosine phosphatase-1B [PTP-1B] inhibitors) are in earlystage development for type 2 diabetes (for more information, see “Emerging Therapies”). Microvascular Complications of Type 2 Diabetes. Chronically elevated blood glucose levels (hyperglycemia) are the cause of several microvascular complications of type 2 diabetes. Diabetic retinopathy, nephropathy, and neuropathy

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can lead to blindness, end-stage renal disease (ESRD), and amputation, respectively, if glycemic control is not maintained over the course of a diabetic patient’s life. The pathophysiology of these microvascular complications stems from the accumulation and deposition of advanced glycated end products (AGEs) in tissues, which occur as a biochemical consequence of chronically elevated glucose levels in the circulation reacting irreversibly with biological macromolecules such as proteins in the endothelial lining of the vasculature or nerve tissue. AGEs thus exert their effect by interfering with blood ßow, disrupting neurotransmission, and inhibiting wound healing. One commonly glycated protein in the circulation, hemoglobin A1c (HbA1c ), has been exploited as a diagnostic measure of long-term (two to three months) glycemic control in diabetic patients. Retinopathy. Diabetic retinopathy is best described as capillary microangiopathy that involves the loss of pericytes and endothelial cells with changes in basement membrane thickness. This condition results in microinfarctions, exudates, retinal edema (ßuid retention), and neovascularization—all of which can progress to vitreous hemorrhage, traction, and retinal detachment, causing severe loss of vision. Diabetes is one of the leading nongenetic causes of blindness in adult populations. Nephropathy. Diabetic nephropathy is a multistage condition that often becomes clinically overt several years after diabetes onset. The Þve stages are as follows: • • • • •

Glomerular hyperÞltration. Microalbuminuria/subclinical nephropathy (albumin excretion rate of 20–200 µg/minute). Clinical/overt nephropathy (albumin excretion rate greater than 200 µg/minute, accompanied by hypertension). Advanced nephropathy (glomerular Þltration rate less than 75 mL/minute and symptoms of uremia). End-stage renal disease (ESRD) requiring dialysis or renal transplantation.

Diabetes is the leading cause of renal dialysis and ESRD worldwide and accounts for a signiÞcant amount of cost to health care systems. Neuropathy. Diabetic neuropathy is one of the most common long-term complications of diabetes. The three major categories of diabetic neuropathy are as follows: • •

•

Peripheral and proximal neuropathy causes tingling, numbness, and often intense pain in the extremities (e.g., hands, hips, legs, feet). Focal neuropathy affects a single nerve or bundle of nerve Þbers and can cause sudden muscle weakness, pain, or visual defects if an optical nerve is affected. Autonomic neuropathy causes changes in many basic bodily functions such as digestion, bladder control, control of blood pressure, and sexual function.

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The numbness caused by peripheral neuropathy has several serious consequences. Patients with peripheral neuropathy commonly develop foot ulcers as a result of small-Þber sensory dysfunction. Because these chronic ulcers heal slowly and are often untreated, they are prone to infection, which can lead to gangrene and ultimately to amputation. Indeed, diabetic neuropathy is the leading cause of nontraumatic amputations in the United States. Macrovascular Complications of Type 2 Diabetes. Type 2 diabetic patients are prone to lipid abnormalities and hypertension, conditions that greatly increase the risk of mortality from cardiovascular complications such as heart attack and stroke. Type 2 diabetes is considered to be a cardiovascular risk equivalent in the Framingham global risk scoring system, which considers a type 2 diabetic to have the equivalent risk for heart attack as an individual who has already had a heart attack (Haffner SM, 1998). Indeed, a prospective study of nearly 350,000 men in the Multiple Risk Factor Intervention Trial (MRFIT) indicated that type 2 diabetes is a stronger predictor of CVD risk than classic CVD risk factors such as hypertension, high total serum cholesterol, and history of smoking tobacco (Stamler J, 1993). Although the medical community continues to debate the relative contributions of hyperglycemia and hyperinsulinemia in the development of CVD, it has become widely accepted that even prediabetic patients who have borderline laboratory values for some of the metabolic complications that accompany diabetes (dyslipidemia and hypertension) are at increased risk for cardiovascular disease (Eckel RH, 2005; Kendall DM, 2002; Reusch JE, 2002; Zimmet PZ, 1999, Haffner SM, 1992; Isomaa B, 2001; Stolar MW, 2002). This cluster of comorbidities is known as metabolic syndrome, a disorder comprising abdominal obesity, insulin resistance, hypertension, elevated triglycerides, and low HDL-cholesterol. Table 8 lists the most widely accepted deÞnitions of metabolic syndrome; they were published by the Third Adult Treatment Panel of the National Cholesterol Education Program and the World Health Organization to identify patients at risk for CVD who do not have classic risk factors. The elevated cardiovascular risk stems in part from insulin resistance, which is associated with elevated plasma levels of C-reactive protein (CRP), Þbrinogen, and plasminogen activator inhibitor-1 (PAI-1) (Festa A, 2002[a]; Festa A, 2002[b]; Folsom AR, 1997). Although not formally recognized as components of metabolic syndrome, these factors increase the risk of CVD by promoting inßammation and thrombosis. CURRENT THERAPIES The discovery of insulin in 1921 by Frederick G. Banting and Charles Best was a major breakthrough in the treatment of diabetes. Innovations in the puriÞcation process and recombinant DNA technology have transformed insulin therapy over the past several decades—from bovine/porcine-derived insulin (1920s) to recombinant human insulin (late 1970s) to human insulin analogues (1990s), which best mimic the physiological effects of endogenous insulin.

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TABLE 8. NCEP-ATP III and WHO Definitions of Metabolic Syndrome According to the National Cholesterol Education Program (NCEP) Adult Treatment Panel III (ATP III), a patient with three or more of the following traits is defined as having metabolic syndrome (Expert Panel on Detection, Evaluation, and Treatment of High Blood Cholesterol in Adults, 2001): • Abdominal obesity: waist circumference >102 cm in men and >88 cm in women. • Hypertriglyceridemia: ≥150 mg/dL (1.7 mmol/L). • Low high-density lipoprotein (HDL) cholesterol: <40 mg/dL (1.04 mmol/L) in men and <50 mg/dL (1.29 mmol/L) in women. • Hypertension, defined as elevated blood pressure (>130 mm Hg systolic or >85 mm Hg diastolic). • Elevated fasting plasma glucosea >110 mg/dL (≥6.1 mmol/L). According to the World Health Organization (WHO), a patient has metabolic syndrome if he or she has type 2 diabetes, impaired fasting glucose (IFG), impaired glucose tolerance (IGT), or impaired glucose uptake (measured by the hyperinsulinemic euglycemic clamp technique) as well as two of the criteria described below (Alberti KG, 1998): • Hypertension, defined as antihypertensive treatment and/or elevated blood pressure (>140 mm Hg systolic or >90 mm Hg diastolic). • Dyslipidemia, defined as elevated plasma triglyceride (≥150 mg/dL, ≥1.7 mmol/L) and/or low HDL cholesterol (<35 mg/dL, <0.9 mmol/L in men; <39 mg/dL, <1.0 mmol/L in women). • Obesity, defined as a high BMI (≥30 kg/m2 ) and/or a high waist:hip ratio (>0.90 in men and >0.85 in women). • Microalbuminuria, defined by urinary albumin excretion rate ≥20 µg/min or albumin:creatine ratio ≥30 mg/g. a In 2004, the American Diabetes Association reduced the cutoff for diagnosis of hyperglycemia to 100

mg/dL. The National Heart, Lung, and Blood Institute and the American Heart Association have not yet revised the NCEP-ATP III guidelines to reflect this change but may do so in the near future (Grundy SM, 2004). BMI = Body mass index. Note: Full source citations appear in ‘‘References.’’

Advances have been made not only in insulin therapies but also in the development of oral antidiabetic agents (OAAs) (Figure 5). ScientiÞc investigators have uncovered new agents that target pathophysiological pathways that contribute to the development of type 2 diabetes. In the 1950s, a novel class of oral insulin secretagogues—the sulfonylureas—was introduced, reßecting the predominant paradigm that type 2 diabetes was a defect in insulin secretion. The biguanide metformin (available Þrst in Europe) and improved sulfonylureas were developed and launched shortly thereafter. The next leap in innovation occurred in the 1990s when several OAAs with various mechanisms of action were approved for type 2 diabetes: metformin (in the United States and Japan), alpha-glucosidase inhibitors (AGIs), meglitinides, and peroxisome proliferator-activated receptorgamma (PPAR-gamma) agonists. Greater knowledge of the pathophysiology of type 2 diabetes, particularly the role of insulin resistance, guided the development

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FIGURE 5. Treatment of type 2 diabetes: a historical perspective.

of these agents. The most recent group of oral agents to enter this market is the Þxed-dose combination (FDC) agents—the Þrst of which, metformin/glyburide (Bristol-Myers Squibb’s Glucovance), was introduced in 2000. In 2005, scientiÞc Þndings that demonstrate that type 2 diabetes is a multihormonal disorder culminated in the launch of two new drugs—the amylin analogue pramlintide (Amylin Pharmaceuticals’ Symlin) and the glucagon-like peptide-1 (GLP-1) analogue exenatide (Amylin Pharmaceuticals/Eli Lilly’s Byetta). As the scientiÞc and medical communities continue to elucidate the role of insulin resistance in cardiovascular risk and the concept of metabolic syndrome becomes more well-deÞned, greater attention is being focused on the effects of current antidiabetic agents on end points other than glucose levels, such as lipid proÞles and inßammatory markers. Many primary and secondary prevention trials

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with existing therapies are under way. These trials have the potential to alter diabetes management as did the United Kingdom Prospective Diabetes Study (UKPDS) and the Diabetes Prevention Program (DPP). This section summarizes select placebo-controlled and comparator clinical trial data that demonstrate the efÞcacy and side effects of key agents used to treat type 2 diabetes. Although many trials have measured several key end points (such as parameters for cardiovascular risks), this section will focus on clinical trials that demonstrate glucose-lowering efÞcacy. Table 9 lists the leading therapies used to treat type 2 diabetes, Table 10 summarizes the advantages and disadvantages of each therapeutic class, and Table 11 summarizes efÞcacy data for oral monotherapies. Figure 6 depicts their sites of action. Peroxisome Proliferator-Activated Receptor-Gamma Agonists Overview. Developed initially in Japan, PPAR-gamma agonists are insulinsensitizing agents that enhance the effect of insulin at peripheral target sites. These agents also have the potential to modify disease progression by preserving pancreatic beta-cell function. PPAR-gamma agonists are sometimes referred to as thiazolidinediones (TZDs), although several nonthiazolidinedione agents are in development in both the PPAR-gamma agonist and dual-acting PPAR agonist classes. Rosiglitazone (GlaxoSmithKline’s Avandia) and pioglitazone (Takeda/Eli Lilly’s Actos) are the only two PPAR-gamma agonists currently available. The Þrst PPAR-gamma agonist, troglitazone (Warner-Lambert’s [now part of PÞzer] Rezulin, GlaxoSmithKline’s Romozin, Sankyo’s Noscal), was withdrawn from Europe in 1998 and from the United States and Japan in 2000 because of cases of fatal liver toxicity. No reports of lethal hepatotoxicity among patients treated with rosiglitazone or pioglitazone have been reported. However, PPARgamma agonists should not be used in patients with active hepatocellular disease or in patients with elevated serum alanine aminotransferase (ALT). Serum ALT monitoring is recommended before therapy is initiated and intermittently thereafter. The PPAR-gamma agonists are associated with weight gain, edema (ßuid retention), and anemia, as their labeling indicates. In a meta-analysis of 22 doubleblind comparator and open-label studies of rosiglitazone, representing a ten-year span of clinical trial experience and more than 5,000 patients, there was no excess reporting of liver enzyme elevations (Krebs J, 2005). Although hepatotoxicity is becoming less of a concern, the possible health consequences of edema (ßuid retention) caused by PPAR-gamma agonists are a topic of debate (Figure 7). In 2002, after a few cases of congestive heart failure were reported, the drug labels for rosiglitazone and pioglitazone were updated to include warnings of cardiac failure and other cardiac events. Head-to-Head Trials of Rosiglitazone and Pioglitazone. Rosiglitazone and pioglitazone are associated with gradual improvements in glycemic control over weeks to months, with concurrent improvement in insulin sensitivity. To date, there have been only a few head-to-head studies of rosiglitazone and

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TABLE 9. Current Therapies Used for Type 2 Diabetes Agent

Company/Brand

PPAR-gamma agonists Rosiglitazone GlaxoSmithKline’s Avandia Pioglitazone Takeda/Eli Lilly’s Actos Biguanides Metformin Bristol-Myers Squibb’s Glucophage, generics Bristol-Myers Squibb’s Glucophage XR, Biovail/DepoMed’s Glumetza Sulfonylureas Glimepiride Sanofi-Aventis’ Amaryl Glyburide/ Sanofi-Aventis’ Diabeta, glibenclaPfizer’s mide Micronase/Glynase, generics Glipizide Pfizer’s Glucotrol, enerics Pfizer’s Glucotrol XL Gliclazide Servier’s Diamicron, Molteni & C.F. LLI Alitti’s Diabrezide, Dainippon’s Glimicron, generics Fixed-dose combinations Metformin/ GlaxoSmithKline’s rosiglitaAvandamet zone Metformin/ Bristol-Myers Squibb’s glyburide Glucovance, Sanofi-Aventis’s Suguan M, Fournier Pharma’s GlicoRest, Abiogen’s Glicnorm, Roche’s BiEuglucon, generics Metformin/ Bristol-Myers Squibb’s glipizide Metaglip Meglitinides Repaglinide

Novo Nordisk’s Prandin/NovoNorm Nateglinide Novartis/ Yamanouchi/ Sanofi-Aventis Japan’s Starlix/Starsis/Fastic Mitiglinide Kissei/Takeda’s Glufast Alpha-glucosidase inhibitors Acarbose Bayer’s Precose/Glucobay, generics Miglitol Bayer/Pfizer/SanofiAventis’s Glyset Voglibose Takeda’s Basen

Daily Dose

Availability

4–8 qd or 2–4 mg bid 15–45 mg qd

US, F, G, I, S, UK US, F, G, I, S, UK, J

500 tid, 850 mg bid

US, F, G, I, S, UK, J

500–1,500 mg qd

US

1–4 mg qd 2.5–5 mg qd or bid

US, F, G, I, S, UK, J US, F, G, I, S, UK, J

10 mg qd, or bid 5–10 mg qd 30–80 mg tid

US, F, G, I, S, UK, J F, G, I, S, UK, J

500 mg/1 mg, 500 mg/ US, F, G, S, UK 2 mg, or 500 mg/4 mg bid 500 mg/2.5 mg bid

US, I

250 mg/2.5 mg, 500 mg/2.5 mg, or 500 mg/5 mg qd or bid

US

1.5–2 mg tid

US, F, G, I, S, UK

60–120 mg tid

US, G, UK, J

5–10 mg tid

J

50–100 mg tid

US, F, G, I, S, UK, J

25–50 mg tid

US, F, G, S

0.2–0.3 mg tid

J

CURRENT THERAPIES

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TABLE 9. (continued) Agent

Company/Brand

Insulin and insulin analogues Insulin lispro Eli Lilly’s Humalog Insulin Sanofi-Aventis’s Lantus glargine GLP-1 analogues Exenatide Amylin Pharmaceuticals/Eli Lilly’s Byetta Amylin analogues Pramlintide Amylin Pharmaceuticals’ Symlin

Daily Dose

Availability

50–60 U, sc injection 2–100 U, sc injection

US, F, G, I, S, UK, J US, G, UK, J

5–10 mg bid, sc injection

US

90–120 mg bid or tid, sc injection

US

bid = Twice daily; qd = Once daily; sc = Subcutaneous; tid = Three times daily. US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan. GLP-1 = Glucagon-like peptide-1; PPAR = Peroxisome proliferator-activated receptor.

pioglitazone. Clinical studies demonstrate that monotherapy with rosiglitazone or pioglitazone offers comparable improvements in glycemic control, although pioglitazone improves triglyceride (TG) and high-density lipoprotein cholesterol (HDL-C) levels as well (Boyle PJ, 2002; Khan MA, 2002; Olansky L, 2003). These Þndings were conÞrmed in a head-to-head study involving 800 type 2 diabetic patients who were randomized to receive either pioglitazone (30 mg/day) or rosiglitazone (4 mg/day) for 24 weeks (Goldberg RB, 2005). Patients in the pioglitazone and rosiglitazone treatment arms experienced similar reductions in HbA1c levels (−0.7% and −0.6%, respectively). Patients receiving pioglitazone also experienced, on average, a 12% decline in TGs and a 15% increase in HDL, whereas those receiving rosiglitazone experienced a 15% increase in TG and an 8% increase in HDL. Patients in both treatment arms experienced elevations in LDL at the end of the study; patients treated with pioglitazone saw a 16% increase in LDL, while those treated with rosiglitazone saw a 23% increase in LDL. Mechanism of Action The PPAR-gamma receptor is found in the nucleus of key target tissues for insulin action, such as skeletal muscle, adipose tissue, and the liver. PPAR-gamma agonists improve glycemic control by enhancing insulin sensitivity, as illustrated in Figure 8. In the presence of endogenous or exogenous insulin, PPAR-gamma agonists increase glucose uptake and utilization in skeletal muscle, increase glucose uptake and reduce fatty-acid output in adipose tissue, and reduce gluconeogenesis, glucose output, and triglyceride synthesis in the liver. Researchers also believe that PPAR-gamma agonists downregulate resistin, which is expressed in diabetic adipocytes and inhibits insulin action in skeletal muscle. In vitro studies have shown that PPAR-gamma agonists trigger an increase in GLUT4 (glucose transporter) cell-surface expression (Hauner H, 2002). In addition, studies have demonstrated anti-inßammatory and antiapoptotic effects that may limit the progression to cardiovascular complications (Artwohl M, 2005; Buffon A, 2005).

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TABLE 10. Efficacy, Benefits, and Drawbacks of Drug Classes Used to Treat Type 2 Diabetes

Drug Class Oral antidiabetic agents Peroxisome proliferator-activated receptor-gamma (PPAR-gamma) agonists Biguanides

Efficacya

Targeted Pathophysiological Defect

Competitive Advantages

1.0–1.6%b

Insulin resistance

Rosiglitazone may have certain short-term benefits on pancreatic beta-cell function.

1.0–2.0%b

Hepatic glucose overproduction Insulin resistance

Sulfonylureas

0.9–2.5%b

Loss of first-phase insulin secretion

Reduce HbA1c levels more rapidly than other marketed antidiabetic agents. Unlike sulfonylureas and meglitinides, biguanides do not cause hypoglycemia or weight gain. Generally well-tolerated other than transient mild side effects (e.g., nausea, vomiting, headache).

Fixed-dose combinations

N.A.

N.A.

The convenience of administering a single pill may increase patient compliance. Co-administration of agents can result in a greater reduction in blood glucose levels, compared with administering either agent alone.

Drawbacks Associated with weight gain, fluid retention, and anemia. Cardiac failure and other cardiac events occur rarely. Gastrointestinal irritation (i.e., nausea and diarrhea) occurs in about one-third of patients during the first few weeks of therapy. Risk of lactic acidosis in some patients. Associated with weight gain (typically five to ten pounds in clinical trials). Hypoglycemia, which occurs most frequently with the long-acting sulfonylureas (i.e., glyburide and chlorpropamide). Efficacy depends upon presence of functioning pancreatic beta cells. Fixed dosing is limiting—some physicians prefer the flexibility of prescribing agents separately.

TABLE 10. (continued)

Efficacya

Targeted Pathophysiological Defect

Competitive Advantages

Drawbacks

Postprandial hyperglycemia

Pose a lower risk for hypoglycemia than sulfonylureas—an advantage in the elderly population. Exert action only in the presence of elevated glucose levels.

0.4–1.0%b

Postprandial hyperglycemia

Do not cause hypoglycemia or weight gain, unlike sulfonylureas and meglitinides.

Generally considered to have a modest therapeutic effect compared with that of other oral antidiabetics. Most common adverse reactions are upper respiratory tract infection and headache. Generally considered to exert a modest therapeutic effect compared with that of other oral antidiabetics. Unpleasant side effects, namely flatulence.

Injectable agents Insulin lispro

>2%c

Complete loss of beta-cell function

Insulin glargine

>2%c

Complete loss of beta-cell function

Results in fewer hypoglycemic events than therapy with regular insulin. Has a shorter onset of action (15 minutes), duration of action (three to four hours), and time of absorption compared with regular insulin. Achieves better glycemic control than regular insulin with fewer hypoglycemic events. Provides a longer, peakless duration of action that more effectively meets basal insulin requirements than other intermediate- and long-acting insulins.

Drug Class Meglitinides

0.6–1.9%b

Alpha-glucosidase inhibitors

Inconvenient mode of administration (injection). May cause hypoglycemia. Inconvenient mode of administration (injection). May cause hypoglycemia.

355

356

TABLE 10. (continued)

Drug Class

Efficacya

GLP-1 analogues

Can be injected without initial resuspension (which is a major cause of variability in other insulins). 0.4–1.5%c

Amylin analogues

0.5%d

Targeted Pathophysiological Defect

Hepatic glucose overproduction, loss of insulin secretion, and insulin resistance Amylin deficiency

Competitive Advantages

Drawbacks

Targets multiple physiological defects. Does not cause weight gain. Potential for beta-cell preservation. Reduces insulin dose needed to sustain glycemic control. Reduces side effects from insulin (i.e., weight gain, hypoglycemia).

Inconvenient mode of administration (injection). GI disturbances, such as nausea.

Inconvenient mode of administration (injection). Must be taken with insulin.

a These figures do not represent comparative efficacy because no trial has investigated the effects of all antidiabetic agents in relation with one another. Because of varying

recruitment criteria, study length, and trial designs, a direct comparison of one agent with another is not feasible. b Efficacy is based on placebo-controlled, short-term monotherapy trials and presented as a placebo-adjusted reduction in HbA from baseline. 1c c Efficacy is based on placebo-controlled, add-on trials in which patients maintained their prestudy treatment regimens. The data are presented as the additional reduction in

HbA1c observed when the new therapy is added to existing therapy. d Efficacy is based on add-on trials in which patients maintained their insulin treatment regimens. The data are presented as the additional reduction in HbA achieved when 1c the amylin analogue was added to insulin therapy.

N.A. = Not applicable.

TABLE 11. Select Monotherapy Trials of Oral Therapies Used to Treat Type 2 Diabetes Oral Antidiabetic Agent

Study Design

PPAR-gamma agonists Rosiglitazone Randomized, double-blind, placebo-controlled (Lebovitz HE, 2001)

Pioglitazone

Randomized, double-blind, placebo-controlled (Phillips LS, 2001) Randomized, double-blind, placebo-controlled (Aronoff S, 2000) Prospective, observational cohort in Japanese patients (Kawamori R, 2004)

Number of Patients

Study Length

Placebo-Adjusted HbA1c Reductiona

493

26 weeks

1.2–1.5%

959

26 weeks

1.1–1.5%

408

26 weeks

1.0–1.6%

22,943

18 months

1% (absolute reduction)

Comments Low-density lipoprotein (LDL) increased threefold and plateaued after two months, but this effect was offset by improvements in high-density lipoprotein (HDL).

Triglyceride (TG) and HDL levels improved in a statistically significant manner. 35% of all patients achieved HbA1c <7%; glucose levels remained stable up to 18 months. 50% of drug-naive patients achieved HbA1c <7%. No cases of hepatic failure were reported. Edema was reported in 8% of patients.

357

358

TABLE 11. (continued) Oral Antidiabetic Agent Biguanides Metformin

Sulfonylureas Glimepiride

Glyburide

Glipizide

Study Design Randomized, double-blind, placebo-controlled (Jones KL, 2002) Randomized, double-blind, placebo-controlled (DeFronzo RA, 1995)

Number of Patients

Study Length

Placebo-Adjusted HbA1c Reductiona

Comments

82 young patients (10–16 years)

16 weeks

1.1%

Metformin is currently the only oral antidiabetic that is indicated for use in children.

289

29 weeks

1.5%

Statistically significant reductions in total cholesterol, LDL, and TGs were observed.

Randomized, double-blind, placebo-controlled (Rosenstock J, 1996) Randomized, double-blind, placebo-controlled (Schade DS, 1998) Randomized, double-blind comparator with glimepiride (Draeger KE, 1996)

416

14 weeks

2.5%

249

22 weeks

1.4%

1,044

52 weeks

Equivalent efficacy

Prospective, randomized, double-blind, comparator with glyburide (Birkeland KI, 1994)

46

15 months

Equivalent efficacy

69% of treated patients achieved HbA1c ≤ 7; 32% of placebo-treated patients achieved HbA1c ≤ 7. Glimepiride patients did, however, achieve significantly lower fasting insulin and C-peptide levels compared with those of the glyburide-treated group.

TABLE 11. (continued) Oral Antidiabetic Agent

Study Design

Gliclazide

Randomized, double-blind comparator trial with glyburide (Tessier D, 1994) Fixed-dose combinations Metformin/break Single-arm, open-label rosiglitazone (Rosenstock J, 2005)

Metformin/break glyburide

Randomized, placebo-controlled (Garber A, 2000) Open-label extension study (Blonde L, 2004)

Metformin/break glipizide

Meglitinides Repaglinide

Randomized, double-blind comparator (Rubin CJ, 2002[a])

359

Randomized, double-blind, placebo-controlled (Jovanovic L, 2000)

Number of Patients

Study Length

Placebo-Adjusted HbA1c Reductiona

Comments

22 untreated elderly patients

24 weeks

Equivalent efficacy

The incidence of hypoglycemia was significantly greater with glyburide.

190 drug-naive patients

24 weeks

4% (absolute reduction)

50% of patients achieved HbA1c < 7%; 38% of patients achieved HbA1c < 6.5%.

806 drug-naive elderly (age ≥60 years) patients 477 patients uncontrolled on sulfonylureas 868 drug-naVve patients

20 weeks

1.3%

52 weeks

1.7% (absolute reduction)

24 weeks

Additional reduction of 0.4–0.7% in HbA1c , compared with metformin or glipizide alone

24 weeks

1.9%

361

60% of patients treated with the fixed-dose combination achieved HbA1c <7%, compared with 40% in the metformin and glipizide groups.

360 TABLE 11. (continued) Oral Antidiabetic Agent Nateglinide

Study Design

Randomized, double-blind, placebo-controlled (Horton ES, 2000) Alpha-glucosidase inhibitors Acarbose Randomized, double-blind, placebo-controlled (Fischer S, 1998) Miglitol Randomized, double-blind, placebo-controlled (Johnston PS, 1998) Voglibose Randomized, crossover, open, comparator with acarbose

Number of Patients

Study Length

Placebo-Adjusted HbA1c Reductiona

701

24 weeks

1.0%

495

24 weeks

1.0%

345

52 weeks

0.4–0.5%

30

8 weeks

Voglibose had slightly less efficacy in reducing post-prandial glucose levels.

Comments

HbA1c was not measured

a The efficacy represents placebo-adjusted HbA 1c reductions from baseline where appropriate. Because of varying recruitment criteria and trial designs used to compile the data in this table, a direct comparison of one agent with another is not feasible. Note: Full source citations appear in ‘‘References.’’

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Rosiglitazone. Developed by SmithKline Beecham (now part of GlaxoSmithKline [GSK]), rosiglitazone (Avandia) (Figure 9) has become GSK’s secondleading brand product behind Advair, a combination inhaler for asthma. In 2004, Avandia garnered 5% of the company’s total pharmaceutical revenue (GSK, Annual Report, 2004). SmithKline Beecham won FDA approval to market rosiglitazone in May 1999 and launched the agent one month later in the United States. The company suffered a setback in its plans to introduce rosiglitazone in Europe when the European Union’s Committee for Proprietary Medicinal Products voted against approval in October 1999. SmithKline Beecham appealed the decision and gained approval for the drug in July 2000. As of 2002, rosiglitazone was available in France, Germany, Italy, Spain, and the United Kingdom. It has yet to be launched in Japan, where the drug has been preregistered since 2001. In preparation for launch, GSK entered into an agreement with Sankyo in 2003 to copromote rosiglitazone. An FDC of rosiglitazone and metformin is also available; it is discussed in detail later. Rosiglitazone is also in late-stage clinical trials for the treatment of psoriasis, Alzheimer’s disease, and rheumatoid arthritis (rosiglitazone has been shown to possess anti-inßammatory and immunomodulatory activities) (Pershadsingh HA, 2004). In the United States, rosiglitazone is approved as both monotherapy and combination therapy with a sulfonylurea, metformin, or insulin. In Europe, rosiglitazone is restricted to second-line therapy; it can be used as a monotherapy or in combination with another OAA. In November 2004, European Union (EU) regulatory authorities approved the use of rosiglitazone as part of a triple-combination therapy with metformin and a sulfonylurea. Although currently contraindicated in insulin-treated patients in Europe, GSK is actively seeking to expand the European labeling to this subpopulation. As noted earlier, rosiglitazone is a selective and potent agonist for PPARgamma that regulates the transcription of insulin-responsive genes involved in the control of glucose production, transport, and utilization as well as the regulation of fatty-acid metabolism. Its half-life is three to Þve hours and does not extend beyond this point when the parent compound is metabolized. Preliminary evidence suggests that rosiglitazone may have certain short-term beneÞts on pancreatic beta-cell function as demonstrated by the reduction in proinsulin:insulin ratios in type 2 diabetics (Lebovitz HE, 2001; Ovalle F, 2004; Smith SA, 2004). In short-term monotherapy trials, rosiglitazone reduced HbA1c in the range of 1.1–1.5% (summarized in Table 11). When combined with other OAAs, rosiglitazone reduces HbA1c by an additional 1–1.2% (Inzucchi SE, 2002). In one study, the combination of metformin and rosiglitazone, as separate agents, reduced HbA1c levels to a greater extent than metformin alone, demonstrating a synergistic effect. This 26-week, multicenter, double-blind, placebo-controlled trial involved 348 type 2 diabetics randomized to receive treatment with a Þxed dose of metformin (2.5 g/day) and placebo or metformin and rosiglitazone (4 mg/day or 8 mg/day) (Fonseca V, 2000). Mean HbA1c fell signiÞcantly from baseline in patients receiving 4 or 8 mg/day rosiglitazone in addition to metformin, by 0.56% and 0.78%, respectively. In contrast, HbA1c increased 0.45% in patients who

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FIGURE 6. Sites of action of the major pharmacotherapies for type 2 diabetes.

received the metformin/placebo combination. In another study, adding rosiglitazone (1 or 2 mg twice daily) to existing sulfonylurea regimens in 574 patients resulted in additional reductions of 0.59% and 1% in HbA1c , respectively, and 24 and 32 mg/dL in fasting plasma glucose (FPG), respectively (Wolffenbuttel BH, 2000). Rosiglitazone’s most common side effect is weight gain, a dose-dependent effect that occurs when the agent is used either as monotherapy or in combination with metformin. At the 4 and 8 mg/day dosages of rosiglitazone, median weight increased 2.6 kg (5.7 lb) and 4.5 kg (10 lb), respectively, after 26 weeks of monotherapy (Lebovitz HE, 2001). Weight increases were lower in the patients

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FIGURE 7. Side effects of PPAR-gamma agonists: peripheral edema and weight gain.

treated with rosiglitazone and metformin. Weight gains of 0.8 kg (1.76 lb) and 2.1 kg (4.62 lb) occurred at the 4 and 8 mg/day dosages of rosiglitazone, respectively, after 26 weeks of therapy (Fonseca V, 2000). The labeling indicates that, in clinical trials, hypoglycemia was recorded in 0.6% of patients treated with rosiglitazone monotherapy versus 0.2% of patients on placebo. Also included in the labeling are reports of anemia in 1.9% of patients

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FIGURE 8. Mechanism of action of PPAR-gamma agonists.

FIGURE 9. Structure of rosiglitazone.

on rosiglitazone monotherapy and in 0.9% of patients taking placebo, and edema in 4.8% of rosiglitazone patients and 1.3% of those receiving placebo. Pioglitazone. In August 1999, Takeda launched pioglitazone (Figure 10) in Japan and the United States, the latter in collaboration with Eli Lilly. The two companies subsequently launched the agent in Europe. The drug received priority review from the FDA, which concluded that pioglitazone has no signiÞcant adverse effects on hepatic function, lipid metabolism, or the heart. It is approved for use in the United States as a monotherapy or in combination with metformin, a sulfonylurea, or insulin. In Europe, pioglitazone is approved as a monotherapy

FIGURE 10. Structure of pioglitazone.

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(when metformin is contraindicated or not tolerated) or as part of combination therapy with other OAAs. Pioglitazone is not approved for combination use with insulin. Pioglitazone is Takeda’s leading international product, accounting for approximately 20% of the company’s net sales in its ethical drugs pharmaceutical segment. The company is taking a leadership role in the Þeld of diabetes by spearheading several international outcomes studies for pioglitazone and partnering with the Oxford Centre for Diabetes, Endocrinology, and Metabolism (OCDEM). In October 2004, Takeda Pharmaceuticals North America (TPNA) submitted a new drug application (NDA) for Actoplus Met, a twice-daily FDC of pioglitazone and metformin; this agent was approved by the FDA at the end of August 2005 (Takeda, press release, August 30, 2005). Meanwhile, Takeda and Andrx are developing a once-daily FDC product, using the latter company’s extended-release formulation of metformin, Fortamet (www.takeda.com, accessed on May 16, 2005). As mentioned earlier, pioglitazone is a selective and potent agonist for PPARgamma that regulates the transcription of insulin-responsive genes involved in the control of glucose production, transport, and utilization and in the regulation of fatty-acid metabolism. Like rosiglitazone, pioglitazone’s parent compound has a half-life of three to Þve hours, but the agent has a total half-life of 16–24 hours because of its active metabolites, as the labeling indicates. In several studies of obese diabetic animal models, investigators observed that pioglitazone prevented beta-cell damage and stress by reducing fat accumulation in the pancreatic islets, thereby improving insulin sensitivity and reducing oxidative stress (Diani AR, 2004; Ishida H, 2004; Kawasaki F, 2004). In short-term monotherapy trials of pioglitazone, HbA1c reductions of 1–1.6% were observed, as summarized in Table 11. Several studies demonstrated that combining pioglitazone with a sulfonylurea, metformin, or insulin potentiates the effects of the latter drugs. In a study of 560 patients treated with a sulfonylurea alone or with another antidiabetic agent, adding pioglitazone (15 or 30 mg once daily) to the sulfonylurea regimen reduced mean HbA1c levels by an additional 0.9% and 1.3%, respectively, and mean FPG by 39 mg/dL and 58 mg/dL, respectively, compared with adding placebo (Kipnes MS, 2001). In a study of 328 metformin-treated patients, adding pioglitazone (30 mg once daily) to metformin signiÞcantly reduced mean HbA1c levels by 0.8% and mean FPG levels by 38 mg/dL, compared with placebo (Einhorn D, 2000). Furthermore, the addition of pioglitazone signiÞcantly reduced TGs by 18.2% and increased HDL by 8.7%. In a combination study described in the package insert, 566 patients were treated with insulin (an average 60.5 units/day, alone or with another antidiabetic agent). Adding pioglitazone (15 or 30 mg once daily) to the insulin therapy signiÞcantly reduced mean HbA1c by 0.7% and 1.0%, respectively, and mean FPG by 35 mg/dL and 49 mg/dL, respectively, compared with placebo. Results from the Þrst long-term outcomes study designed to assess the effect of pioglitazone on macrovascular complications—the Prospective Pioglitazone Clinical Trial in Macrovascular Events (PROACTIVE) trial—were presented at

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the 41st meeting of the European Association for the Study of Diabetes in September 2005 (Takeda, press release, September 12, 2005; www.proactive-results.com, accessed September 13, 2005). More than 5,000 high-risk type 2 diabetic patients aged 35–75 with a history of cardiovascular disease (i.e., prior heart attack or stroke, acute coronary syndrome, previous bypass surgery) were enrolled in this prospective, multicenter, double-blind, randomized, placebo-controlled study. Patients were followed for a mean period of 2.5 years. The primary end points of this study included time from randomization to the occurrence of new macrovascular events or death. Patients were maintained on existing medications (e.g., OAAs, insulins, antihypertensive drugs, statins) and randomized to receive either pioglitazone (15 mg/day, eventually titrated up to 45 mg/day) or placebo. Patients in the pioglitazone group had a 16% reduced risk of heart attacks, stroke, and death compared with patients in the placebo group. Additionally, patients receiving pioglitazone exhibited a 0.8% reduction in HbA1c , 11.4% reduction in TG, 19% increase in HDL-C, and 7.2% increase in LDL-C at the end of the study. In comparison, patients given placebo saw 0.3% reduced HbA1c levels, 1.8% increased TG, 10.1% increased HDL-C, and 4.9% increased LDL-C. Excluding predeÞned cardiovascular end points, the most common serious adverse events included heart failure (5.7% of patients in the pioglitazone group versus 4.1% in the placebo group), hypoglycemia (27.9% versus 20.1%, respectively), and edema (21.6% versus 13.0%, respectively). Treatment with pioglitazone did not result in increased risk of liver toxicity or cancer. However, patients did experience an average weight gain of 3.6 kg. Pioglitazone’s labeling indicates that the most frequently reported adverse events in placebo-controlled trials were headache, upper respiratory tract infection, myalgia, pharyngitis, and tooth disorder. Edema was reported in 4.8% of pioglitazone-treated patients and 1.2% of placebo patients. Edema occurred more frequently in patients treated with a combination of pioglitazone and insulin (15% of pioglitazone/insulin-treated patients versus 7% of placebo patients). Weight gain ranging from 0.5 kg to 2.8 kg was also observed with pioglitazone. Biguanides Overview. Only one biguanide is available for the treatment of type 2 diabetes: metformin (Bristol-Myers Squibb [BMS]’s Glucophage and Glucophage XR, Biovail/DepoMed’s Glumetza, generics). Since the publication of the results of the landmark UKPDS in the 1990s, metformin has become the preferred Þrst-line therapy for type 2 diabetes, particularly in obese patients. Mechanism of Action. Metformin’s precise mechanism of action is unknown, although research shows that its primary action is to reduce hepatic glucose production—which is elevated in diabetic patients—by inhibiting the activity of glucose-6-phosphatase (Hundal RS, 2000; Mithieux G, 2002; Stumvoll M, 1995), as depicted in Figure 11. Other studies indicate that unlike with the sulfonylureas and meglitinides, which target pancreatic beta-cell insulin secretion, metformin’s secondary effect is insulin sensitization (Lupi R, 1999; Mather KJ, 2001). A

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FIGURE 11. Metformin’s mechanism of action.

preliminary study suggests that metformin may also reduce the risk of CVD by easing insulin resistance, which is thought to be central to the pathogenesis of endothelial function in diabetics (Mather KJ, 2001). Improvements in vascular endothelial function have positive implications for treating vascular abnormalities, such as the impaired endothelium-dependent responses commonly observed in type 2 diabetes patients. Metformin. Metformin (Figure 12) plays a central role in pharmacological therapy for type 2 diabetes. Introduced in 1959 in France, it gradually became available throughout Europe and Japan. In 1995, the agent was launched in the

FIGURE 12. Structure of metformin.

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TYPE 2 DIABETES

United States, where it is approved for use as a monotherapy or in combination with other OAAs and with insulin therapy. In 1999, the FDA asked BMS to test metformin in children with type 2 diabetes, and in December 2000, the agency approved it for diabetics younger than age 18. The Þrst formulation of metformin that became available was the immediaterelease (IR) formulation (BMS’s Glucophage), which requires dosing two to three times daily. BMS subsequently released an extended-release (XR) formulation, which can be administered once daily. (Andrx also has an XR formulation of metformin, Fortamet.) The most recent once-daily formulation of metformin to enter the market is metformin GR (Biovail/DepoMed’s Glumetza). Biovail licensed U.S. and Canadian manufacturing and marketing rights for metformin GR in May 2002. The two companies Þled an NDA with the FDA for the once-daily, extended-release formulation of metformin in April 2004 and received approval to market these formulations of metformin in June 2005 (Biovail, press release, June 3, 2005). Metformin GR was developed using DepoMed’s gastric retention (GR) technology, which promises to cause a lower incidence of gastrointestinal side effect because of preferential absorption in the upper gastrointestinal tract. In February 2003, DepoMed reported positive results from its Þrst randomized, double-blind, dose-ranging, parallel-design Phase III study. In this study, 536 type 2 diabetics received an undisclosed regimen of BMS’s metformin IR or one of three undisclosed regimens of DepoMed’s metformin GR for six months. According to DepoMed, all three dosing regimens of metformin GR signiÞcantly reduced glycosylated hemoglobin levels as effectively as metformin IR. Additionally, an open-label follow-up study with 250 subjects from the original trial demonstrated that metformin GR consistently lowered levels of glycosylated hemoglobin and FPG (DepoMed, press release, December 10, 2003). The evidence to support the widespread use of metformin is bolstered by the results of the landmark UKPDS. The UKPDS is the largest and longest outcomes trial ever undertaken in the study of diabetes treatment. The UKPDS involved more than 7,600 patients with a median follow-up time of ten years. Major Þndings from the UKPDS include the following: •

•

•

Good control of blood pressure (<150/85 mmHg) reduced the risks of macrovascular (e.g., stroke, heart failure) and microvascular complications (e.g., diabetic retinopathy, nephropathy, and neuropathy). HbA1c levels equivalently achieved by intensive pharmacological therapy (7.0%) with glyburide, metformin, and insulin reduced microvascular complications by 25%, compared with HbA1c levels achieved by conventional therapy (i.e., lifestyle modiÞcations) (7.9%). Over the length of the study, pancreatic beta-cell function declined in patients and combination therapy (polypharmacy) was needed to achieve optimal blood glucose control. Treatment regimens must be periodically adjusted to prevent progressive worsening of hyperglycemia in patients taking OAAs.

Aside from the sulfonylureas and insulin, no other antidiabetic agents have published outcomes data at this time. Results from this long-term clinical trial suggest

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that metformin monotherapy reduces the risk of diabetic complications and mortality (UKPDS, 1998). The UKPDS 34 trial involved 1,704 newly diagnosed obese type 2 diabetes patients and an average follow-up period of ten years. Subjects were randomized to one of two arms: (1) conventional therapy with diet alone; or (2) intensive pharmacological therapy with metformin, aiming for an aggressive FPG below 6 mmol/L. Results indicate that intensive glucose therapy with metformin reduces the risk of diabetes-related complications: •

•

•

Metformin-treated subjects exhibited a risk reduction of 32% (compared with patients treated with diet alone) for any diabetes-related end points (p = 0.002), including death from hyperglycemia or hypoglycemia, cardiac events, stroke, renal failure, amputation, and retinopathy. SpeciÞcally, metformin-treated patients demonstrated a 30% lower risk than the diet-treated group for developing macrovascular complications. However, microvascular complications were not signiÞcantly reduced, presumably because of the small sample size. The metformin-treated group experienced risk reductions of 42% for diabetes-related deaths (p = 0.02) and 36% for all-cause mortality (p = 0.01), compared with the diet-treated group.

During the ten-year follow-up period of the UKPDS, the median FPG and HbA1c levels of metformin-treated patients (8 mmol/L and 7.4%, respectively) were consistently lower than those of diet-treated patients (9.5 mmol/L and 8.0%, respectively). Although both parameters dropped from baseline levels in the metformin-treated group in the Þrst year, they rose gradually over the next ten years—but remained below the levels of the diet-treated group. For example, the median HbA1c levels of the metformin- and diet-treated groups during the Þrst Þve years were 6.7% and 7.5%, respectively, rising to 7.9% and 8.5%, respectively, in the last Þve years (UKPDS, 1998). Short-term monotherapy trials with metformin show that, on average, metformin leads to a 1.1–1.5% reduction in HbA1c levels compared with placebo, as summarized in Table 11. Because metformin is frequently used as a Þrst-line therapy and current medical practice dictates a stepwise, add-on approach, there are many clinical trials in which metformin is a component of the combination therapy. An additional reduction of approximately 0.5–1.5% in HbA1c has been observed when the drug is used in combination therapy with either insulin or another OAA (Inzucchi SE, 2002). Metformin is as effective as a sulfonylurea in reducing HbA1c but does not cause hypoglycemia or weight gain because it does not act on insulin secretory mechanisms. In the UKPDS, the rate of any hypoglycemic episode was lower in metformin-treated patients than in sulfonylurea- or insulin-treated patients (UKPDS, 1998). The percentages of patients per year who experienced one or more major hypoglycemic episodes in the conventional sulfonylurea, insulin, and metformin groups were 0.7%, 3.1%, and 0.3%, respectively. As a monotherapy, metformin’s chief adverse effect is the risk of lactic acidosis. Although the incidence of this condition is rare, it is fatal in approximately

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50% of patients, as stated in the package insert. Therefore, metformin is contraindicated in patients who are prone to lactic acidosis and unable to metabolize lactate. This group includes patients with hepatic and renal insufÞciencies, severe cardiac or respiratory disease, chronic metabolic acidosis, or a history of alcohol abuse. Caution is advised for elderly patients as well. Metformin use is temporarily halted for surgical procedures and for procedures that require iodinated contrast media. Gastrointestinal irritation (nausea, diarrhea, abdominal cramps, and altered taste) has been reported in about one-third of patients during the Þrst few weeks of therapy. These side effects tend to diminish with continued use and can be minimized by initiating therapy with low doses of metformin and by taking the drug with food. Sulfonylureas Overview. Derived from sulfonic acid and urea, sulfonylureas are insulin secretagogues that control glycemia by inducing insulin secretion. Developed in the 1950s, sulfonylureas have long been a preferred Þrst-line therapy for managing type 2 diabetes, either as monotherapy or in combination with another OAA or insulin. Patients who generally respond well to this treatment are those diagnosed before age 40 who have less than Þve years of symptomatic disease before therapy initiation and an FPG level below 300 mg/dL. Non-obese patients with insulinopenia are also good candidates for this therapy. Three generations of sulfonylureas are on the market, each different from the others in potency and pharmacokinetics. First-generation agents, which are rarely used today, include tolbutamide (generics), tolazamide (generics), acetohexamide (generics), and chlorpropamide (PÞzer’s Diabinese, generics). The secondgeneration agents are short-acting compounds with half-lives ranging from two to ten hours; as a group, they are 100 times more potent than Þrst-generation agents because of improvements in their binding properties. Members of this group include glyburide/glibenclamide (PÞzer’s Micronase, SanoÞ-Aventis’s Diabeta, generics), glipizide (PÞzer’s Glucotrol, Glucotrol XL, generics), and gliclazide (Servier’s Diamicron, Molteni & C.F. LLI Alitti’s Diabrezide, Dainippon’s Glimicron, generics). Glimepiride (SanoÞ-Aventis’s Amaryl) is the only third-generation sulfonylurea available; this agent offers greater potency, has a longer half-life (i.e., it can be dosed once-daily), and carries a lower risk of causing hypoglycemia. In 1970, the University Group Diabetes Program found early evidence of cardiovascular morbidity from tolbutamide; this controversial study was highly criticized and its Þndings were never reproduced. The UKPDS refuted those Þndings and deemed sulfonylureas safe (UKPDS Group, 1998). Nevertheless, the package inserts of tolbutamide and glyburide contain a special warning regarding the increased risk of cardiovascular mortality. Mechanism of Action. Sulfonylureas induce Þrst-phase insulin secretion. Figure 13 illustrates their mechanism of action. These agents bind to the 140 kDa sulfonylurea receptors (SUR-1) on the cell surface of pancreatic beta cells. The SUR-1 receptors are linked to Kir6.2 adenosine triphosphate (ATP)-sensitive

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FIGURE 13. Mechanisms of action of the sulfonylureas and meglitinides.

FIGURE 14. Structure of glimepiride.

potassium channels; binding of sulfonylureas to the SUR-1 receptor thereby reduces potassium conductance and depolarizes the cell membrane to induce insulin secretion. Preliminary research in type 2 diabetic and nondiabetic patients suggests that these agents may have insulin-sensitizing properties—that is, they may increase peripheral glucose utilization (Clark HE, 1996; Raptis SA, 1999). Glimepiride. Glimepiride (SanoÞ-Aventis’s Amaryl) (Figure 14) is a thirdgeneration sulfonylurea available in all seven major pharmaceutical markets. As the second-highest-grossing product in SanoÞ-Aventis’ diabetes franchise, behind insulin glargine (Lantus), it is an important component of the company’s diabetes portfolio. In addition, the company is developing an FDC of glimepiride and metformin and is working closely with its generics arm, Winthrop Medicine, to provide generic formulations of Amaryl to certain markets. These markets were not disclosed in the company report (SanoÞ-Aventis, annual report, 2004). Glimepiride works by binding to SUR-1 receptors, which are coupled to the Kir6.2 ATP-sensitive potassium channels. Although its efÞcacy resembles that of a second-generation drug, it controls glucose at slightly lower doses because of its higher afÞnity for the sulfonylurea receptor (Draeger KE, 1996). Glimepiride is also more speciÞc for pancreatic beta cells, thereby reducing unwanted binding with the ATP-sensitive potassium channels of the heart (Lewis J, 2001; Smits P, 1996). Glimepiride may also reduce insulin resistance. A 24-week, open-label

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trial involving 14 type 2 diabetic men found that glimepiride lowered HbA1c through improved insulin secretion and insulin sensitivity, as indicated by Þrstphase insulin secretion and the insulin sensitivity index (Kabadi MU, 2004). Double-blind, randomized, one-year, placebo-controlled trials of glimepiride demonstrated 1.4–2.5% reductions in HbA1c , compared with placebo, as summarized in Table 11. The addition of glimepiride to the regimens of type 2 diabetes patients uncontrolled with metformin improves glycemic control: an HbA1c reduction of 0.8–0.9% was observed in combination-treated patients compared with metformin or sulfonylurea monotherapy (Charpentier G, 2001). Like many sulfonylureas, glimepiride may cause hypoglycemia. This side effect is most likely to occur in patients who are elderly, who have reduced renal functioning, or who have irregular eating schedules (Inzucchi SE, 2002). Glimepiride does, however, have a pharmacokinetic proÞle that suggests it is less likely than other sulfonylureas, particularly glibenclamide, to cause this condition (McCall AL, 2001). Patients on glimepiride are less at risk of experiencing cardiovascular complications than are patients on other sulfonylureas. Unlike other sulfonylurea derivatives, no signiÞcant weight gain is linked to glimepiride use (Jasik M, 2000; Lewis J, 2001). Fixed-Dose Combinations Overview. For patients who require therapy with more than one agent, singlepill FDCs provide a convenient alternative to taking two separate pills. All available combination agents contain metformin plus an OAA, thus reßecting metformin’s core position in the type 2 diabetes therapeutic regimen. Mechanism of Action. Each of the combination agents discussed in this section possesses two mechanisms of action: that of metformin and that of the agent with which metformin is combined. Although metformin’s precise mechanism of action is unknown, research shows that its primary action is to reduce hepatic glucose production by inhibiting the activity of glucose-6-phosphatase (Hundal RS, 2000; Mithieux G, 2002; Stumvoll M, 1995). Metformin/Rosiglitazone. In November 2002, GlaxoSmithKline launched Avandamet, an FDC of metformin/rosiglitazone, in the United States. It is also available in France, Germany, Spain, and the United Kingdom. This FDC is indicated for patients with type 2 diabetes who already receive rosiglitazone and metformin as separate tablets or who are not adequately controlled on metformin alone. In March 2005, the FDA and the Department of Justice seized Avandamet supplies because of violations in good manufacturing practices (GMP) in the Puerto Rico and Tennessee manufacturing facilities. FDA inspections revealed that some lots of Avandamet were not always manufactured uniformly such that there might be inconsistencies in the dosing of rosiglitazone. GlaxoSmithKline was allowed to continue manufacturing Avandamet following reviews by the FDA and an independent expert (GlaxoSmithKline, press release, April 27, 2005).

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A selective and potent agonist for PPAR-gamma, rosiglitazone regulates the transcription of insulin-responsive genes involved in the control of glucose production, transport, and utilization as well as in the regulation of fatty-acid metabolism. Metformin reduces hepatic glucose production by inhibiting the activity of glucose-6-phosphatase. Although metformin and rosiglitazone were administered concomitantly in several randomized, placebo-controlled studies, only one trial has evaluated the efÞcacy of the metformin/rosiglitazone FDC product. A 24-week, single-arm, open-label study examined the efÞcacy of an FDC of metformin/rosiglitazone in 190 drug-naive patients with poorly controlled type 2 diabetes (i.e., HbA1c >11% or FPG > 270 mg/dL) (Rosenstock J, 2005[a]). At the end of the study, patients exhibited an average HbA1c reduction of 4% and an FPG reduction of 139 mg/dL. Furthermore, 50% of the patients achieved HbA1c <7% and 38% of patients achieved HbA1c <6.5%. Only 2% of patients reported hypoglycemia. Diarrhea was the most common adverse effect (12%), and the incidence of edema was 3%. Median weight gain from baseline was 2.2 kg. In dual-therapy trials, concomitant use of metformin and rosiglitazone has proven to be beneÞcial. A randomized, double-blind, multicenter, Phase III trial enrolled 348 type 2 diabetic patients who received 250 mg of metformin plus placebo, a 250 mg/4 mg (metformin/rosiglitazone) combination, or a 250 mg/8 mg combination for 26 weeks (Fonseca V, 2000). Mean levels of HbA1c fell 1.0% in the 4 mg metformin/rosiglitazone group and 1.2% in the 8 mg metformin/rosiglitazone group compared with the metformin/placebo group. The combination treatment also improved insulin sensitivity and beta-cell function more effectively than treatment with metformin alone. Metformin/Glyburide. BMS’s Glucovance is an FDC of metformin and glyburide (Figure 15), a sulfonylurea. Metformin reduces hepatic glucose production by inhibiting the activity of glucose-6-phosphatase, while glyburide increases insulin secretion. This FDC is available only in the United States and Italy. A randomized, placebo-controlled, Phase III clinical trial involved 806 drugnaive type 2 diabetic patients aged 60 or older who received either glyburide alone, metformin alone, a 1.25 mg/250 mg (glyburide/metformin) FDC, a 2.5 mg/500 mg FDC, or placebo. After Þve months, absolute reductions in HbA1c were 1.0% with glyburide, 0.8% with metformin, and 1.3% for either FDC, compared with placebo (Garber A, 2000). Similar results were obtained in a 52-week

FIGURE 15. Structure of glyburide.

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TYPE 2 DIABETES

FIGURE 16. Structure of glipizide.

open-label extension study of 477 patients who had failed to maintain glycemic control using sulfonylurea monotherapy. Patients were divided into different treatment arms and were treated once- to four-times daily with 2.5 mg/500 mg FDC of glyburide/metformin. Patients experienced an average reduction in HbA1c of 1.7% at the conclusion of the study (Blonde L, 2004). Other studies of metformin/glyburide combination therapy, in which each was given separately, produced similar Þndings. A 29-week, randomized, double-blind study of 632 type 2 diabetes patients demonstrated that metformin/glyburide combination therapy, administered concurrently as separate pills, provided a synergistic reduction in blood glucose levels compared with either agent alone (DeFronzo RA, 1995). Metformin/Glipizide. The FDC of metformin and glipizide (BMS’s Metaglip) (Figure 16) reached the U.S. market in 2002. Metformin reduces hepatic glucose production by inhibiting the activity of glucose-6-phosphatase. Glipizide is a sulfonylurea that improves glycemic control by increasing insulin secretion by pancreatic beta cells. In a multi-center, double-blind, 24-week study, 868 drug-naive type 2 diabetes patients were randomized into Þve treatment arms: 1.25/250 mg glipizide/metformin; 2.5/250 mg glipizide/metformin; 2.5/500 mg glipizide/metformin; 500 mg metformin; or 5 mg glipizide (Rubin CJ, 2002[a]). Results demonstrated that the single-pill combinations effected an additional reduction of 0.4–0.7% in HbA1c , compared with metformin or glipizide alone. Approximately 60% of patients treated with the FDC achieved an HbA1c of less than 7%, compared with 40% in the metformin and glipizide groups. In a similar study, 247 type 2 diabetes patients who were inadequately controlled with sulfonylureas were randomized into three groups: 5/500 mg glipizide/metformin; 30 mg glipizide; or 500 mg metformin (Rubin CJ, 2002[b]). Patients who took the FDC experienced additional reductions of 1% in HbA1c and 37 mg/dL in FPG, compared with patients taking metformin or glipizide alone. Approximately 36% of patients achieved HbA1c of less than 7% in the single-pill combination arm, compared with 10% in the metformin and glipizide groups. Meglitinides Overview. The meglitinides, or benzoic acid derivatives, are a novel class of insulin secretagogues that includes repaglinide (Novo Nordisk’s Prandin/NovoNorm), the Þrst in its class; nateglinide (Novartis’s Starlix/Starsis);

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FIGURE 17. Structure of repaglinide.

and mitiglinide (Kissei/Takeda’s Glufast), the newest member. These agents, characterized by rapid onset and short duration of action, target postprandial hyperglycemia and demonstrate efÞcacy comparable to that of an older class of drugs, the sulfonylureas, in reducing HbA1c (Wolffenbuttel BH, 1999). Meglitinides, commonly used in adjunct therapies, have found a niche market in the elderly population because they pose a lower risk for hypoglycemia. Mechanism of Action. Meglitinides bind to a putative nonsulfonylurea receptor on the pancreatic beta cell and induce insulin secretion in a manner similar to that of the sulfonylureas, as illustrated in Figure 13. Although both sulfonylureas and meglitinides induce secretion, the latter exert their effects only in the presence of elevated glucose levels, thus minimizing the risk of hypoglycemia. Repaglinide. Boehringer Ingelheim developed repaglinide (Figure 17) with Novo Nordisk. The compound was approved in December 1997 and launched in the United States in June 1998 as monotherapy and in combination with metformin for type 2 diabetes. Repaglinide became available in Europe in 1998 and is in Phase III development in Japan by Sumitomo Pharmaceuticals, which licensed the rights to develop and market rapaglinide from Novo Nordisk in 2004 (Sumitomo Pharmaceuticals, press release, September 16, 2004). Repaglinide binds to a putative nonsulfonylurea receptor on the pancreatic beta cell and induces insulin secretion in the presence of elevated glucose levels. Repaglinide has been shown to reduce HbA1c levels by 1.6–1.9% in randomized, placebo-controlled clinical trials (Goldberg RB, 1998; Jovanovic L, 2000). In a 24-week, multicenter, double-blind, randomized trial, 361 patients received preprandial treatment with placebo or repaglinide (1 or 4 mg daily) (Jovanovic L, 2000). In the repaglinide-treated groups, changes in HbA1c levels were 1.9% lower than in the placebo group. According to the labeling, mild or moderate hypoglycemia occurred less often in patients treated with repaglinide (16%) than in patients treated with sulfonylureas (20%); severe hypoglycemia with coma, seizure, or other neurological impairment occurred rarely. Nateglinide. Nateglinide (Novartis/Yamanouchi/SanoÞ-Aventis Japan’s Starlix/Starsis/Fastic) (Figure 18), a D-phenylalanine derivative, is a rapid, shortacting insulin secretagogue that targets postprandial hyperglycemia. It was

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TYPE 2 DIABETES

FIGURE 18. Structure of nateglinide.

approved in the United States in December 2000 as a monotherapy and in combination therapy with metformin and was launched in February 2001. Nateglinide is marketed by Novartis in Germany and the United Kingdom. Yamanouchi and Morishita originally developed the agent in Japan and then licensed it to Novartis for development outside Asia. Nateglinide was launched in Japan by Ajinomoto and Novartis in August 1999 under the trade name Starsis. Like repaglinide, nateglinide binds to a putative nonsulfonylurea receptor on the pancreatic beta cell and induces insulin secretion in the presence of elevated glucose levels. The reduction in the risk of type 2 diabetes in patients taking the antihypertensive agent valsartan (Novartis’s Diovan, an angiotensin II receptor antagonist [AIIRA]) has led to the initiation of a large-scale prospective trial: the Nateglinide and Valsartan in Impaired Glucose Tolerance Outcomes Research (NAVIGATOR) study. In addition to reducing blood pressure, studies have shown that AIIRAs may improve insulin sensitivity. The NAVIGATOR study will investigate the combined efÞcacy of nateglinide and valsartan in preventing CVD and type 2 diabetes in 39,000 individuals previously diagnosed with impaired glucose tolerance. The trial is expected to conclude in 2008. Nateglinide has been shown to lower HbA1c by 0.6–1.0% in randomized placebo-controlled clinical trials (HaneÞeld M, 2000; Horton ES, 2000). Additional HbA1c reductions of 0.6–0.9% were observed when nateglinide was used in combination with metformin (Horton ES, 2000). In a 24-week, randomized, double-blind study, patients received either 120 mg nateglinide before meals, 500 mg metformin three times a day, combination therapy, or placebo. Researchers found that HbA1c levels fell 0.5% more with nateglinide, 0.8% more with metformin, and 1.4% more with combination therapy than with placebo (Horton ES, 2000). As nateglinide’s labeling indicates, hypoglycemia occurred in 2% of treated patients in placebo-controlled clinical trials; 0.3% discontinued therapy because of hypoglycemia. Mitiglinide. In August 2002, Kissei and Takeda agreed to comarket mitiglinide∗ in Japan and to launch the product under the single brand name Glufast. It was approved in Japan in January 2004. Mitiglinide is in Phase II development in the United States by Kissei and in Phase III in Europe by Servier. Data from latephase mitiglinide trials in type 2 diabetics have not been published nor have data comparing mitiglinide’s effect on glucose levels with that of currently available members of this class. In announcing that it would comarket mitiglinide with

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Takeda, Kissei described the results of a completed double-blind comparative Phase III clinical trial (Kissei, media release [electronic], August 5, 2002). In this study, mitiglinide signiÞcantly improved three indices of glucose levels: glycosylated hemoglobin, postprandial plasma glucose, and FPG. Kissei also claimed that the incidence of hypoglycemia in this study was equivalent to that of placebo. Alpha-Glucosidase Inhibitors Overview. The primary effect of alpha-glucosidase inhibitors (AGIs) is to lower postprandial glucose and improve glycemic control without triggering weight gain or hypoglycemic events. AGIs generally exert only a modest effect, reducing HbA1c levels by about 0.5–1%, and approximately 50% of patients experience ßatulence, abdominal pain, and/or diarrhea (Coniff RF, 1994; Fischer S, 1998; Holman R, 1999; Inzucchi SE, 2002; Johnston PS, 1998; Vichayanrat A, 2002). AGIs are used as alternatives for patients, particularly the elderly, who have mild-to-moderate hyperglycemia and for patients who are prone to hypoglycemia or lactic acidosis. The AGIs on the market today are acarbose (Bayer’s Precose/Glucobay, generics), miglitol (Bayer/PÞzer/SanoÞ-Aventis’s Glyset), and voglibose (Takeda’s Basen). The Þrst AGI introduced for type 2 diabetes, acarbose is a complex oligosaccharide obtained from the fermentation processes of the microorganism Actinoplanes utahensis. Although its glucose-lowering efÞcacy is modest, acarbose is one of the safest oral antidiabetics available. Therefore, several studies have investigated its therapeutic role in patients with impaired glucose tolerance. These trials demonstrated that acarbose is effective in reducing cardiovascular risk and can prevent the progression to type 2 diabetes in prediabetic individuals. The most prominent of these studies is the Study to Prevent Non-Insulin-Dependent Diabetes Mellitus (STOP-NIDDM) trial, a multicenter, placebo-controlled, randomized study of 1,492 patients with impaired glucose tolerance who were followed for three years. Results demonstrated that acarbose reduced the risk of progressing from impaired glucose tolerance to developing type 2 diabetes by 25% (Chiasson JL, 2002). Mechanism of Action. AGIs hinder carbohydrate digestion at the brush border of the intestinal epithelium, as illustrated in Figure 19, effectively delaying carbohydrate absorption and allowing the delayed insulin secretion characteristic of type 2 diabetics to catch up with carbohydrate absorption. Consequently, AGIs reduce PPG levels. These agents concentrate in the intestinal enterocytes, where they act locally, minimizing the risk of hypoglycemia. Acarbose. The Þrst AGI introduced, acarbose (Figure 20) was launched in Europe and Japan in 1990 and in the United States in 1996. It is a complex oligosaccharide obtained from the fermentation processes of the microorganism Actinoplanes utahensis. As illustrated in Figure 19, acarbose inhibits an enzyme target distinct from that of miglitol.

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FIGURE 19. Mechanism of action of the alpha-glucosidase inhibitors.

FIGURE 20. Structure of acarbose.

A multicenter, double-blind, placebo-controlled trial evaluating the efÞcacy and safety of acarbose (25, 50, 100, or 200 mg three times daily) in 495 patients found that the agent lowers HbA1c levels by 1% in comparison with placebo (Fischer S, 1998). In clinical trials of acarbose, 45% of patients experienced increased ßatulence, 10% reported abdominal pain, and 19% reported diarrhea (Coniff RF, 1994; Holman R, 1999). Following reports of mortality due to fulminant hepatitis, Korosho, the advisory board to the Japanese Ministry of Health, Labor, and Welfare, recommended that the acarbose package insert include a warning that it may cause serious hepatic function disorders, such as fulminant hepatitis. Miglitol. Miglitol (Bayer/PÞzer/SanoÞ-Aventis’s Glyset) (Figure 21) is a pseudomonosaccharide with potency like that of acarbose and many of the same side effects. Bayer licensed miglitol to SanoÞ-Synth´elabo (now SanoÞ-Aventis) in November 1997 for all markets outside of North America and Japan and to Pharmacia & Upjohn (now PÞzer) in August 1998 for North America, Australia,

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FIGURE 21. Structure of miglitol.

FIGURE 22. Structure of voglibose.

and New Zealand. Miglitol has been launched in the United States and in most European countries but is not yet available in Japan. Like acarbose, miglitol inhibits carbohydrate digestion at the brush border of the intestinal epithelium. However, miglitol targets a different digestive enzyme than does acarbose. In a randomized, double-blind trial, 345 patients received miglitol (50 or 100 mg three times daily) or placebo (Johnston PS, 1998). Researchers found that therapy with miglitol reduced HbA1c levels by 0.7% in comparison with placebo. According to the product’s labeling, therapy with miglitol, as with acarbose, is associated with increased ßatulence, abdominal pain, and diarrhea. Voglibose. Voglibose (Takeda’s Basen) (Figure 22) is available only in Japan, where it launched in 2005. This agent’s mechanism of action is the same as that of acarbose and miglitol. In a randomized, 30-patient clinical trial to compare the safety and efÞcacy of voglibose (0.2 mg three times daily) and acarbose (100 mg three times daily), voglibose was associated with slightly less efÞcacy for postprandial glucose reduction. Among patients receiving voglibose, 57% reported increased ßatulence and 10% reported diarrhea (Vichayanrat A, 2002). Insulin and Insulin Analogues Overview. In the research and development of insulin, pharmaceutical companies have followed two complementary strategies for improving glycemic control: improving the pharmacokinetic proÞles of insulins and creating moreconvenient devices to administer insulin. Currently, type 2 diabetics use insulin for maintaining basal insulin levels between meals and/or achieving appropriate increases in circulating insulin levels at mealtimes. The purpose of the steady release of low levels of insulin is to maintain normal blood glucose levels during

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fasting (between meals and during sleep), whereas bolus insulin administration is required to control postprandial hyperglycemia. Products designed for strictly basal use are divided into two release proÞles: intermediate-acting and long-acting. To administer a pre-meal bolus of insulin, patients use short-acting insulins. Insulin was historically derived from animal sources (bovine or porcine insulins), but over the past decade, the market has shifted to recombinant human insulin, which is expressed in either Escherichia coli or Saccharomyces cerevisiae and causes fewer immune reactions. These genetically modiÞed bacteria produce recombinant human insulin; the Þnal product may be modiÞed chemically to attain the desired activity proÞle (short-, intermediate-, or long-acting). Insulin analogues, also produced as recombinant peptides, achieve speciÞc release proÞles by substitution of key amino acids in the native insulin peptide. Thus far, marketed insulin analogues include short- and long-acting insulins. The shortacting insulin glulisine (SanoÞ-Aventis’s Apidra) is the latest analogue to enter the diabetes market, joining insulin lispro (Eli Lilly’s Humalog) and insulin aspart (Novo Nordisk’s NovLog/NovoRapid). Insulin glargine (SanoÞ-Aventis’s Lantus) was the only available long-acting insulin analogue until insulin detemir (Novo Nordisk’s Levemir) launched in 2005 in Germany and the United Kingdom. This discussion focuses on the insulin analogues lispro and glargine because of their growing popularity in type 2 diabetes treatment. Formulations. A wide variety of injectable recombinant human insulin and insulin analogue products with different durations of action are available (Table 12). The onset and duration of action of recombinant human insulin can be unpredictable, depending on the patient’s eating and exercise habits; therefore, glycemic control and dosing adjustment may be problematic. By mimicking the body’s natural insulin response, insulin analogues improve glycemic control while causing fewer hypoglycemic events. Long-term improvements in glycemic control should reduce patients’ risk of developing diabetic complications. Historically, PCPs referred patients who require insulin therapy to endocrinologists, who were better equipped to titrate patients’ requirements for human insulins, which are subject to tremendous interpatient variability. Endocrinologists therefore did not see many patients until the late stages of disease. The development of insulin analogues, particularly the long-acting basal insulins, has made it easier for PCPs to prescribe insulin and, therefore, has resulted in earlier use of insulin in the management of type 2 diabetes. Devices. Insulin can be administered via three types of devices: syringes, pens, and pumps. Table 13 describes the various devices used to deliver insulin. Insulin is provided as vials or cartridges, depending on the type of device used. Unopened vials or cartridges are stored in the refrigerator up to the expiration date or at room temperature up to 28 days. Opened vials should be used within 28 days and can be stored in the refrigerator or at room temperature. Opened insulin cartridges (which are typically insulin analogues) should also be used within 28 days and must be kept at room temperature.

TABLE 12. Insulin Formulations Description Short-acting recombinant insulin Rapid-acting insulin analogues

Intermediate-acting recombinant insulin Long-acting recombinant insulin Long-acting insulin analogues

Agent

O nset of Action

Peak of Action

Duration of Action

Regular human insulin (Lilly’s Humulin R; Novo Nordisk’s Novolin R) Insulin lispro (Lilly’s Humalog); insulin aspart (Novo Nordisk’s NovoLog/NovoRapid); insulin glulisine (Sanofi-Aventis’ Apidra) Neutral protamine Hagedorn (NPH) insulin (Lilly’s Humulin N; Novo Nordisk’s Novolin) Ultralente insulin (Lilly’s Humulin U) Insulin glargine (Sanofi-Aventis’ Lantus); insulin detemir (Novo Nordisk’s Levemir)

30–60 min

2–3 hr

5–8 hr

5–15 min

30–90 min

3–5 hr

1–3 hr

4–10 hr

14–18 hr

6–10 hr

8–12 hr

20–36 hr

1 hr

None

24 hours

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Mechanism of Action. In type 2 diabetes, insulin therapy supplements the patient’s own insulin supply. Insulin-resistant patients are unable to produce sufÞcient quantities of insulin to maintain serum glucose levels as a result of the exhaustion of the insulin-producing capabilities of the pancreatic beta cells. The addition of exogenous insulin compensates for the body’s inability to produce an adequate insulin response. The manufactured insulin products mimic the physiological actions of endogenous insulin. However, because they must be introduced into the body, they are not able to mimic perfectly the dynamic response of a healthy pancreas to variations in blood glucose levels. Insulin Lispro. Insulin lispro (Lilly’s Humalog), a rapid-acting (bolus) insulin, was the Þrst insulin analogue on the market. It has a reversed order of amino acids at positions 28 and 29 and is produced in Escherichia coli as a recombinant peptide. Although as potent as regular insulin, lispro has a shorter onset of action (15 minutes) and duration of action (three to four hours). Clinical studies show that lispro, in combination with a sulfonylurea, offers improvements to FPG and postprandial glucose levels as well as reductions in HbA1c levels, compared with treatment with a sulfonylurea alone. The biphasic Humalog Mix 75/25 (75% insulin lispro protamine suspension, an intermediate-acting agent, and 25% insulin lispro solution, a rapid-acting agent) has the same duration of action as regular insulin but a faster onset. Insulin lispro is indicated for the treatment of diabetes (type 1 or 2) in conjunction with long-acting insulin. In a 5.5-month, randomized, open-label, parallel trial involving type 2 diabetics who failed sulfonylurea therapy, 148 patients were randomized to twice-daily insulin lispro or rapid-acting human regular insulin, both in combination with neutral protamine hagedorn (NPH), an intermediate-acting insulin (Ross SA, 2001). Results demonstrated that postprandial glucose elevations after breakfast and dinner were signiÞcantly lower in the lispro-treated arm than in the regular insulin-treated arm. However, an average HbA1c reduction of 2.5% was similar in both groups. During the day, rates of hypoglycemia were similar in both arms, but nocturnal hypoglycemia occurred less often in patients treated with lispro than with regular human insulin. Insulin Glargine. Insulin glargine (SanoÞ-Aventis’s Lantus) is a recombinant protein that differs from human insulin by three amino acids. It provides a longer, peakless duration of action that more effectively meets basal insulin requirements than currently available intermediate- and long-acting insulins. In May 2003, the FDA approved its ßexible administration schedule, meaning that patients can now administer the agent at any time during the day rather than only at bedtime. A 28-week Phase III study compared insulin glargine with NPH insulin in 518 type 2 diabetics who were not taking oral agents and had previously received a long-acting basal insulin (with or without preprandial injections of rapid-acting regular human insulin) (Rosenstock J, 2001). Researchers reported no differences in reductions of HbA1c between the study groups: mean HbA1c levels were 8.6% and 8.5% in the insulin glargine and NPH groups, respectively. Similar percentages of patients exhibited an FPG of less than 6.7 mmol/L: 29.6% of the insulin

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TABLE 13. Devices Used to Administer Insulin Device

Description

Comments

Syringe

Syringes are fine-gauge hypodermic needles attached to hollow barrels. Needles have very sharp points, most of which have a special coating to ease entry through the skin upon injection. Insulin is provided in vials from which the patient draws up the insulin into the syringe. Insulin pens are a type of syringe to which a cartridge containing insulin is attached. Some devices offer replaceable cartridges with reusable pens, while others have disposable pens with prefilled cartridges. Pens are attached to a fine, short needle, similar to the needle on an insulin syringe. Patients turn a dial to select the desired dose of insulin and press a plunger on the end to inject the insulin subcutaneously. Insulin pumps are the size of a deck of cards and are designed to be worn on a belt or carried in a pocket. The pump injects insulin into the subcutaneous tissue of the abdomen via an attached catheter. Pumps can be programmed to administer a bolus of short-acting insulin (typically, an insulin analogue) in response to meals. Pumps can also be programmed to deliver small amounts of insulin throughout the day to maintain a basal level of insulin.

Lowest cost option for administering insulin. Patients must measure volume of insulin to be injected at each administration. High dose-to-dose variability can lead to hypoglycemia.

Pen

Pump

Cartridges of insulin are more expensive than vials of insulin. Pens offer great convenience thanks to their small size and portability. Use of a dial to control the dose of insulin results in less dose-to-dose variability and, therefore, a lower risk of hypoglycemia.

Insulin pumps are the most expensive insulin delivery devices available. If adjusted properly, these pumps carry the lowest risk of hypoglycemia because they allow close control of insulin levels without multiple injections.

glargine group and 27.1% of the NPH insulin group. This trial demonstrated that insulin glargine is as efÞcacious as NPH insulin. A one-year Phase III study compared insulin glargine with NPH insulin in 426 type 2 diabetics who had poor glycemic control on oral agents (Yki-J¨arvinen H, 2000). Patients were allowed to continue taking their oral agents during the study. Researchers observed similar improvements in HbA1c levels in both treatment groups: HbA1c levels in the insulin glargine group dropped to an average of 8.34%; HbA1c levels in the NPH insulin group fell to an average of 8.24%. However, the investigators observed signiÞcantly lower blood glucose concentrations in the insulin glargine group compared with the NPH group, both before and after dinner. Therapy with insulin glargine has an incidence of hypoglycemia similar to or less than that of NPH insulin and a reduced incidence of nocturnal hypoglycemia

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compared with that of NPH insulin (Levien TL, 2002). Researchers conducted a 28-week Phase III study comparing insulin glargine with NPH insulin in 518 type 2 diabetes patients who were not taking oral agents and who had previously received basal insulin (with or without preprandial injections of regular insulin) (Rosenstock J, 2001). SigniÞcantly fewer patients experienced episodes of nocturnal hypoglycemia in the insulin glargine group (31.3%) than in the NPH group (40.2%). Insulin glargine patients maintained a 25% reduction in nocturnal hypoglycemia throughout the study. Weight gain was also less in the insulin glargine group compared with the NPH insulin group (0.4 kg and 1.4 kg, respectively). However, injection site pain was reported more frequently in the insulin glargine group. A retrospective analysis combining the results of a nine-month observational study and a 24-week, open-label, parallel study showed that the early introduction of insulin glargine in combination with existing oral antidiabetic therapy (with glimepiride and metformin) reduced HbA1c by approximately 1.5% with minimal weight gain (up to 1 kg) (Janka HU, 2005). Results also demonstrate that aggressive therapy with insulin and OAAs is more effective than treatment with insulin alone while offering equivalent safety. Glucagon-Like Peptide-1 Analogues Overview. The glucagon-like peptide-1 (GLP-1) analogues have received much attention from the diabetes community over the course of their development. The Þrst of its kind—exenatide (Amylin Pharmaceuticals/Eli Lilly’s Byetta)—was Þnally launched in June 2005. Preliminary evidence suggests that incretin hormones have a trophic effect on beta cells, resulting in proliferation and maturation of existing and new beta cells, respectively (Holst JJ, 2003). However, GLP-1 analogues do have signiÞcant drawbacks. For example, they are available only by injection. In addition, exogenous GLP-1 is susceptible to cleavage by dipeptidyl peptidase IV (DPP-IV), an enzyme in the bloodstream that renders the compound inactive. As a result, the early versions of GLP-1 analogues have only short half-lives and so require multiple daily doses. Mechanism of Action. GLP-1 is an intestinal peptide hormone produced by L cells of the small intestines; it acts upon multiple organs to induce glucoselowering. It plays the following roles: • • • •

Induces insulin secretion in a glucose-dependent manner. Controls gastric emptying. Inhibits appetite. Modulates islet secretion of glucagon and somatostatin, thereby reducing hepatic glucose production.

Exenatide. In April 2005, the FDA approved exenatide (Amylin Pharmaceuticals/Eli Lilly’s Byetta) as an adjunct therapy for type 2 diabetes. Exenatide is a 39-amino-acid GLP-1 analogue derived from the venom of the Gila monster

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lizard Heloderma suspectum, whose GLP-1 shares 50% homology with human GLP-1. According to Amylin, exenatide mimics the actions of GLP-1 by binding to the GLP-1 receptor, thus initiating the hormone’s biological effects. Because exenatide is resistant to degradation by DPP-IV, an enzyme present in the bloodstream, it has a longer half-life and duration of action (several hours) than recombinant human GLP-1. It is administered subcutaneously twice a day; an extended-release formulation is being developed in collaboration with Alkermes. Diabetic patients are thought to exhibit defective incretin signaling, which is characterized by reduced levels of active GLP-1 (amino acids 7–36) and impaired GLP-1 secretion in response to an oral glucose challenge. Impairments in the incretin response thus contribute to postprandial hyperglycemia. Administration of exogenous GLP-1 enhances the incretin response in type 2 diabetic patients. Three Phase III clinical trials of exenatide—known as the AC2993: Diabetes Management for Improving Glucose Outcomes (AMIGO) development program—have been completed in the United States. The Þrst trial, which investigated exenatide’s potential as monotherapy, began in December 2001 and was completed in August 2003. The second and third Phase III trials studied the efÞcacy of exenatide in combination with sulfonylureas and in combination with both sulfonylureas and metformin, respectively. Both trials were completed in November 2003, and data from all three trials are strikingly similar. In August 2003, Amylin and Eli Lilly reported data from the Þrst seven-month AMIGO study (Amylin, media release [electronic], August 6, 2003). Subjects (who were failing to achieve target HbA1c levels with metformin) received 10 mg exenatide twice daily as monotherapy. In patients receiving exenatide, mean HbA1c levels dropped from 8.6% to 7.2% by week 20 of the trial. The most common side effect was mild-to-moderate nausea, which resulted in six patients leaving the program. Results from the second AMIGO study were reported in early November 2003 (Amylin, media release [electronic], November 10, 2003). In this seven-month study, two-thirds of the 377 subjects received 10 mg of exenatide twice daily (after a one-month introductory course of 5 mg exenatide, twice a day) and continued ongoing treatment with sulfonylureas for the duration of the trial; the remainder received placebo with sulfonylureas. Of the subjects receiving exenatide, 41% experienced reductions in HbA1c levels to 7% or less and signiÞcant weight loss. Although severe hypoglycemia was not recorded, 36% of subjects receiving exenatide experienced mild-to-moderate hypoglycemia, compared with only 3% of the subjects receiving placebo. Nausea was the most frequent side effect and resulted in eight patients leaving the study. In late November 2003, Amylin and Eli Lilly reported positive results from the third of the AMIGO trials (Amylin, media release [electronic], November 24, 2003). The trial involved 734 type 2 diabetics who were failing to achieve target HbA1c levels with combinations of sulfonylureas and metformin. Patients were maintained on OAAs, but approximately two-thirds were randomized to receive exenatide, while the remainder received placebo (both arms remained on metformin and sulfonylureas). Subjects in the exenatide group received 5 mg

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subcutaneous doses of the agent (twice daily at breakfast and dinner) for a month, followed by a six-month phase with twice-daily doses of either 5 or 10 mg. Thirtyfour percent of patients receiving twice-daily administrations of 10 mg exenatide reduced their HbA1c levels to ≤7% and experienced an average reduction in body weight of 2 kg. HbA1c and weight data for patients in the placebo arm were not provided. There were no patient withdrawals due to hypoglycemia, and the most common side effect was nausea, which was experienced by 21% of the placebo arm, 39% of the 5 mg exenatide arm, and 49% of the 10 mg exenatide arm. Surprisingly, nausea accounted for less than 3% of subjects leaving the study. A one-year, Phase III, open-label trial observed similar glucose-lowering effects (Poon TH, 2003). The study examined the efÞcacy and safety of exenatide in combination with OAAs (i.e., metformin and sulfonylurea) in 150 patients with type 2 diabetes treated with the OAAs for at least three months prior to enrollment. Exenatide was injected subcutaneously, twice daily (10 mg), and OAAs were continued. After 12 weeks of treatment, patients exhibited an HbA1c reduction of 1.5% from baseline. Less than 5% of patients withdrew because of nausea. In an open-label, 82-week extension of Phase III clinical trials, 265 type 2 diabetics taking exenatide in combination with metformin or a sulfonylurea demonstrated sustained glycemic control (mean 1.2% reduction in HbA1c ) and weight loss (mean loss of 4.6 kg) (Kendall DM, 2005). Further analysis showed that those patients who experienced the greatest weight reduction experienced the greatest improvements in blood pressure and lipid proÞles. Adverse events, predominantly gastrointestinal in nature, were consistent with those observed during placebo-controlled trials. To investigate whether the glycemic control and weight reduction observed in the aforementioned trials are correlated with the gastrointestinal side effects of nausea and vomiting, researchers performed a post ad-hoc analysis using the data from the three randomized, placebo-controlled, 30-week clinical trials and a 52-week extension study (Maggs D, 2005). Statistical analysis showed that exenatide-induced HbA1c and weight reductions did not correlate with the severity of nausea or vomiting. One of the greatest unmet needs in type 2 diabetes treatment is for a diseasemodifying agent. Such an agent would likely become Þrst-line therapy and garner signiÞcant sales. Exenatide may be just that agent, but its injectable formulation and the current lack of long-term outcomes data are major hurdles to achieving this therapeutic role. A recent animal study corroborates Þndings from earlier research suggesting that exenatide may promote beta-cell neogenesis. Insulinresistant obese fa/fa rats treated with exenatide for six weeks experienced a 50% increase in beta-cell mass that was independent of body weight and glycemia (Nikoulina S, 2003). Furthermore, tremendous unmet need remains for agents that improve glycemic control without causing weight gain. Exenatide joins a small cadre of antidiabetic agents that can lower HbA1c while also causing a small amount of weight loss—both factors that will contribute to its success in the market.

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Amylin Analogues Overview. The role of amylin in diabetes has been under study for decades. Although many questions remain unanswered, amylin appears to work in concert with insulin as a key player in glucose regulation. Amylin Pharmaceuticals has developed pramlintide (Symlin), a synthetic human amylin analogue, for the treatment of type 2 diabetes. Mechanism of Action. Amylin is produced and secreted by the beta cells of the pancreas and, in concert with insulin, appears to be a key player in glucose management (Figure 23). The hormone slows glucose absorption from the gut into the blood, as well as postprandial glucose release from the liver. Although there have been conßicting reports relating to associations between type 2 diabetes and amylin levels, it is believed that a deÞciency of amylin may result in higher levels of postprandial glucose in the blood. Pramlintide. In March 2005, the FDA approved pramlintide (Amylin Pharmaceuticals’ Symlin) for combination use with insulin in the treatment of type 2 diabetics whose glucose levels remain uncontrolled on insulin alone. Pramlintide is also in Phase II development for obesity. The agent is an amylin analogue containing substitutions at positions 25, 28, and 29, which inhibit the otherwise problematic self-aggregation observed with native amylin. Development of pramlintide in Europe was suspended in January 2004 after the European Medicines Agency requested additional information not provided in the original Þling. The company is working to clarify the request and is actively seeking partnership for further development in Europe and Japan.

FIGURE 23. Mechanism of action of amylin analogues.

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To test the efÞcacy and tolerability of pramlintide and insulin compared with insulin alone, researchers performed a randomized, double-blind, parallel study involving 499 type 2 diabetics (Gottlieb A, 1999). Prior to randomization, all subjects were stabilized on insulin for two months and then underwent a onemonth placebo lead-in phase. Subjects receiving pramlintide were administered subcutaneous injections of 90 µg two or three times a day or 120 µg twice a day for 26 weeks (for a total of 180, 240, or 270 µg a day). The addition of pramlintide to insulin therapy signiÞcantly increased the number of subjects meeting “response” criteria (a reduction in HbA1c of ≥0.5% at four weeks); 47%, 49%, and 50% of subjects were responsive to total daily doses of 180, 240, and 270 µg, respectively. Subjects receiving insulin and 240 µg/day pramlintide experienced statistically signiÞcant reductions in HbA1c (0.4%) versus baseline (9.4%) compared with those receiving insulin alone (0.1%). Interestingly, the highest dose of pramlintide was no more effective according to this measure. Although the investigators state that the frequency of severe hypoglycemia in the pramlintide groups was not statistically different from that of the insulin and placebo group, they noted an increase in overall hypoglycemia in those receiving pramlintide of 7.0%, 6.0%, and 10% in the 180, 240, and 270 µg groups, respectively, compared with insulin and placebo (3.0%). In a multicenter, open-label study, 166 insulin-using type 2 diabetics were administered 120 µg of pramlintide two or three times a day, and their insulin use was subsequently adjusted to achieve standard glycemic targets (Karl DM, 2005). After the 26-week treatment period, patients exhibited an HbA1c reduction of 0.56%, weight loss of 2.8 kg, and short-acting insulin dose reduction of 10%. The most common adverse events were nausea (mild to moderate, 31%; severe, 2%) and vomiting (mild to moderate, 7%; severe, <1%). Pramlintide is also under Phase II development for obesity. A dose-ranging study of pramlintide in 204 obese patients (of whom 44 had type 2 diabetes) demonstrated that the agent elicited an average 3.5 kg weight loss after 16 weeks of treatment. Pramlintide may thus Þnd use in niche patient populations who require lower doses of insulin or who want to limit weight gain while on insulin therapy and do not mind additional daily injections. Emerging Therapies Development of compounds for the treatment of type 2 diabetes is highly competitive. As of this writing, more than 250 investigational compounds are in active preclinical and clinical development, and in 2005, more than 30 drug-licensing and collaboration deals took place among pharmaceutical and biotechnology companies in the United States, Europe, and Japan. Several factors are fueling this activity: •

There is an alarming increase in the number of individuals who are both physically inactive and either overweight or obese—conditions that are key risk factors for developing type 2 diabetes. In addition, type 2 diabetes preva-

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lence correlates strongly with advancing age, and the elderly populations in the seven major pharmaceutical markets are expected to increase. Physicians are increasingly turning to polypharmacy to treat type 2 diabetes by introducing multiple agents with complementary actions at the onset of drug treatment. Existing antidiabetic therapies have shortcomings in the areas of safety, tolerability, and long-term efÞcacy. Of particular concern are the lack of disease-modifying agents that can slow the progression of type 2 diabetes and the small number of weight-neutral agents (most antidiabetic agents cause weight gain). Researchers have a better understanding of the pathophysiology of type 2 diabetes.

Over the past ten years, the peroxisome proliferator-activated receptor (PPAR) and incretin pathways have been a major hub of research and development efforts, culminating in a late-stage pipeline dominated by PPAR agonists, GLP-1 analogues, and dipeptidyl peptidase (DPP-IV) inhibitors. In 2005, two products—pramlintide (Amylin Pharmaceuticals’ Symlin) and exenatide (Eli Lilly/Amylin’s Byetta)—emerged from this pipeline. Other compounds in latestage development are Þxed-dose combination agents that seek to improve patient compliance by combining two antidiabetic agents into one pill. The development of Þxed-dose combination agents has become a popular practice among many leading companies such as Bristol-Myers Squibb (BMS), GlaxoSmithKline (GSK), and Takeda to protect their diabetes franchises in the wake of impending patent expirations. Early-stage investigational drugs target an array of pathways and processes—glycogen metabolism (glycogen synthase kinase-3 inhibitors and glycogen phosphorylase inhibitors), insulin signaling (protein tyrosine phosphatase-1B [PTP-1B] inhibitors), glucose transport (sodium glucose cotransporter inhibitors), and hepatic lipid catabolism (11-beta hydroxysteroid dehydrogenase inhibitors). Because of the extensive number of investigational agents for type 2 diabetes and the lack of human clinical trial data, no early-stage drugs are discussed here. In the next ten years, physicians will have even more therapeutic options for their patients. Uptake of these drugs may be limited for the following reasons: •

•

•

These agents will likely be used as adjunctive therapy with the wellestablished, inexpensive Þrst-line agents metformin or the sulfonylureas because of general safety concerns with new therapies. General practitioners (GPs) and primary care physicians (PCPs) are generally more conservative than specialists in treating type 2 diabetes and so may be slower in incorporating the newer agents into their diabetes treatment regimens. Outcomes data are crucial for future, long-term uptake, but time and expense make it unfeasible to conduct these types of trials at the clinical development

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stage. Companies are instead performing these trials as postmarketing studies. Long-term safety data are needed. Most clinical trials for type 2 diabetes drugs have been 16-week, placebo-controlled trials, although more companies are performing 24- to 52-week-long studies to address safety issues.

Many of the clinical trials of emerging therapies mentioned in this section also report effects that pertain to dyslipidemia and metabolic syndrome. This section focuses on glucose-lowering efÞcacy, which is speciÞc to type 2 diabetes. Dual-Acting Peroxisome Proliferator-Activated Receptor Agonists Overview. Dual-acting PPAR agonists have afÞnity for both the alpha and gamma subtypes of PPAR. These agents not only stimulate PPAR-gamma, thereby improving glycemic control, but also activate PPAR-alpha, thereby favorably altering lipid proÞles. The latter effect may have signiÞcant implications in preventing the cardiovascular complications associated with type 2 diabetes. An effective dual-acting PPAR agonist with a side-effect proÞle that is comparable to, if not better than, that of currently available PPAR-gamma agonists would be a strong competitor to the currently marketed agents rosiglitazone (GSK’s Avandia) and pioglitazone (Takeda’s Actos). The development of several drugs in this class has been discontinued because of potentially serious side effects—such as ßuid retention with GSK’s GI262570 (GSK, press release, October 23, 2001), bladder tumors that were detected in preclinical trials with Novo Nordisk’s ragaglitazar (NN-622) (Novo Nordisk, press release, July 26, 2002), and rare malignant tumors found in mice receiving Merck’s MK-767 (Merck, media release [electronic], November 2003). These Þndings raised signiÞcant safety concerns and prompted the Executive Carcinogenicity Assessment Committee of the U.S. Center for Drug Evaluation and Research (CDER) to collect and review rodent data from 11 different PPAR agonists under development. The committee concluded that this class of agents is a “multispecies, multistrain, multisex, multisite carcinogen” (www.fda.gov, accessed June 29, 2005). As a result, the FDA and European regulatory bodies issued new guidance requiring two-year rodent carcinogenicity studies before Phase III (human) clinical trials lasting longer than six months can be conducted (www.fda.gov/cder/present/DIA2004/elhage Þles/frame.htm, accessed July 30, 2005). The latest casualty is Takeda’s TAK-559, which was discontinued in 2004 during Phase III testing because of abnormal liver function observed in a small number of patients during a review of interim trial results (Takeda, press release, March 30, 2005). Late-stage dual-acting PPAR agonists still under development include muraglitazar (Merck/BMS’ Pargluva), tesaglitazar (AstraZeneca’s Galida), and Eli Lilly/Ligand Pharmaceuticals’ naveglitazar. BMS and AstraZeneca have already completed the required two-year rodent carcinogenicity data. Ligand and Eli Lilly will complete the necessary studies later in 2005. Esai’s E-3030, Ono’s ONO-5129, SanoÞ-Aventis’ AVE-0847, and

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Plexxikon’s PLX-204. Compounds that target all three isoforms of PPAR (alpha, gamma, and delta), such as Perlegen/Mitsubishi Pharma’s netoglitazone, are in early investigation. These pan-PPAR agonists are not discussed here because they are in the early stages of development and no human clinical data are available yet. Mechanism of Action. Activation of PPAR-gamma increases glucose uptake by skeletal muscle and enhances adipocyte differentiation. PPAR-alpha receptors are expressed in skeletal muscle, liver, kidney, and vascular endothelial cells. Activation of PPAR-alpha affects fatty-acid oxidation and lipoprotein metabolism and increases cellular uptake of fatty acids (Ide T, 2000; Murakami K, 1998), as depicted in Figure 24. PPAR-alpha has also been implicated in the regulation of inßammatory responses in vascular endothelial cells and atherosclerosis (Loviscach M, 1999). Dual-acting PPAR agonists, therefore, function similarly to PPAR-gamma agonists by improving beta-cell function and lowering serum glucose levels but have the added effect of lowering serum lipid levels.

FIGURE 24. Effects of PPAR-alpha activation.

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Muraglitazar. BMS and Merck are developing muraglitazar (Pargluva)∗ , a dual activator of PPAR-alpha and PPAR-gamma. Due to recent news that muraglitazar may cause severe cardiovascular adverse events, Merck has withdrawn from this partnership and BMS is currently deliberating on the drug’s fate. Two-year carcinogenicity studies in rodents were done early in development and have been reviewed by the FDA. Tumor incidence in rodents was found either to be species-speciÞc or to occur at dosing levels 48 times higher than what was used in human studies (BMS company presentation, R&D review, November 17, 2004). Promising Phase III clinical data were presented at the 65th ScientiÞc Session of the American Diabetes Association (ADA) in June 2005 (Kendall DM, 2005[a]; Kendall DM, 2005[b]; Mohideen P, 2005). Results suggested that muraglitazar is able to induce glycemic control in patients who were uncontrolled on monotherapy with a sulfonylurea or metformin. Side effects included slightly elevated risk of hypoglycemia, weight gain, and edema (ßuid retention). Findings from a comparison trial with pioglitazone in type 2 diabetic patients who failed metformin monotherapy indicated that muraglitazar (at high doses) is more effective in lowering glucose levels than pioglitazone (at moderate doses). Although both agents have a similar side-effect proÞle, muraglitazar caused a slightly higher rate of edema and greater weight gain than pioglitazone. Edema is a concern with both the PPAR-gamma agonists and the dual-acting PPAR agonists because of the potential for congestive heart failure (CHF). Several cases of CHF were observed during clinical trials, but the overall number was small and mostly involved patients with a prior history of cardiovascular disease. All cases were resolved by either treatment with diuretics or discontinuation of therapy with muraglitazar. A double-blind, randomized, placebo-controlled, 24-week Phase III study involving 583 type 2 diabetic patients with hyperglycemia uncontrolled on sulfonylurea monotherapy demonstrated muraglitazar’s efÞcacy as a second-line therapy (Mohideen P, 2005). Patients were randomized into three arms: (1) high-dose muraglitazar (5 mg); (2) low-dose muraglitazar (2.5 mg); or (3) placebo once daily (in addition to existing glyburide therapy). Patients treated with muraglitazar exhibited reductions in HbA1c levels of 1.0% (low-dose group) and 1.2% (high-dose group), compared with an increase of 0.2% HbA1c in the placebo arm. However, a greater percentage of patients in the muraglitazar treatment arm experienced weight gain and hypoglycemia compared with those treated with glyburide and placebo: 3% (low dose) and 7% (high dose) exhibited weight gain versus 2% in the glyburide/placebo arm; 15% (low dose) and 19% (high dose) experienced hypoglycemia versus 8% in the glyburide/placebo arm. The frequency of edema was also slightly elevated in the muraglitazar arm (9% in the low-dose group, 10% in the high-dose group, compared with 7% in the glyburide/placebo group). The 5 mg dose of muraglitazar is therefore associated with a higher level of side effects but only slightly better efÞcacy in reducing HbA1c levels. In a 24-week, double-blind, randomized comparator trial with pioglitazone, 1,159 patients who remained uncontrolled on metformin monotherapy were

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randomized to once-daily muraglitazar (5 mg) or once-daily pioglitazone (30 mg) (Rubin CJ, 2005). During their presentation at the 65th ScientiÞc Sessions of the ADA in June 2005, the investigators noted that the 30 mg pioglitazone dose was chosen rather than the 45 mg dose because it is the clinically approved dosage for use in combination with metformin and is the most commonly used dosage. HbA1c reduction was more pronounced with 5 mg muraglitazar than with 30 mg pioglitazone (1.14% versus 0.85% reduction, respectively), although side effects of edema (9.2% versus 7.2%) and weight gain (1.4 kg versus 0.6 kg) were slightly more elevated with muraglitazar than with pioglitazone. Muraglitazar was also shown to be effective in lowering HbA1c levels in a greater proportion of patients compared with pioglitazone. Additionally, patients treated with 5 mg muraglitazar exhibited a 28% reduction in triglyceride (TG) levels and a 19% increase in highdensity lipoprotein (HDL) cholesterol. In comparison, patients receiving 30 mg pioglitazone exhibited a 14% reduction in TG and a 14% increase in HDL levels. Also at the 65th ScientiÞc Sessions of the ADA in June 2005, long-term clinical trial data for muraglitazar were presented during the Late-Breaking Clinical Trial Symposium (Kendall DM, 2005[b]). The Phase III pioglitazone comparator study continued with a 26-week extension, and preliminary analyses indicate that the differences in glycemic control and lipid-lowering efÞcacy between muraglitazar 5 mg and pioglitazone 30 mg were maintained after 50 weeks—muraglitazar-induced HbA1c reductions of 1.12% versus 0.74% for pioglitazone. The overall long-term proÞle of adverse events was similar to that observed in the short-term study. Four additional deaths were reported for patients on muraglitazar (stroke, myocardial infarction, sudden cardiac death, and previously diagnosed pancreatic cancer). Although details were not provided, investigators reported that the deaths were not drug-related. Three cases of CHF occurred in the muraglitazar group compared with one case in the pioglitazone group. At the same session, data from a Phase II study were presented. The randomized, double-blind, dose-ranging trial compared the efÞcacy of muraglitazar and pioglitazone in drug-naive type 2 diabetes patients inadequately controlled with diet and exercise (BMS and Merck, press release, June 12, 2005). The 1,477 patients were randomized to one of Þve muraglitazar doses (0.5 mg, 1.5 mg, 5 mg, 10 mg, or 20 mg) or pioglitazone 15 mg. Patients treated with muraglitazar exhibited dose-dependent reductions in HbA1c ranging from 0.57% to 1.76%, compared with a reduction of 0.57% in patients treated with 15 mg pioglitazone. After the initial 24-week dose-ranging study, patients continued in an ongoing long-term extension phase with their current dose of study medication except for two exceptions: patients who received muraglitazar 0.5 mg in the shortterm phase were given a dose increase to 1.5 mg and patients who received muraglitazar 20 mg in the short-term phase were given a lower dose of 10 mg (BMS and Merck, press release, June 12, 2005). Patients who were maintained on one dose (5 mg) without the need for titration throughout the 104-week study experienced a 1.52% reduction in HbA1c . Glycemic results for pioglitazone during the extension phase of the study are not available. However, results of

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changes to lipid parameters are available for both groups. Patients treated with 5 mg muraglitazar exhibited a 22% reduction in TG and a 29% increase in HDL levels, compared with a 12% reduction in TG and an 18% increase in HDL levels in patients treated with 15 mg pioglitazone. Muraglitazar was associated with a slightly lower frequency of edema than pioglitazone: 25% of patients in the 5 mg muraglitazar group experienced edema compared with 30% of patients in the 15 mg pioglitazone group. However, muraglitazar caused signiÞcantly more weight gain than pioglitazone: the average amount of weight gained by patients taking 5 mg muraglitazar for 104 weeks was 5.9 kg, compared with a gain of 1.9 kg in patients taking 15 mg pioglitazone. Tesaglitazar. AstraZeneca is seeking to enter the type 2 diabetes market with its Þrst antidiabetic agent, tesaglitazar (Galida)∗ , an activator of both PPARalpha and -gamma subtypes. Tesaglitazar has completed carcinogenicity studies in rodents and has been in Phase III development in the United States since 2003. AstraZeneca is also developing this compound as a treatment for metabolic syndrome. Clinical trial data for tesaglitazar were lacking until the presentation of Phase II results at the 65th ScientiÞc Sessions of the ADA in June 2005. The randomized, double-blind, placebo-controlled, dose-ranging trial involved 418 patients with type 2 diabetes (Goldstein BJ, 2005). Subjects were randomized into the following treatment arms: placebo; tesaglitazar (0.1, 0.5, 1.0, 2.0, or 3.0 mg); or 45 mg pioglitazone once daily for 12 weeks. Fasting plasma glucose (FPG) was reduced in a dose-dependent manner, by 30 mg/dL in the 0.5 mg/day tesaglitazar group up to 61 mg/dL in the group receiving 3 mg/day. Patients treated with 45 mg/day pioglitazone experienced an average FPG reduction of 38.5 mg/dL. Secondary end points in this study included changes in TG and HDL and were observed to occur in a dose-dependent manner: patients treated with tesaglitazar exhibited 5–41% reduced TG levels and 1–13% increased HDL levels. In comparison, patients treated with 45 mg/day pioglitazone experienced an 8% reduction in TG levels and a 6% increase in HDL. Side effects increased in frequency in a dosedependent manner. Edema occurred in 4.2–6.8% of tesaglitazar-treated patients, compared with 4.2% in the pioglitazone group and 3% in the placebo arm. No cases of CHF were reported. Naveglitazar. Naveglitazar∗ is a dual-acting PPAR agonist developed by Ligand Pharmaceuticals and Eli Lilly. It is in Phase II/III development in the United States. Its development was delayed for 18–24 months pending completion of two-year rodent carcinogenicity data, as requested by the FDA. Few data are available for naveglitazar, but some promising early Phase II data were presented at the 64th ScientiÞc Sessions of the ADA in June 2004 (Prince M, 2004). The randomized, double-blind, six-arm parallel study was conducted in 151 patients with type 2 diabetes. Patients continued their existing therapy (i.e., diet and exercise alone or monotherapy with metformin or a sulfonylurea) and were treated once daily for 12 weeks with placebo, 8 mg rosiglitazone, or naveglitazar (0.04, 0.2, 0.8, or 1.2 mg). Results showed that 0.8 and 1.2 mg/day

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naveglitazar reduced HbA1c in a dose-dependent manner (by 1.01% and 1.20%, respectively). In comparison, patients taking 8 mg/day rosiglitazone saw a 0.79% reduction in HbA1c . However, the study was not sufÞciently powered to provide statistically signiÞcant results for rosiglitazone. Secondary end points included TG and HDL levels. Patients in the 0.8 and 1.2 mg/day naveglitazar groups saw 60 mg/dL and 87 mg/dL reductions in TG levels, respectively, compared with placebo, and increases in HDL ranging from 9.0% to 10.7%. Naveglitazar was generally well tolerated and did not cause statistically signiÞcant weight gain over the course of the 12-week study. Peroxisome Proliferator-Activated Receptor-Gamma Agonists Overview. The key commercial opportunity that remains for an investigational PPAR-gamma agonist is to improve upon the safety proÞle of the existing PPARgamma agonists rosiglitazone and pioglitazone. These agents’ two major drawbacks are their propensity to cause weight gain and edema in patients. To tackle this issue, Metabolex is developing metaglidasen, a selective PPAR modulator (SPPARM) that does not act on adipose tissue—the main site from which the aforementioned side effects originate. Other companies that have PPAR-gamma agonists in early-stage development include Calyx’s CLX-092, Sankyo’s rivoglitazone (CS-011), and Amgen’s AMG-131 (a SPPARM). These agents are not discussed in this section because human clinical data are not yet available. Because several high-proÞle PPAR-gamma agonists have been discontinued in Phase III over the past Þve years as a result of tumor Þndings in rodents, the CDER Executive Carcinogenicity Assessment Committee collected and reviewed rodent data from 11 PPAR agonists (-gamma and dual-acting) and concluded that this class of agents is a cross-species, multi-tissue carcinogen (www.fda.gov, accessed June 29, 2005). As stated earlier, the FDA now requires two-year rodent carcinogenicity studies before Phase III clinical trials lasting longer than six months in humans can be conducted. As a result, the development of all PPARgamma agonists will be delayed pending completion of rodent carcinogenicity studies. Mechanism of Action. The gamma isotype of the PPAR receptor is found in the nucleus of key target tissues for insulin action, such as skeletal muscle, adipose tissue, and the liver. PPAR-gamma agonists improve glycemic control by enhancing insulin sensitivity, as illustrated in Figure 8. In the presence of endogenous or exogenous insulin, PPAR-gamma agonists increase glucose uptake and utilization in skeletal muscle, increase glucose uptake and reduce fatty-acid output in adipose tissue, and reduce gluconeogenesis, glucose output, and triglyceride synthesis in the liver. Researchers also believe that PPAR-gamma agonists downregulate resistin, which is expressed in diabetic adipocytes and inhibits insulin action in skeletal muscle. In vitro studies have shown that PPAR-gamma agonists trigger an increase in the expression of the GLUT4 glucose transporter on the cell-surface of different tissues (Hauner H, 2002). In addition, studies have

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demonstrated anti-inßammatory and antiapoptotic effects that may limit the progression to cardiovascular complications of diabetes (Artwohl M, 2005; Buffon A, 2005). Metaglidasen. Metaglidasen∗ , also known as MBX-102, is a nonthiazolidinedione, selective PPAR modulator (SPPARM) in Phase II development by Metabolex. The agent is chemically related to halofenate, which was in development in the 1970s as a uric acid treatment and lipid-lowering agent. Halofenate comprises two enantiomers, one of which was associated with gastrointestinal side effects caused by the inhibition of cyclooxygenase-1 (COX-1). The other enantiomer—reformulated as metaglidasen—is thought to be responsible for halofenate’s glucose- and lipid-lowering effects. SPPARMs have a mechanism of action similar to that of the PPAR-gamma agonists and function by modulating genes needed for insulin sensitization, glucose metabolism, lipid metabolism, and inßammation. Metabolex has developed an analogue of metaglidasen designated MBX-2044, which recently completed Phase I trials, and claims that this agent exhibits greater potency than that of metaglidasen. Meanwhile, plans for Phase III trials of metaglidasen are under way following promising Phase II results. The most recent Phase II trial results demonstrate that metaglidasen (in combination with existing insulin therapy) offers good efÞcacy with less weight gain compared with insulin therapy alone (Rosenstock J, 2005). The randomized, double-blind, placebo-controlled, 12-week study involved 217 patients with type 2 diabetes whose blood glucose levels were inadequately controlled on insulin monotherapy. This population is considered to be at greater risk for edema and weight gain from the use of PPAR-gamma agonists, compared with those using PPAR-gamma agonists with other oral antidiabetic agents. All patients continued on their insulin therapy and were randomized into one of three treatment arms: 200 mg/day metaglidasen, 400 mg/day metaglidasen, or placebo. The highest dose of metaglidasen demonstrated the greatest efÞcacy, and overall side effects were slightly less frequent than observed in the active control group (insulin plus placebo): • • •

•

400 mg metaglidasen (plus insulin) reduced HbA1c by 1% from baseline, compared with 0.3% in the active control group (placebo plus insulin). 400 mg metaglidasen lowered TGs by 38% compared with placebo. Weight gain in the two insulin plus metaglidasen groups ranged from 0.3 kg to 0.7 kg compared with 0.8 kg gained by patients in the insulin plus placebo group. 11% of patients in the 200 mg metaglidasen arm developed edema compared with 5.8% in the 400 mg metaglidasen arm and 16% in the placebo arm.

Fixed-Dose Combinations Overview. As type 2 diabetes progresses, patients usually require more than one medication to maintain glycemic control. Clinical evidence increasingly indicates that combination therapy at the onset of drug treatment may be more effective

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than stepwise regimens (Table 11). For patients who require therapy with more than one agent, single-pill, Þxed-dose combinations (FDCs) provide a convenient alternative to taking two separate pills. GSK is developing its second FDC, Avandaryl, which is a combination of the PPAR-gamma agonist rosiglitazone (GSK’s Avandia) and the sulfonylurea glimepiride (SanoÞ-Aventis’s Amaryl). Takeda is introducing its Þrst FDC, Actoplus Met, a combination of pioglitazone (Takeda/Eli Lilly’s Actos) and metformin, and it is following up with an FDC of pioglitazone and glimepiride (SanoÞ-Aventis’s Amaryl). Although these new formulations offer more-convenient dosing regimens, the availability of generic versions of the traditional Þrst-line therapies, metformin and sulfonylureas, will make it difÞcult to introduce FDCs at an earlier point during the course of therapy. Also hampering the uptake of FDCs as Þrst-line therapy is the need for dose titration, which cannot be effectively done with FDCs. Mechanism of Action. FDCs are single-pill agents that combine therapies with complementary therapeutic effects. FDCs under development include PPARgamma agonists in combination with either metformin or a sulfonylurea, two of the most commonly used oral antidiabetic agents. The mechanism of action of metformin has not yet been fully elucidated, but investigators postulate that it improves insulin sensitivity and reduces hepatic glucose production. Sulfonylureas reduce hyperglycemia by inducing insulin secretion. (For more information about the mechanisms of action of the individual agents found in FDCs, see “Current Therapies.”) Pioglitazone/Metformin. Pioglitazone/metformin (Actoplus Met) will be Takeda’s Þrst FDC in the type 2 diabetes market; the agent was approved by the FDA in August 2005 (Takeda, press release, August 30, 2005). In February 2004, the company Þled a marketing authorization application (MAA) in Europe. Actoplus Met is a twice-daily formulation. In January 2004, Takeda and Andrx entered into an agreement to develop a once-daily formulation that will combine pioglitazone (Figure 10) with Andrx’s extended-release formulation of metformin (Fortamet) (Figure 12). To date, no data have been published evaluating the efÞcacy and safety of Takeda’s twice-daily FDC. However, studies have evaluated the combination regimen of pioglitazone and metformin as separate pills. In a study of 328 metformin-treated patients, adding pioglitazone (30 mg once daily) signiÞcantly reduced mean HbA1c levels by 0.8% and mean FPG levels by 38 mg/dL compared with metformin and placebo (Einhorn D, 2000). The addition of pioglitazone also signiÞcantly reduced TGs by 18.2% and increased HDL by 8.7%, and investigators observed a statistically nonsigniÞcant downward trend in low-density lipoprotein (LDL) levels. Pioglitazone/Glimepiride. In June 2005, Takeda submitted an NDA for an FDC of pioglitazone (Figure 10) and glimepiride (Figure 14). To date, no data have been published evaluating the efÞcacy and safety of this FDC. However, a 12-month, double-blind, randomized, parallel-group study comparing the effects

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of pioglitazone and rosiglitazone in patients treated with glimepiride demonstrated that combination therapy with either PPAR-gamma agonist improves HbA1c by 1.3% after 12 months of therapy (Derosa G, 2005). Both PPAR-gamma agonists and sulfonylureas are known to cause considerable weight gain. In fact, this study showed that patients treated with either combination of pioglitazone/glimepiride or rosiglitazone/glimepiride experienced an average increase of 4.9% and 6.2%, respectively, in their body mass index (BMI) after one year of treatment. The pioglitazone/glimepiride FDC will therefore face considerable competition from FDCs containing PPAR-gamma agonists and metformin, which are not associated with as signiÞcant an increase in weight. Rosiglitazone/Glimepiride. In October 2003, GSK submitted Avandaryl, an FDC of rosiglitazone (GSK’s Avandia) (Figure 9) and glimepiride (SanoÞAventis’s Amaryl) (Figure 14), for FDA approval. In August 2004, the company announced that the FDA had issued an approvable letter for the FDC pending resolution of manufacturing issues for rosiglitazone in GSK’s manufacturing plant in Puerto Rico. The drug was launched in February 2006 after securing Þnal regulatory approval. SanoÞ-Aventis will receive a portion of the proÞts from U.S. sales, but the company will not be copromoting the agent in the United States. Clinical trials evaluating the efÞcacy of the rosiglitazone/glimepiride FDC have not been published. Therefore, this section reviews data from trials in which rosiglitazone and glimepiride were administered as separate agents. One study found that the introduction of rosiglitazone to existing glimepiride therapy induced better glycemic control than increasing the dose of glimepiride monotherapy (Rosenstock J, 2005). In the 24-week, randomized, double-blind, placebo-controlled study, 361 type 2 diabetic patients were treated with 2 mg glimepiride for six weeks and then were randomized to one of two treatment arms: 4 mg rosiglitazone + 2 mg glimepiride (submaximal dose) or placebo + 4 mg glimepiride (maximum dose), which was titrated to 8 mg if FPG exceeded 110 mg/dL. The rosiglitazone-treated group exhibited statistically signiÞcant HbA1c reductions of 0.68% from baseline compared with 0.08% with high-dose glimepiride alone. Glucagon-Like Peptide-1 Analogues Overview. In 2005, the highly anticipated GLP-1 analogue exenatide (Amylin Pharmaceuticals/Eli Lilly’s Byetta) entered the U.S. market, becoming the Þrst agent of its class and the Þrst approved agent that modulates the incretin pathway. Several GLP-1 compounds are in clinical development. Leading the pipeline are Alkermes/Amylin/Lilly’s exenatide LAR and Novo Nordisk’s liraglutide. Earlystage GLP-1 analogues include Ipsen’s BIM-51077, Theratechnologies’ TH-0318, and Suntory’s SUN-E7001. This section does not discuss the early-stage analogues because no human clinical data are yet available. GLP-1 analogues reduce hyperglycemia via multiple mechanisms, and preliminary evidence suggests that they have a trophic effect on beta cells, resulting in

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proliferation and maturation of existing and new beta cells, respectively (Holst JJ, 2002). However, current GLP-1 analogues have signiÞcant drawbacks. First, they are available only by injection. Second, exogenous GLP-1 is susceptible to cleavage by dipeptidyl peptidase IV (DPP-IV), an enzyme in the bloodstream that renders the compound inactive. As a result, early versions of GLP-1 analogues have short half-lives and require high doses injected several times daily. Third, the side effect of nausea affects a signiÞcant percentage of patients, although many patients report that it subsides after several weeks of therapy. Agents in late-stage development are chemically modiÞed to have extended half-lives and so can be injected less frequently than exenatide. Mechanism of Action. GLP-1 is an intestinal peptide hormone produced by L cells of the small intestines. It plays the following roles: • • •

It induces insulin secretion in a glucose-dependent manner (and so is associated with a lower risk of hypoglycemia). It slows gastric emptying and suppresses appetite. It modulates islet secretion of glucagon and somatostatin, thereby reducing hepatic glucose production.

Diabetic patients are thought to exhibit reduced levels of active GLP-1 (amino acids 7–36) and impaired GLP-1 response to a glucose load, Þndings that suggest this impairment may contribute to postprandial hyperglycemia as a result of defective activity of the incretin pathway. Exogenous GLP-1 analogues substitute for the lack of functional endogenous GLP-1 or enhance the effects of GLP-1 in type 2 diabetic patients. Exenatide LAR. In collaboration with Alkermes, Amylin Pharmaceuticals and Eli Lilly are developing an extended-release formulation of exenatide, known as exenatide LAR (AC-2993 LAR) *, which is in Phase II trials in the United States and Phase I in Europe. A multidose Phase II trial using a once-weekly regimen began in the Þrst quarter of 2005 (Amylin, 2004 annual report). Exenatide LAR uses Alkermes’s Medisorb injectable sustained-release drug delivery system. In contrast to the two- to three-times daily formulation of exenatide, LAR is administered once a week. The technology has the potential to reduce administration to once a month. Preliminary results from the ongoing Phase II study indicate that exenatide LAR is safe and effective (Amylin, press release, August 22, 2005). The doubleblind, placebo-controlled study enrolled 45 patients who were unable to achieve glycemic control using metformin or lifestyle modiÞcations (average HbA1c ≈ 8.5). Patients were randomized to receive one of two once-weekly dosing regiments (low-dose or high-dose) or placebo. A signiÞcant number of patients receiving the high dose of exenatide LAR (12 of 14) had HbA1c < 7.0 after 15 weeks of treatment. In comparison, no patients in the placebo arm achieved HbA1c levels <7.0 during this period. Additionally, patients in the high-dose treatment arm lost an average of 9 pounds; weight-loss data is available for

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patients in the placebo arm was not available. Nausea was the most common adverse event, affecting 20% of patients in the high-dose group. In comparison, only 7% of patients in the placebo group experienced nausea. No incidents of severe hypoglycemia or gastrointestinal discomfort were reported in this study. Other data are available only from small-scale safety trials. A single-dose study involved 60 type 2 diabetic patients whose blood glucose levels were poorly controlled with lifestyle modiÞcations or metformin (Amylin Pharmaceuticals, press release, November 2, 2004). Patients in this study were randomized to one injection of exenatide LAR (four concentrations) or placebo and followed for 90 days. No dose-limiting side effects were observed. These results served as an impetus to start the multidose trial. A Phase I study found exenatide LAR to be well tolerated by type 2 diabetic patients. Researchers demonstrated that 23-hour continuous subcutaneous infusions of exenatide LAR, over the course of ten days, reduced preprandial, postprandial, and fasting glucose levels (Taylor K, 2002). Liraglutide. Under license from Scios, Novo Nordisk is developing a oncedaily subcutaneous analogue of GLP-1 called liraglutide∗ , which was previously known as insulinotropin or NN-2211. Liraglutide is in Phase II development in the United States, Europe, and Japan. Boston’s Massachusetts General Hospital, holder of the patent for insulinotropin, licensed the agent to Scios in 1988. Scios, in turn, licensed the agent to PÞzer. Scios’s agreement with PÞzer was terminated in 1996, and the agent was subsequently licensed to Novo Nordisk. Similar to the native GLP-1 peptide, liraglutide stimulates insulin secretion in a glucose-dependent manner. However, liraglutide has a signiÞcantly longer duration of action than that of GLP-1 because of chemical modiÞcations rendering the agent resistant to cleavage by DPP-IV (Juhl CB, 2002). Once-daily liraglutide has been shown to increase beta-cell sensitivity as measured by elevated levels of insulin and C-peptide nine hours after administration in type 2 diabetics (Chang AM, 2003). Preliminary in vitro studies in human pancreatic islet cells suggest that liraglutide’s effects may be regulated through the transforming growth factor (TGF)-beta pathway (Perfetti P, 2005). TGF-beta is thought to be a key regulator of islet-cell regeneration (Luo X, 2005). Other preclinical studies of liraglutide suggest that it may prevent free fatty-acid (FFA)-induced apoptosis in primary beta cells, thus contributing to the preservation of the beta cells and potentially delaying the onset and progression of type 2 diabetes (Bregenholt S, 2001). Data are limited at this time, but one study found that liraglutide’s efÞcacy is comparable to that of glimepiride and that liraglutide is relatively safe and well tolerated (Madsbad S, 2004). The 12-week, double-blind, placebo-controlled study with an open-label comparator (glimepiride) was conducted in 193 type 2 diabetic patients. Patients were randomized to the following treatment groups: variable dosages of liraglutide (0.045, 0.225, 0.45, 0.6, or 0.75 mg/day), placebo, or glimepiride (1–4 mg/day). Placebo-adjusted HbA1c changes from baseline in each liraglutide-treated group ranged from an increase of 0.2% in the lowestdose group to a reduction of 0.75% in the highest-dose group. The latter group

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achieved the same level of HbA1c reduction as glimepiride-treated patients who exhibited an average reduction of 0.74% HbA1c . Results also showed that 59% of patients in the two highest-dose groups (0.6 and 0.75 mg) achieved HbA1c ≤ 7% (mean baseline HbA1c was 7.6%). Liraglutide did not cause weight gain. In fact, the 0.45 mg liraglutide treatment induced a statistically signiÞcant weight loss of 1.2 kg. The incidence of nausea increased with dose: 10 out of 135 patients treated with liraglutide were affected by nausea compared with 1 out of 29 patients in the placebo arm. Other gastrointestinal side effects were vomiting, diarrhea, and constipation—most of which were resolved in three days. No antibody formation was detected. Liraglutide is given in a once-daily injection, an advantage over the multiple daily injections of the currently marketed GLP-1 analogue, exenatide (Lilly/Amylin’s Byetta). CJC-1131. CJC-1131 is an injectable GLP-1 analogue developed using ConjuChem’s Drug AfÞnity Complex (DAC) bioconjugation technology. The compound is in Phase II development in the United States and Europe. DAC extends the half-life of the GLP-1 peptide up to ten days in humans, a signiÞcant increase compared with native GLP-1, which has a half-life of less than Þve minutes. The DAC construct is composed of albumin, the carrier protein that enables the longer half-life, and a connector that permanently attaches the peptide to the albumin molecule. Preliminary data from primate studies show that CJC-1131 is not immunogenic (Wen SY, 2005). A Phase II study demonstrated that CJC-1131 reduces HbA1c more than metformin monotherapy in type 2 diabetes patients (Ratner RE, 2005). In this 12week, double-blind, placebo-controlled study, 81 type 2 diabetes patients were initially treated with metformin during the four-week baseline period. Patients were subsequently randomized to three treatment arms while continuing their metformin therapy: once-daily low-dose CJC-1131 (2.1 µg/kg); once-daily highdose CJC-1131 (2.6 µg/kg); or placebo (metformin only). CJC-1131 lowered HbA1c by 1.1% (high) and 0.6% (low) compared with placebo. In addition, 58% of patients whose baseline HbA1c was higher than 7% reduced their HbA1c to less than 7%. Weight loss was observed in all three groups but CJC-1131 led to an additional reduction of 2.5 kg (high-dose arm) and 1.6 kg (low-dose arm) over metformin. Gastrointestinal discomfort was the most common adverse event noted. SpeciÞcally, 25% of patients experienced moderate nausea during the Þrst four weeks, but the incidence of nausea subsided as the study continued. AVE-0010. SanoÞ-Aventis is developing AVE-0010 (ZP-10)∗ , a GLP-1 receptor agonist, which is in Phase II development in the United States. In June 2003, Aventis signed a licensing agreement with Zealand Pharma for the development and worldwide commercialization of AVE-0010. An early Phase I study showed that AVE-0010 was safe and well tolerated (Zealand Pharma, press release, September 18, 2002). This double-blind, placebocontrolled study assessed the safety, tolerability, and efÞcacy of AVE-0010 in 28 subjects with type 2 diabetes. Each subject received a single dose of AVE-0010

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(ranging from 1 to 40 µg) or placebo and was given a glucose-loaded meal one hour after dosing. No safety issues were identiÞed during the study, and the data indicated that a maximum of 20 µg of AVE-0010 was tolerated as a single dose. Investigators observed a dose-dependent lowering of plasma glucose concentrations following AVE-0010 administration compared with placebo. A 28-day, 64-patient, placebo-controlled Phase IIa study in which AVE-0010 was administered once or twice daily to type 2 diabetic patients (in addition to existing therapy) demonstrated a dose-dependent improvement in postprandial blood glucose compared with placebo added to existing therapy (SanoÞ-Aventis, press release, March 3, 2005). AVE-0010 appeared to be well tolerated. The incidence of nausea and vomiting was similar in treated and placebo groups. Dipeptidyl Peptidase IV Inhibitors Overview. Agents that target the incretin pathway are now taking center stage as the next innovation in the treatment of type 2 diabetes. Exenatide, a GLP1 analogue and the Þrst agent to target the incretin pathway, was launched in the United States in 2005. Another high-proÞle class of agents that targets this pathway are the DPP-IV inhibitors. Available in oral formulations, the DPP-IV inhibitors are more convenient than the GLP-1 analogues, which are available only in injectable formulations. The lead DPP-IV inhibitors in the type 2 diabetes pipeline are Novartis’ vildagliptin (LAF-237) and Merck’s sitagliptin (MK-431), both of which are in Phase III development. Probiodrug’s PSN-9301, and BMS’s saxagliptin. GSK has three DPP-IV inhibitors in its pipeline—823093, 815541, and 825964. The lack of data for these early-stage compounds precludes a detailed discussion of them. Mechanism of Action. DPP-IV cleaves and inactivates GLP-1, a peptide involved in increasing insulin secretion in a glucose-dependent manner. As a result, DPP-IV inhibitors potentiate endogenous GLP-1 activity by preventing the cleavage and inactivation of GLP-1. Vildagliptin. Novartis is developing vildagliptin (formerly LAF-237)∗ , an oral, long-acting DPP-IV inhibitor that is administered once daily (Ahren B, 2003). This compound entered Phase III trials for type 2 diabetes in the Þrst half of 2004. Inhibition of DPP-IV increases basal and postprandial GLP-1 levels, an action that inhibits glucagon secretion, culminating in improved glycemic control (Ahren B, 2005). Rodent data also demonstrate that vildagliptin increases pancreatic-cell neogenesis and reduces apoptosis in neonatal rats (a model for rapid beta-cell turnover and growth) (Duttaroy A, 2005). One-year efÞcacy data from an extension study are available for vildagliptin, but the number of patients participating in the study is relatively small. Results demonstrate that the addition of vildagliptin to metformin therapy can further reduce HbA1c , compared with treatment with metformin alone. This result was based on a 12-week, randomized, double-blind, placebo-controlled study that was extended 40 weeks to measure one-year efÞcacy (Ahren B, 2004). At the start

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of the trial, 107 type 2 diabetic patients who continued taking their metformin therapy participated in the Þrst 12 weeks. Seventy-one patients who completed the 12-week study consented to participate for an additional 40-week extension of the study. In the 12-week efÞcacy trial, the vildagliptin group exhibited a 0.6% reduction of HbA1c from baseline, compared with an increase of 0.1% in the placebo group (metformin only). In the extension population, treated patients experienced a placebo-adjusted HbA1c reduction of 1.1%. In addition, 42% of vildagliptin-treated patients achieved an HbA1c < 7%, compared with only 11% in the placebo group. Safety and tolerability did not differ between the treated and placebo groups. In the 12-week study, 52% of treated patients reported an adverse event compared with 55% in the placebo arm. In the longterm study, 69% of treated patients experienced an adverse event compared with 59% in the placebo arm. No hypoglycemic episodes occurred in the extension study. A small-scale study investigated pharmacokinetic interactions between vildagliptin and the PPAR-gamma agonist pioglitazone (Serra DB, 2005). Twelve type 2 diabetic patients were maintained on 45 mg/day pioglitazone for eight weeks and then randomized to one of two treatment arms: 100 mg/day vildagliptin plus pioglitazone, or placebo plus pioglitazone. Patients were maintained on combination therapy for 21 days before the 7-day open-label phase of the study, during which all patients were treated with 100 mg/day vildagliptin alone. Results of the study showed that vildagliptin was safe and well tolerated in combination with pioglitazone. Coadministration of vildagliptin and pioglitazone reduced average FPG levels, compared with treatment with pioglitazone and placebo. Vildagliptin appears to be well tolerated and safe as monotherapy, according to a study reported at the 64th ADA meeting in June 2004. The investigators calculated that the percentage of drug-related adverse events in subjects receiving vildagliptin or placebo was 16% and 11%, respectively (Pratley R, 2004). Another study demonstrated that vildagliptin was safe and well tolerated as an add-on therapy to metformin (Ahren B, 2004). In the study, 5.5% of subjects receiving vildagliptin and 4.0% of subjects receiving placebo experienced mild, drug-related adverse events. No serious adverse events were reported. In a placebo-controlled, double-blind trial, 40 type 2 diabetic patients whose blood glucose levels were controlled by diet alone were randomized to 100 mg/day vildagliptin or placebo (Ahren B, 2003). Results indicated that patients receiving vildagliptin experienced elevated postprandial GLP-1 levels and reductions in FPG and postprandial glucose levels. In addition, insulin levels remained unaffected throughout the study despite demonstrable reductions in glucose levels, which indicates that insulin sensitivity improved in patients treated with vildagliptin. Subjects tolerated vildagliptin relatively well; 12 patients receiving the drug experienced adverse events compared with 9 in the placebo group. Vildagliptin will be the Þrst agent within the highly anticipated DPP-IV inhibitor class to reach the market. As an orally available agent targeting the GLP1 pathway, vildagliptin offers a signiÞcant advantage over the injectable GLP-1 analogues. Clinical trials indicate that vildagliptin has a favorable side-effect

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proÞle, does not cause weight gain, and offers a level of efÞcacy comparable to that of existing oral antidiabetic agents. Sitagliptin. Sitagliptin (MK-431)∗ is a twice-daily oral DPP-IV inhibitor in development by Merck. It entered Phase III trials in the second half of 2004 in the United States. In November 2003, Merck subsidiary Banyu Pharma entered into an agreement with Ono Pharmaceuticals to codevelop and comarket sitagliptin in Japan, where the agent is in Phase II trials. A randomized, double-blind, 12-week study involving 743 type 2 diabetic patients demonstrated that sitagliptin can reduce HbA1c levels in a dose-dependent manner. Patients were randomized to the following treatment arms: placebo; twice-daily sitagliptin (5, 12.5, 25, or 50 mg); or glipizide (5 mg, uptitrated if necessary) (Scott R, 2005). Placebo-adjusted HbA1c reductions in the sitagliptintreated group ranged from 0.4% to 0.8% (the highest dose achieved the greatest reduction), compared with a reduction of 1.0% in the glipizide-treated group. Hypoglycemia was much more common in the glipizide arm: 21 patients (17%) experienced at least one hypoglycemic event compared with 2 patients in the placebo group and 2 patients in the sitagliptin group. No signiÞcant weight gain was observed in the sitagliptin-treated groups compared with a 1.1 kg weight gain in the glipizide arm. Sitagliptin was slightly less effective than glipizide in reducing HbA1c levels but its side-effect proÞle was more favorable. Another study demonstrated that sitagliptin is effective in lowering blood glucose levels without causing weight gain. The double-blind, randomized, eightweek crossover trial involved 28 metformin-treated patients who were not reaching their glycemic targets (Brazg R, 2005). The patients continued with their existing metformin therapy and were randomized to one of two regimens: placebo for four weeks, then the addition of twice-daily sitagliptin for the next four weeks; or twice-daily sitagliptin for four weeks, then the addition of placebo. Generally, adding sitagliptin to metformin monotherapy reduced glucose levels (24-hour glucose levels: 157.9 mg/dL in the metformin-only group versus 125 mg/dL in the sitagliptin group; FPG levels: -3.4 mg/dL in the metformin-only group versus -24 mg/dL in the sitagliptin group). Sitagliptin was not associated with weight gain, increased gastrointestinal side effects, or increased hypoglycemia in this study. The investigators did not provide speciÞc data. Other studies have shown that sitagliptin is generally safe and well tolerated. One study demonstrated that the agent does not cause hypoglycemia in healthy subjects (Bergman A, 2005; Herman GA, 2005; Stevens C, 2005). Inhaled Insulins Overview. Several major pharmaceutical companies have licensed proprietary inhalation technologies from biotechnology companies to develop inhaled formulations of insulin. The following inhaled insulins are in late-stage development for type 2 diabetes: Nektar/PÞzer/SanoÞ-Aventis’ Exubera, Aradigm/Novo Nordisk’s AERx, and Alkermes/Eli Lilly’s AIR. Kos Pharmaceuticals and Mannkind (formerly Pharmaceutical Discovery) also have inhaled insulins in advanced stages of

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development. The most important features distinguishing these devices are their relative size and weight. Other companies developing inhaled insulins include Coremed and BMS (in collaboration with QDose); in view of the paucity of data on these products, they are not discussed in detail here. Patients who have used the various inhalation devices welcome this mode of delivery and comment that if it were available they would be more receptive to insulin therapy (Freemantle N, 2005; Hayes RP, 2005; Testa M, 2002). Nevertheless, despite the generally positive outlook for inhaled insulin therapy, there are several reasons for caution in predicting how widely inhaled insulins will be adopted: •

•

•

•

Absorption of inhaled insulin is less efÞcient than direct injection, so it requires more insulin. Therefore, although the cost of inhaled insulin therapy is not yet known, it is likely to be signiÞcantly more expensive than injectable insulin. Because insulin is a growth factor for many cell types, some physicians are concerned that long-term use might damage lung tissues (e.g., pulmonary Þbrosis). Although the convenience and noninvasive nature of inhaled insulin are generally regarded as signiÞcant advantages, the inhalation devices in development are bulkier than asthma inhalers and less convenient to use than an insulin pen. Because the inhaled insulins in development are short-acting insulins, many diabetic patients will still need to administer injections of basal insulin.

Mechanism of Action. In type 2 diabetes, the function of the pancreatic beta cell becomes impaired such that insufÞcient amounts of insulin are released. Therefore, exogenous insulin is needed to substitute for the lack of endogenous insulin. Insulin inhalers work in a manner similar to that of asthma inhalers. The insulin is propelled into the lungs upon inhalation and is absorbed into the bloodstream through the alveoli. Exubera . Nektar (formerly Inhale Therapeutic Systems), in collaboration with PÞzer and SanoÞ-Aventis, is developing a dry-powder insulin formulation called Exubera that uses Nektar’s Inhance dry-powder inhaler technology. Unlike conventional dry-powder inhalation devices, Nektar’s system is designed to deliver the correct dosage independent of the patient’s inspiratory ßow rate. Exubera will likely be the Þrst inhaled insulin to reach the market. PÞzer and SanoÞAventis will commercialize the product worldwide and are preparing additional safety information to augment the interim safety data submitted to the European Medicines Agency (EMEA). Exubera is preregistered in the United States and Europe. The FDA’s Endocrinologic and Metabolic Drugs Advisory Committee reviewed this agent in September 2005 and recommended approval for both type 1 and type 2 diabetes (PÞzer, press release, September 8, 2005).

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In June 1999, Phase III trials involving type 1 and 2 diabetics began at 117 sites in the United States and Germany; they were completed in July 2001. In December 2001, the developers postponed the NDA so as to collect additional long-term pulmonary safety data. In October 2002, the collaborators announced that additional long-term clinical trials in type 1 and 2 diabetic patients were under way and that they were in ongoing discussions with regulatory authorities regarding the timing of their submissions (SanoÞ-Aventis, press release, October 11, 2002). In early 2004, the EMEA sent a 70-day review letter to PÞzer and SanoÞ-Aventis. After responding, PÞzer and SanoÞ-Aventis received a further 120-day review letter in July 2004. At the May 2001 meeting of the American Thoracic Society in San Francisco, researchers reported that Exubera was found to be safe after two years’ administration in a trial involving 140 patients: no evidence of lung function impairment was observed in the trial. In the same month, however, Aventis and PÞzer disclosed one case of pulmonary Þbrosis in the 1,000 patients who participated in various trials lasting more than six months—although a direct link to Exubera has not been established. If more cases of Þbrosis emerge, the FDA may require warnings on the product label, a move that would discourage physicians from prescribing Exubera as a Þrst-line therapy. At the 62nd ScientiÞc Sessions of the ADA in June 2002, Nektar presented the results of a Phase III trial. The three-month multicenter study investigated the efÞcacy of inhaled insulin in 309 patients who were poorly controlled with oral antidiabetic agents (i.e., HbA1c > 8%) (Rosenstock J, 2002). Patients were randomized into three treatment arms: inhalation therapy alone (one to two inhalations before meals); inhalation therapy with existing oral antidiabetic agents; or oral antidiabetic agents alone. Patients taking inhaled insulin—either as monotherapy or in combination—exhibited HbA1c reductions of 1.4% and 1.9%, respectively, from baseline, compared with a 0.2% reduction in patients treated with oral agents alone. Postprandial glucose reductions of 43 mg/dL and 24 mg/dL were observed in the inhaled insulin monotherapy and combination groups, respectively, compared with a 2 mg/dL reduction in the oral monotherapy group. On the downside, hypoglycemia occurred more frequently in the inhaled insulin group (1.3–1.7 events per subject-month, compared with 0.1 event per subject-month in the oral monotherapy group). In addition, patients on inhaled insulin therapy experienced an average weight gain of 2.8 kg; no weight gain was observed in the oral monotherapy group. Studies presented at the 63rd ScientiÞc Sessions of the ADA in June 2003 addressed concerns about pharmacodynamics. One assessment found that smoking alters the absorption of Exubera and concluded that patients should abstain before and during treatment (Becker RHA, 2003[a]). In another study involving elderly, obese type 2 diabetics, investigators showed that the absorption proÞle of Exubera has “within-subject variability” comparable to that of subcutaneous insulin (Mudaliar S, 2003). At the same meeting, researchers presented the results of a three-month, headto-head trial of Exubera and rosiglitazone (DeFronzo RA, 2003). In this study,

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investigators randomized 145 type 2 diabetic patients who were not optimally controlled on diet and exercise to preprandial inhaled insulin (one to two inhalations of 1 or 3 mg) or rosiglitazone (4 mg twice daily). At the end of the treatment period, 83% of Exubera-treated patients achieved HbA1c < 8% compared with 58% of rosiglitazone-treated patients. Additionally, Exubera induced average HbA1c reductions of 2.3%, compared with 1.4% for rosiglitazone. However, inhaled insulin caused greater weight gain (1.9 kg versus 0.8 kg) and more hypoglycemic events (0.7 event/subject-month versus 0.05 event/subject-month). A six-month, randomized trial compared the efÞcacy of preprandial doses of Exubera (plus ultralente) with that of two subcutaneous injections of mixed insulin in 299 type 2 diabetic patients on stable insulin therapy (Hollander PA, 2004). Patients on Exubera experienced HbA1c reductions of 0.7% and greater treatment satisfaction compared with 0.6% in the subcutaneous insulin group. Furthermore, 47% of the Exubera-treated patients reached HbA1c <7%, compared with 32% in the subcutaneous insulin arm. Patients treated with Exubera experienced coughing at a greater rate than did patients treated with mixed insulin injections, but all other adverse events, including hypoglycemia, were similar between the two arms. Increased immunogenicity was detected with Exubera but did not correlate with dose, glycemic control, or adverse events. Most importantly, no difference in pulmonary function was detected. Overall, clinical trials of Nektar’s Exubera in type 2 diabetics demonstrate the following: •

•

Exubera (as monotherapy or in combination with oral antidiabetic agents) improves glycemia in patients who are inadequately controlled with oral antidiabetic agents but it is associated with a greater risk for hypoglycemia. Exubera achieves the same level of glycemic control as conventional injected insulin with a comparable level of side effects.

AERx Insulin Diabetes Management System . Novo Nordisk is developing an inhaled insulin based on Aradigm’s AERx Insulin Diabetes Management System (iDMS) (NN-1998) for pulmonary delivery of insulin. In September 2004, Novo Nordisk expanded the licensing agreement to assume full responsibility for the development of the AERx inhaled insulin. It is in Phase II development for type 2 diabetes in the United States and Europe. AERx is also in Phase III development for type 1 diabetes in these two regions and in Phase I development for type 1 diabetes in Japan. Novo Nordisk has rights to the worldwide marketing of AERx inhaled insulin. Aradigm’s proprietary AERx system has approximately the same dimensions and weight as a small video camera. It releases a Þne, controlled aerosol mist of insulin while a handheld electronic inhaler with computerized sensors adjusts to the patient’s breathing to automatically deliver the drug at the optimal point during inhalation. Aradigm has been issued a U.S. patent relating to the regulation of patient breathing for effective and reproducible pulmonary delivery of insulin. AERx is by far the most technologically advanced of the insulin

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inhalers in development and so will likely be the most expensive of the inhaled insulins. Because of the importance of dosage precision and reproducibility in insulin-treated patients, particularly type 1 diabetics, this product will provide a major advantage over Exubera. Additionally, the electronic compliance monitoring feature will offer valuable information to physicians. A pharmacokinetic study in nonsmoking type 1 diabetic patients demonstrated that AERx insulin has a faster onset of action than that of subcutaneous regular insulin and an onset of action similar to that of insulin aspart, a rapid-acting insulin analogue (Petersen AH, 2005). Two clinical studies showed that the AERx system is safe and well tolerated but that it requires specialized dosing in diabetics with asthma and those who smoke. In the Þrst trial, researchers examined a group of 45 nondiabetic, nonsmoking subjects, 17 of whom were diagnosed with mild-to-moderate asthma (Henry RR, 2003). Results indicated that patients with asthma consistently exhibited less insulin absorption and, consequently, lower reduction of blood glucose levels compared with the control group. Researchers did not observe signiÞcant changes in lung function measurements after administration of AERx iDMS. Although the pulmonary delivery of insulin was well tolerated in this trial, the data suggest that type 2 diabetic patients with asthma—because of variability in the size of their airway openings, blood ßow, and airway secretions—may require a higher dose of insulin to control their blood glucose levels. The second trial investigated the pharmacokinetics of this system in 27 nondiabetic smokers and 16 nondiabetic nonsmokers (Himmelmann A, 2003). Because a signiÞcant percentage of the type 2 diabetic population consists of smokers, the researchers sought to evaluate the effect of smoking on the absorption of inhaled insulin. Results indicated that approximately 60% more insulin was absorbed in smokers compared with nonsmokers because of the increased permeability of the cells comprising the capillary-alveoli border in smokers. A 12-week, open-label, parallel-group trial demonstrated that AERx iDMS reduces HbA1c levels to the same degree as subcutaneous injections (Hermansen K, 2004). In this trial, 107 patients with insulin-treated type 2 diabetes were randomized to preprandial AERx iDMS or short-acting insulin injections, both in combination with NPH insulin. Both groups demonstrated reductions in HbA1c (−0.69% in the AERx arm and −0.77% in the insulin injections arm). In addition, subjects in both groups experienced the same frequency and types of adverse events: 54% in AERx versus 55% in the insulin injections group. The most frequent adverse events were headache, upper respiratory tract infection, and diarrhea—all of which were characterized as mild to moderate. AIR. In April 2001, Lilly entered into a partnership with Alkermes to develop short- and long-acting inhaled formulations of insulin using Alkermes’s Advanced Inhalation Research (AIR) delivery system. Under the terms of the agreement, Lilly holds exclusive worldwide rights to products resulting from this collaboration and has responsibility for conducting clinical trials, securing regulatory approvals, overseeing large-scale manufacturing, and marketing the drug. In a January 2003 company presentation, Alkermes announced that a formulation of

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the AIR delivery system had entered Phase II trials in the United States for type 1 diabetes. Although development is for type 1 diabetes, AIR insulin has the potential to be used off-label for type 2 diabetes. As a compact inhaler that can deliver small peptides and complex macromolecules, the AIR system has the potential to deliver sustained-release formulations of drugs such as long-acting insulin, representing an advantage over other inhaled insulin formulations. The unique AIR insulin formulation is a lowdensity, porous particle that can be delivered using small, mechanically operated inhalers that can accommodate high drug doses. Because of the characteristics of the AIR formulation, a simple, inexpensive, breath-actuated inhaler can be used without the need for fans or motors, as are required for the Exubera and AERx inhaled insulins. Clinical data from large-scale trials are limited, but a few small-scale studies demonstrated AIR insulin’s safety and efÞcacy. Phase I data presented at the ADA’s June 2001 ScientiÞc Sessions show that, in a study of 12 healthy subjects, AIR insulin has a faster onset of action than that of insulin lispro or regular insulin (Osborn C, 2001). Results from an open-label randomized crossover trial involving 20 healthy nonsmoking subjects indicated that AIR insulin’s bioavailability and biopotency were consistent across the various doses and that the system had a prolonged duration of effect compared with insulin lispro (Rave K, 2005). Lung function as measured by forced expiratory volume in one second (FEV1 ) was similar before and after dosing. Findings from a 12-week, randomized, open-label study suggest that AIR is as safe and effective as subcutaneous insulin (Garg S, 2005). In this trial, 250 type 1 patients were randomized to AIR insulin plus glargine or subcutaneous shortacting insulin plus glargine. Both groups achieved the same level of HbA1c after treatment (7.9% with AIR versus 8.0% with subcutaneous insulin). However, AIR demonstrated better reduction in FPG, an action that may have triggered the greater incidence of nocturnal hypoglycemia (4.2 episodes per 30 days with AIR versus 2.7 per 30 days with subcutaneous insulin). Overall hypoglycemic rates were the same between the groups (0.17 with AIR versus 0.13 with subcutaneous insulin). Although AIR is likely to be the third-to-market inhaled insulin for diabetes, the device is by far the smallest inhaler in late-stage development. Furthermore, according to Alkermes’s Web site, the AIR inhaler may be priced more favorably than the other two insulin inhalers because of its smaller size and simpler mechanism. The cost issue is a major consideration with this of class of agents and for type 2 diabetics, who are often taking multiple agents. Kos’s Breath-Actuated Insulin Inhaler. Kos is developing a breath-actuated insulin (BAI) inhaler for type 2 diabetes. The device is a metered-dose inhaler with a Vortex nozzle actuator and electronic dose counter. At this time, the company is actively seeking to outlicense the development of this drug, which is in Phase II development in the United States (Kos Pharmaceuticals, 2004 annual report).

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A small-scale, randomized, 28-day comparator trial involving 24 type 2 diabetic patients poorly controlled with metformin and/or sulfonylureas found that the addition of Kos’s BAI inhaler to oral antidiabetic therapy was more effective than adding subcutaneous insulin glargine (Hausmann M, 2004). Patients were randomized to preprandial Kos inhaled insulin or bedtime subcutaneous glargine as add-on therapies to existing oral regimens. An HbA1c reduction of 1.23% was achieved by the Kos inhaled insulin arm compared with a reduction of 1.05% in the glargine arm. Four episodes of hypoglycemia were reported with the Kos inhaler compared with three with glargine. Technosphere Inhaled Insulin. Mannkind (formerly Pharmaceutical Discovery) is developing a dry-powder inhaled insulin based on its Technosphere formulation of insulin and its MedTone inhaler. In 2004, Mannkind completed Phase II trials in the United States and met with the FDA (Mannkind, 2004 annual report). Phase III trials began in June 2005 in the United States and at the end of 2004 in Europe. The Technosphere inhaled formulation of insulin has been shown to mimic the early-phase insulin response in type 2 diabetes better than subcutaneous insulin as demonstrated by its ability to induce faster disposal of postprandial glucose (Boss AH, 2005). According to the company, Technosphere insulin enters the bloodstream within 10–14 minutes of dosing, approximating Þrst-phase insulin release—a Þrst-phase spike in a healthy person normally occurs within 5–10 minutes once food enters the digestive tract. The MedTone inhaler device is compact and can Þt into the palm of the patient’s hand. Limited data are available at this time, but data from a recent trial presented in June 2005 at the 65th ScientiÞc Sessions of the ADA show some promise. The randomized, double-blind, placebo-controlled trial involved 119 type 2 diabetics whose hyperglycemia was not controlled on diet or oral antidiabetics alone (Rosenstock J, 2005). Patients were randomized to Technosphere insulin or placebo. After 12 weeks, Technosphere insulin reduced HbA1c by 0.76% compared with a reduction of 0.32% in the placebo arm. The greatest effect was observed among patients with baseline HbA1c between 8% and 10.5%; in this subpopulation, Technosphere insulin reduced HbA1c by 1.37% compared with −0.51% in the placebo group. No severe hypoglycemia was reported in any of the groups and lung function tests showed no differences between the groups. Oral Insulin Overview. Although inhaled insulin is garnering the attention of the medical community, several companies are developing oral formulations of insulin in the form of buccal spray or oral tablets. Generex Biotechnology’s buccal spray Oralin is the most clinically advanced of these formulations. Other companies pioneering the development of oral insulin are Nobex/Biocon, BioSante, Emisphere, and Flamel. Of note, Nobex has developed an oral insulin tablet HIM-2 up to Phase II. However, in 2004, the company engineered a more-potent analogue of HIM2 and is seeking to develop that compound, which is at the preclinical stage (www.nobexcorp.com, accessed July 12, 2005).

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In addition to its convenient mode of administration, oral insulin offers numerous physiological beneÞts compared with inhaled insulins. Mimicking the natural trafÞcking pattern of endogenous insulin, oral insulin enters the liver via the portal vein after being absorbed from the gastrointestinal tract. Oral insulins therefore have the potential to improve hepatic glucose control and thus improve FPG levels. The development of orally administered insulin presents many challenges as well: the technology must offer protection from the hostile environment of the digestive tract; the patient’s eating and drinking behavior will greatly inßuence the effective doses that are absorbed into the bloodstream; and long-term safety still needs to be determined. Mechanism of Action. In type 2 diabetes, the function of the pancreatic beta cell becomes impaired such that insufÞcient amounts of insulin are released. Oral insulins are delivered into the bloodstream by the liver (following absorption by the gastrointestinal tract) or directly into the circulation via the buccal mucosa by spraying the insulin formulation directly into the mouth. Buccal delivery of insulin bypasses the portal vein, thereby limiting the effect of insulin on the liver. Oralin Until May 2003, Generex Biotechnology and Eli Lilly were codeveloping an oral insulin formulation using RapidMist, Generex’s proprietary buccal drug delivery technology. Following Lilly’s withdrawal from the partnership, Generex announced its intention to continue developing the product, named Oralin, with other partners (Eli Lilly and Generex Biotechnology, press release, May 23, 2003). Generex subsequently partnered with Kinexum and announced that the latter company would manage the development of Generex’s clinical and regulatory divisions and oversee clinical activities in all therapeutic areas under study at Generex (Generex Biotechnology, press release, December 2003). Oralin has been in Phase IIb trials for type 1 and type 2 diabetes in the United States and Europe since 2003. It was approved in Ecuador in 2005. The RapidMist drug delivery system allows patients to spray a rapid-acting insulin into the buccal cavity; this in-mouth spray has been shown to have a faster absorption and elimination proÞle than that of injected short-acting insulin (Modi P, 2002[b]). Pharmacodynamic studies in healthy individuals and type 1 diabetics show that Oralin’s onset of action is faster than that of subcutaneous regular insulin and its time to peak onset is similar to that of insulin lispro (Guevara-Aguirre J, 2003[a]; Raz I, 2003). In type 1 diabetics, researchers observed no differences in absorption on three different occasions (Guevara-Aguirre J, 2003[b]). Preliminary efÞcacy studies in type 1 diabetic patients demonstrated that Oralin controls glucose levels in a manner comparable to that of subcutaneous insulin (Modi P, 2003). Most of the data that are available are based on small-scale clinical trials, making it difÞcult to assess Oralin’s efÞcacy. Data presented at the 62nd ScientiÞc Sessions of the ADA in June 2002 conÞrm intriguing early reports that Generex’s oral insulin can be used effectively in combination with currently available

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oral antidiabetic agents to achieve glycemic control. A single-blind, randomized, crossover study assessed the efÞcacy of oral insulin as an adjunct therapy to oral antidiabetic agents in controlling postprandial hyperglycemia in type 2 diabetics after a standard meal challenge (Guevara-Aguirre J, 2002). The trial involved 13 patients randomized to combination therapy with oral antidiabetic agents and oral insulin or oral antidiabetic agents alone. Patients inhaled 15 puffs 15 minutes prior to a 360-calorie Ensure meal. Glucose, insulin, and C-peptide levels were measured at one, two, and four hours after the meal. Results indicated that the addition of Oralin reduced postprandial glucose and C-peptide levels and elevated serum insulin concentrations to a greater extent than oral antidiabetic agents alone. A proof-of-concept study was conducted in type 2 diabetics on multiple daily injections with the goal of demonstrating that Oralin can replace subcutaneous injections. This open-label, crossover, randomized study involved only 23 type 2 diabetes patients (Guevara-Aguirre J, 2004). Patients were randomized to one of two groups: one dose of subcutaneous injection and then three to seven days later Oralin (100 U) or Oralin administered Þrst, followed by the subcutaneous injection. Patients were given a standard breakfast ten minutes after the dose, and 30- and 60-minute postprandial glucose levels were measured. Results indicated that postprandial glucose levels were lowered to a greater degree with Oralin than with subcutaneous injections. During the Þrst-hour after dosing, C-peptide dropped to a greater extent in Oralin-treated patients than in those patients who were subcutaneously injected, indicating a reduction in endogenous insulin production. Insulin levels were signiÞcantly higher with Oralin during the Þrst 30 minutes compared with the injection. Investigators concluded that Oralin can be used instead of premeal injections to control postprandial glucose levels. In a double-blind, randomized, 12-week study in type 2 diabetics with poorly controlled glucose on metformin plus glargine, 26 patients were randomized to seven puffs of Oralin or seven puffs of placebo to be taken ten minutes before each meal in addition to existing therapy (Raz I, 2005). Only eight-week interim results were available for presentation at the 65th ScientiÞc Sessions of the ADA. No differences in FPG were observed, but postprandial glucose levels fell by 15.4% in the treated group compared with an increase of 3.9% in the placebo group. No large-scale human trial data are available regarding Oralin’s safety, but Generex announced the results of a two-year safety and toxicology animal study (Generex, press release, August 15, 2002). Researchers found that chronic administration of Oralin did not change the epithelial cells of the mouth or induce toxic injury. Previously, the company announced the results of studies suggesting that the oral spray poses no risk to the lungs because its droplets are too large to enter deep lung regions (Generex, press release, November 8, 2001). REFERENCES Adler AI, et al. Association of systolic blood pressure with macrovascular and microvascular complications of type 2 diabetes (UKPDS 36): prospective observational study. British Medical Journal. 2000;321(7258):412–419.

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Botas P, et al. Prevalence of diabetes mellitus and glucose intolerance in the population aged 30 to 75 years in Asturias, Spain. Revista Clinica Espanola. 2002;202:423–429. Bourdel-Marchasson I. Five-year mortality in elderly French subjects from the PAQUID epidemiological survey: the burden of diabetes. Diabetic Medicine. 1998;830–835. Bouret SG, et al. Trophic action of leptin on hypothalamic neurons that regulate feeding. Science. 2004;304(5667):108–110. Bouskila M, et al. Adiponectin: a relevant player in PPAR-gamma-agonist-mediated improvements in hepatic insulin sensitivity? International Journal of Obesity and Related Metabolic Disorders. 2005;29(suppl 1):S17–S23. Boyle PJ, et al. Effects of pioglitazone and rosiglitazone on blood lipid levels and glycemic control in patients with type 2 diabetes mellitus: a retrospective review of randomly selected medical records. Clinical Therapeutics. 2002;24(3):378–396. Brazg R, et al. Effect of adding MK-0431 to ongoing metformin therapy in type 2 diabetic patients who have inadequate glycemic control on metformin. 65th ScientiÞc Sessions of the American Diabetes Association Abstracts. 2005;54(supp 1). Abstract 11–OR. Brecchia M, et al. Imatinib mesylate may improve fasting blood glucose in diabetic Ph+ chronic myelogenous leukemia patients responsive to treatment. Journal of Clinical Oncology. 2004;22(22):4653–4655. Bregenholt S, et al. The GLP-1 analogue NN2211 inhibits free fatty acid-induced apoptosis in primary rat beta cells. European Association for the Study of Diabetes. September 10, 2001. Abstract 65. Brunmair B, et al. RWJ241947: an antidiabetic thiazolidinedione with reduced adipogenic action? Diabetologia. 2002;45(suppl 2):235. Bruno G. A population-based prevalence survey of known diabetes mellitus in northern Italy based upon multiple independent sources of ascertainment. Diabetologia. 1992;35:851–856. Buffon A. Anti-inßammatory effects of peroxisome proliferator-activated receptor-gamma agonist in type 2 diabetic patients. 65th ScientiÞc Sessions of the American Diabetes Association Abstracts. 2005;54(supp 1). Abstract 655–P. Bulpitt CJ, et al. Association of symptoms of type 2 diabetic patients with severity of disease, obesity, and blood pressure. Diabetes Care. 1998;21(1):111–115. Burgers JS, et al. Comparative analysis of recommendations and evidence in diabetes guidelines from 13 countries. Diabetes Care. 2002;25:1933–1939. Carpentier A, et al. Acute enhancement of insulin secretion by FFA in humans is lost with prolonged FFA elevation. American Journal of Physiology. 1999;276:E1055–E1066. Castell C, et al. Prevalence of diabetes in Catalonia (Spain): an oral glucose tolerance test-based population study. Diabetes Research and Clinical Practice. 1999;43:33–40. Centers for Disease Control and Prevention (CDC). Behavioral risk factor surveillance system survey data. Atlanta, Georgia: U.S. Department of Health and Human Services, Centers for Disease Control and Prevention; 2004. Available at: www.cdc.gov/brfss. Accessed August 30, 2005. Centers for Disease Control and Prevention (CDC). National Center for Health Statistics (NCHS). National Health and Nutrition Examination Survey Data, 1999–2002. Hyattsville, MD: U.S. Department of Health and Human Services, Centers for Disease Control and Prevention, 2005. Available at: www.cdc.gov/nchs/nhanes.htm. Accessed: May 24, 2005.

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Chang AM, et al. The long-acting GLP-1 derivative, NN2211, restores beta-cell sensitivity to glucose in subjects with type 2 diabetes. 63rd ScientiÞc Sessions of the American Diabetes Association Abstracts. 2003;52(suppl 1). Abstract 486–P. Charbonnel B, et al. The prospective pioglitazone clinical trial in macrovascular events (PROactive); can pioglitazone reduce cardiovascular events in diabetes? Study design and baseline characteristics of 5,238 patients. Diabetes Care. 2004;27(7):1647–1653. Charles MA. Revision des criteries diagnostiques du diabetes. Les raisons et les consequences. Diabetes Metab. 1998;24:75–79. Charpentier G, et al. Improved glycemic control by addition of glimepiride to metformin monotherapy in type 2 diabetic patients. Diabetic Medicine. 2001;18(10):828–834. Cheng PT, et al. BMS-298585 is a novel, uniquely balanced dual activator of peroxisome proliferator-activated receptors (PPAR)-alpha and -gamma, with an excellent ADME proÞle. 62nd ScientiÞc Sessions of the American Diabetes Association. 2002;51(suppl 2). Abstract 381–P. Chevre JC, et al. Insulin promoter factor 1 gene is not a major cause of maturity-onset diabetes of the young in French Caucasians. Diabetes. 1998;47:843–844. Chiasson JL, et al. STOP-NIDDM Trial Research Group. Acarbose for prevention of type 2 diabetes mellitus: the STOP-NIDDM randomised trial. Lancet. 2002;359(9323):2072–2077. Clark HE, et al. The effect of glimepiride on pancreatic beta-cell function under hyperglycemic clamp and hyperinsulinemic, euglycemic clamp conditions in non-insulindependent diabetes mellitus. Hormone and Metabolism Research. 1996;28(9):445–450. Cline GW, et al. Impaired glucose transport as a cause of decreased insulin-stimulated muscle glycogen synthesis in type 2 diabetes. New England Journal of Medicine. 1999;341:240–246. Clore JN, et al. Glucose-6-phosphatase ßux in vitro is increased in type 2 diabetes. Diabetes. 2000;49:969–974. Colhoun HM. The scope for cardiovascular disease risk factor intervention among people with diabetes mellitus in England: a population-based analysis from the Health Survey for England 1991–1994. Diabetic Medicine. 1999;16:35–40. Committee of Japan Diabetes Society (JDS) for the Diagnostic Criteria of Diabetes Mellitus. Report of the Committee of Japan Diabetes Society on the classiÞcation and diagnostic criteria of diabetes mellitus. 2000. Available at: www.jds.or.jp. Coniff RF, et al. Long-term efÞcacy and safety of acarbose in the treatment of obese subjects with non-insulin-dependent diabetes mellitus. Archives of Internal Medicine. 1994;154(21):2442–2448. Cooksey RC, et al. Mechanism of hexosamine-induced insulin resistance in transgenic mice overexpressing glutamine: fructose-6-phosphate amidotransferase: decreased glucose transporter GLUT4 translocation and reversal by treatment with thiazolidinedione. Endocrinology. 1999;140(3):1151–1157. Cowie CC. Prevalence of diabetes and impaired fasting glucose in adults—United States, 1999–2000. Morbidity and Mortality Weekly Report. 2003;52:833–837. Cramer JA, et al. Compliance with inhaled insulin treatment using AERx iDMS Insulin Diabetes Management System. 63rd ScientiÞc Sessions of the American Diabetes Association Abstracts. Diabetes. 2003;52(suppl 1). Abstract 442–P.

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Cricelli C, et al. Prevalence estimates for chronic diseases in Italy: exploring the differences between self-report and primary care databases. Journal of Public Health Medicine. 2003;25:254–257. Cusi K, et al. Insulin resistance differentially affects the PI3 kinase- and Map kinase-mediated signaling in human muscle. Journal of Clinical Investigation. 2000;105(3):311–320. Dabelea D, et al. Type 2 diabetes mellitus in minority children and adolescents: an emerging problem. Pediatric Endocrinology. 1999;28:709–729. Davidson JA, et al. EfÞcacy of orlistat in patients with type 2 diabetes with baseline HbA1c levels >8%. 62nd ScientiÞc Sessions of the American Diabetes Association Abstracts. 2002;51(suppl 2). Abstract 384–P. Davies MJ. Screening for type 2 diabetes mellitus in the U.K. Indo-Asian population. Diabetic Medicine. 1999;16:131–137. DECODA Study Group. Age- and sex-speciÞc prevalence of diabetes and impaired glucose regulation in 11 Asian cohorts. Diabetes Care. 2003;26:1770–1780. DECODE Study Group. Will new diagnostic criteria for diabetes mellitus change phenotype of patients with diabetes? Reanalysis of European epidemiological data. British Medical Journal. 1998;317;371–375. DECODE Study Group. Age- and sex-speciÞc prevalences of diabetes and impaired glucose regulation in 13 European cohorts. Diabetes Care. 2003;26:61–69. DECODE Study Group on behalf of the European Diabetes Epidemiology Study Group. Is fasting glucose sufÞcient to deÞne diabetes? Epidemiological data from 20 European studies. Diabetologia. 1999;42:647–654. [a] DECODE Study Group on behalf of the European Diabetes Epidemiology Study Group. Consequences of the new diagnostic criteria for diabetes in older men and women. Diabetes Care. 1999;22:1667–1671. [b] DeFronzo RA, et al. EfÞcacy of metformin in patients with non-insulin-dependent diabetes mellitus. The Multicenter Metformin Study Group. New England Journal of Medicine. 1995;333:541–549. DeFronzo RA, et al. EfÞcacy and safety of inhaled insulin (Exubera) compared with rosiglitazone in type 2 diabetes patients not optimally controlled on diet and exercise: results of a 3-month, randomized, comparative trial. 63rd ScientiÞc Sessions of the American Diabetes Association Abstracts. 2003;52(suppl 1). Abstract 162–OR. Degn KB, et al. One week’s treatment with NN2211, a long-acting GLP-1 derivative, markedly ameliorates 24-h glycemia and b-cell function and reduces fasting endogenous glucose release in type 2 diabetic patients. 63rd ScientiÞc Sessions of the American Diabetes Association Abstracts. 2003;52(suppl 1). Abstract 498–P. De Marco R, et al. Cause-speciÞc mortality in type 2 diabetes. The Verona Diabetes Study. Diabetes Care. 1999;22(5):756–761. Derosa G, et al. A comparison of the effects of pioglitazone and rosiglitazone combined with glimepiride on prothrombotic state in type 2 diabetic patients with metabolic syndrome. Diabetes Res Clin Pract. 2005;69(1):5–13. Detournay B, et al. Managing type 2 diabetes in France: the ECODIA survey. Diabetes Metab. 2000;26(5):363–369. Diabetes Prevention Program Research Group. The Diabetes Prevention Program (DPP): description of lifestyle intervention. Diabetes Care. 2002;25(12):2165–2171.

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World Health Organization (WHO) Consultation: DeÞnition, diagnosis, and classiÞcation of diabetes mellitus and its complications. I. Diagnosis and classiÞcation of diabetes mellitus. Geneva, World Health Organization. 1999;99:2. Wu G, et al. Presence of glutamine:fructose-6-phosphate amidotransferase for glucosamine-6-phosphate synthesis in endothelial cells: effects of hyperglycemia and glutamine. Diabetologia. 2001;44(2):196–202. Yamagata K, et al. Mutations in the hepatocyte nuclear factor-4a gene in maturity-onset diabetes of the young (MODY1). Nature. 1996;384:458–460. [a] Yamagata K, et al. Mutations in the hepatocyte nuclear factor-1a gene in maturity-onset diabetes of the young. Nature. 1996;384:455–458. [b] Yazdani-Biuki B, et al. Improvement of insulin sensitivity in insulin-resistant subjects during prolonged treatment with the anti-TNF-alpha antibody inßiximab. European Journal of Clinical Investigation. 2004;34:641–642. Yki-J¨arvinen H, et al. Less nocturnal hypoglycemia and better postdinner glucose control with bedtime insulin glargine compared with bedtime NPH insulin during insulin combination therapy in type 2 diabetes. HOE 901/3002 Study Group. Diabetes Care. 2000;23(8):1130–1136. Zhu F, et al. The role of urotension II gene in the genetic susceptibility to type 2 diabetes in Chinese population. Zhonghua Yi Xue Za Zhi. 2002;82(21):1473–1475. Zimmermann R, et al. Fat mobilization in adipose tissue is promoted by adipose triglyceride lipase. Science. 2004;306(5700):1383–1386. Zimmet PZ, et al. Etiology of the metabolic syndrome: potential role of insulin resistance, leptin resistance, and other players. Annals of the New York Academy of Sciences. 1999;892:25–44. Zimmet P, et al. Global and societal implications of the diabetes epidemic. Nature. 2001;414(6865):782–787.

Acute Exacerbations of Chronic Bronchitis ETIOLOGY AND PATHOPHYSIOLOGY Introduction Acute exacerbations of chronic bronchitis (AECB) are acute episodes of cough, increased dyspnea (shortness of breath), and increased sputum production that occur in patients with chronic bronchitis (CB). CB is deÞned as inßammation of the lung, daily cough, and sputum production during at least three consecutive months for more than two successive years (Sethi S, 2000). Patients with CB typically have chronic overproduction of sputum, airway inßammation, and cough, most often owing to a longstanding history of tobacco use. CB patients are prone to suffering acute exacerbations, typically due to a trigger (such as a bacterial infection) that increases airway inßammation, resulting in increased airway secretions and symptoms of AECB. These symptoms include dyspnea, excess sputum production, and purulent sputum. Because no characteristic laboratory, radiographic, or physiological tests have been developed for AECB, clinical deÞnitions have traditionally been used (e.g., Anthonisen and Seemingal criteria). Anthonisen and colleagues proposed three clinical criteria to deÞne AECB: increased sputum volume, increased sputum purulence, and increased dyspnea over baseline (Anthonisen NR, 1987). These criteria, experts contend, continue to be commonly used by general practitioners. Based on their Þndings, Anthonisen and colleagues proposed three levels of severity: •

Type I is the most severe category, comprising all three cardinal symptoms: worsening dyspnea, increased sputum volume, and increased sputum purulence.

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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Type II is a lower-grade exacerbation in which patients suffer from only two of the three cardinal symptoms. Type III is the least severe grade; patients experience only one of the three cardinal symptoms.

Type I and II appear to beneÞt from antibiotic therapy while antibiotic therapy for patients with type III AECB does not appear to offer signiÞcant improvements over placebo (Anthonisen NR, 1987). Seemungal and colleagues also proposed major and minor criteria to deÞne AECB (Seemungal TA, 2000). The major criteria are the three proposed by Anthonisen and colleagues. Minor criteria include wheeze, sore throat, or symptoms of a common cold such as nasal discharge or congestion. Their deÞnition of AECB is at least two major symptoms or one major symptom and one minor symptom for at least two consecutive days. AECB differs from acute bronchitis in that the former typically occurs as an acute infection in an individual with longstanding underlying chronic obstructive lung disease. In contrast, acute bronchitis typically occurs in an otherwise healthy individual as an acute infection, without any relation to underlying lung disease. Further, AECB is most often bacterial in origin, whereas acute bronchitis is typically caused by viruses. The term acute exacerbations of chronic obstructive pulmonary disease (COPD, described further below) is sometimes used to refer to an acute episode of dyspnea (airßow limitation) with or without infection or sputum production in patients with COPD. COPD is a broader term than CB that also includes emphysema. In practical terms, however, COPD patients often have a combination of emphysema and CB, and clinically, physicians group these patients under the umbrella term of COPD. For the purposes here, the term AECB refers to acute episodes of dyspnea (airßow limitation) with infection and sputum production in patients with CB. This section focuses on the infectious etiology of acute exacerbations. Etiology AECB is a disease that occurs in patients who suffer from chronic underlying airway disease, typically related to tobacco smoking. It does not commonly occur among healthy children or adults. Experts believe that infections are the most common triggers of AECB episodes (50–80%), while inhaled allergens or pollutants trigger approximately 20% of episodes. A critical factor in understanding the etiology of AECB is the underlying lung disease; therefore, a discussion of chronic airway disease leading to AECB is warranted. Chronic Underlying Airway Disease. COPD is an umbrella term that includes the entities of CB and emphysema when they are associated with airßow limitation. Ordinarily, asthma, cystic Þbrosis, bronchiectasis, and bronchiolitis obliterans (other bronchitic syndromes) are excluded from this deÞnition. Most

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deÞnitions consider COPD to be a disease state characterized by airßow limitation that is not fully reversible. The airßow limitation is usually progressive and associated with an abnormal inßammatory response of the lungs to inhaled particles or gases. Individuals with chronic lung diseases such as CB, emphysema, and asthma develop structural damage to their lung architecture as a result of exposure to various environmental or genetic stimulants. These stimulants include cigarette smoke, pollutants, infectious pathogens, and allergens. When pulmonary tissue is exposed to these stimuli, the physiological response is inßammation. When the exposure becomes chronic, the inßammatory response becomes chronic (Quillen DM, 2001). An abnormal inßammatory response may also predispose individuals to chronic airway disease because excessive secretions of the inßammatory cells can cause occlusion of airway passages and/or contraction of airway smooth muscle. The airways subsequently narrow and become obstructed, preventing normal airßow to the lungs (i.e., obstructive lung disease). This impaired airßow compromises the patient’s breathing and other vital processes, reducing quality of life and increasing the risk of life-threatening complications such as pneumonia and myocardial infarctions. CB, emphysema, and asthma differ in their pathophysiological mechanisms but result in the same outcome: impedance of airßow to the lungs and a predisposition to acute respiratory exacerbations and infection. CB is a clinically deÞned disease characterized by inßammation of the lining of the bronchial tubes resulting in diminished airßow to and from the lungs and production of heavy mucus or phlegm. Emphysema is deÞned in terms of anatomic pathology and is characterized by destruction of the walls of the air sacs of the lung, resulting in diminished elasticity of the lungs and decreased respiratory function. It is caused primarily by external stimuli such as cigarette smoking. Emphysema and CB often occur concurrently in the same individuals. Asthma is an inßammatory disorder of the airways, inßuenced by environmental and genetic mechanisms. Asthma is characterized by periodic attacks of wheezing, shortness of breath, chest tightness, and coughing. When any of these three conditions results in irreversibly obstructed airways, they become classiÞed under the umbrella term COPD. As a result, the signs and symptoms of these diseases overlap considerably. In fact, many patients may suffer concomitantly from two or three of these syndromes. The schematic shown in Figure 1 illustrates the overlap in CB, emphysema, asthma, and COPD. Clinically, the vast majority of patients diagnosed with CB and emphysema are classiÞed as having COPD. The frequency of acute exacerbations among patients is variable, similar to the varying rate of progression of CB, emphysema, and asthma among patients. It is important to note that these diseases can reach a “stable” state, with minimal or steady disease progression. However, AECB occurs when this “steady state” is somehow disrupted by a trigger, such as an acute infection. For example, patients suffering from CB ultimately reach a steady state of the disease that is characterized by chronic sputum production, some degree of chronic dyspnea, and limitation of strenuous activity. However, stable patients exposed to a threshold level of stimulants such as cigarette smoke, pollutants, allergens, or

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FIGURE 1. Overlap of obstructive pulmonary diseases.

infectious organisms can suffer from ßare-ups or acute exacerbations whereby the excess inßammation and bronchial secretion results in symptoms of coughing and shortness of breath. Bacterial Involvement. The role of bacteria in CB and AECB has been controversial and a topic of debate for the past 40 years. Bacteria are commonly recovered from the sputa of both clinically stable patients with CB and patients with AECB. However, serological studies comparing serum antibody titers with respiratory pathogens have yielded confusing and often contradictory results (Murphy TF, 1992). These Þndings raised two important questions: •

•

Are the bacteria recovered during exacerbations normal inhabitants (commensals) of the lower respiratory tract (LRT) that contaminate the sputa, or do they truly cause the exacerbation? If the bacteria are responsible for the acute exacerbation, how do they adversely affect lung function during the exacerbation and beyond?

Evidence now exists to support the role of bacteria in acute exacerbations. First, bacteria can be recovered from the LRT not only from sputum samples but also

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from LRT samples obtained using Þberoptic bronchoscopy with a protected specimen brush (PSB) (Monso E, 1995; Soler M, 1998). Sampling the LRT using this method avoids contaminating the sample with any upper respiratory secretions that may harbor commensal organisms. The potential for contamination by commensal organisms remains the chief factor limiting the reliability of sputum samples. The presence of an immune response, identiÞed by the appropriate increase in serum antibodies to bacteria isolated from patients with an exacerbation, further supports the role of bacteria in the pathophysiology of the disease. Studies testing for antibodies against homologous infecting strains have demonstrated that serum collected from patients following an exacerbation contain neutralizing, bactericidal activity against the homologous infecting strain; this activity was absent in serum collected before the exacerbation (Bakri F, 2002; Yi K, 1997). These studies avoid the shortcomings of earlier studies by using immunoassays that are speciÞc to antibodies of surface-exposed epitopes (speciÞc sites on the bacterial antigen), rather than measuring antibody levels to whole bacteria. Additional evidence of the role bacteria play in AECB comes from a number of placebo-controlled studies examining the role of antibiotic treatment in AECB; these studies demonstrate an overall clinical beneÞt to patients treated with antibiotics (Allegra L, 2001; Saint S, 1995). Typically, antibiotic treatment results in a shorter exacerbation, a more rapid return of peak ßow rate, and high overall clinical improvement. Antibiotics may also increase the interval between exacerbations, decrease hospitalization, and prevent the development of secondary bacterial infections or progression to pneumonia. Patients who derive the greatest beneÞt from antibiotic therapy are those with more severe functional impairment and a higher number of exacerbations per year. Pathogens Associated with AECB. Among episodes of infectious etiology, aerobic gram-positive and gram-negative bacteria (50–75%), atypical pathogens (1–10%), and respiratory viruses (30%) are the primary pathogens associated with AECB (Ball P, 1998; Sethi S, 2000; Wilson R, 2000). The relative contribution of these pathogens differs depending on the severity of the underlying airway disease. Approximately 30% of all acute exacerbations are precipitated by respiratory viruses. However, many exacerbations triggered by bacteria are secondary to the viral infection. Clinically, physicians do not differentiate between viral and bacterial causes, owing to a lack of practical viral diagnostics or treatments. Table 1 lists the common pathogens associated with AECB. Pathogenic bacteria most commonly isolated from sputum during an exacerbation include nontypeable Haemophilus inßuenzae (NTHI), Streptococcus pneumoniae, and Moraxella catarrhalis. These bacteria are responsible for the majority (70%) of exacerbations caused by infectious agents, with NTHI usually isolated twice as often as S. pneumoniae (Ball P, 1995[a]; Sethi S, 2002). Atypical organisms such as Mycoplasma pneumoniae and Chlamydia pneumoniae (identiÞed by serology) also play a role in AECB; these organisms are estimated to be associated with 1–10% and 4–5% of all AECB, respectively (Sethi S, 2000).

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TABLE 1. Pathogens Associated with Acute Exacerbations of Chronic Bronchitis Bacteria Haemophilus influenzae Streptococcus pneumoniae Moraxella catarrhalis Staphylococcus aureus Pseudomonas aeruginosa Opportunistic gram-negatives Atypical organisms Mycoplasma pneumoniae Chlamydia pneumoniae Viruses Rhinoviruses Influenza/parainfluenza viruses Respiratory syncytial virus Coronaviruses Herpes simplex virus Adenoviruses

Presencea (%) H. influenzae, S. pneumoniae, and M. catarrhalis account for 70% of exacerbations.

Accounts for ≤10% of exacerbations.

Accounts for 30% of exacerbations.

a Sethi S. Infectious etiology of acute exacerbations of chronic bronchitis. Chest. 2000;117:380S–385S.

Additionally, Enterobacteriaceae and Pseudomonas species play an important role in acute exacerbations, chießy among patients with severe COPD. Table 2 lists the dominating bacteria isolated from patients with varying stages of lung disease in one AECB study, where the composition of pathogens differs by CB severity: group 1 bacteria include S. pneumoniae and Staphylococcus; group 2 bacteria include H. inßuenzae and M. catarrhalis; and group 3 bacteria include Enterobacteriaceae and Pseudomonas species (Eller J, 1998). Haemophilus influenzae. H. inßuenzae is a commensal organism in the upper respiratory tract. It is the most common cause of AECB and tends to cause infection when it descends into the LRT in patients with underlying respiratory illness, such as COPD. H. inßuenzae is a gram-negative bacterium and can also cause pneumonia (infection in the LRT) (Murphy TF, 1992). Furthermore, H. inßuenzae triggers a direct immune reaction in the airways that signiÞcantly reduces mucociliary clearance. H. inßuenzae is isolated in 30–70% of all bacterial exacerbations of chronic bronchitis (Ball P, 1994). H. inßuenzae is increasingly resistant to several classes of antibiotics such as certain macrolides, ßuoroquinolones, cephalosporins, and beta-lactam antibiotics (Doern GV, 2004). Moraxella catarrhalis. M. catarrhalis and S. pneumoniae are the next most common isolates; together, they account for another 24–33% of isolates in AECB (Ball P, 1995[a]). M. catarrhalis causes bronchitis or pneumonia in children and adults with underlying chronic lung disease and is occasionally a cause of bacteremia or meningitis, especially in patients who are immunocompromised. Bacteremia can be complicated by local infections such as osteomyelitis or septic arthritis. M. catarrhalis is also associated with nosocomial infections.

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TABLE 2. Dominating Bacteria in Patients with Acute Infective Exacerbations of Chronic Obstructive Pulmonary Disease

Bacterial Pathogen Group 1 S. pneumoniae S. aureus Group 2 H. influenzae M. catarrhalis Group 3 P. aeruginosa Pseudomonas spp. S. marcescens Klebsiella pneumoniae Proteus vulgaris Escherichia coli Citrobacter spp. Enterobacter spp. S. maltophilia

All Patients

Stage I FEV1 = 50% predicted)

Stage II FEV1 > 35% to <50% predicted)

Stage III FEV1 > 35% predicted)

∼30%

46%

26%

23%

∼20%

24%

33%

14%

∼48%

30%

40%

64%

Note: Table represents the dominating bacteria recovered in studies of patients with varying lung disease severities. Stages I, II, and III are lung disease severities based on the FEV1 values of the patients. (FEV1 = Forced expiratory volume predicted in one second.) The severity of the airway disease is measured based on the forced expiratory volume, which is the measure (in seconds) of the maximum volume of gas that can be expelled from the lungs during a forced vital capacity test. This test indicates the extent of airway obstruction a patient experiences. A lower FEV is indicative of a higher degree of airway obstruction and greater severity of lung disease.

Streptococcus pneumoniae. S. pneumoniae is an important cause of bacterial AECB (accounting for 15–20% of cases) and other respiratory infections. S. pneumoniae is encased in a polysaccharide capsule that resists phagocytosis by macrophages, so the antibody-dependent killing mechanisms (i.e., complement) are important in the immune system’s defense against this organism. S. pneumoniae also contains the enzyme IgA1 protease, which cleaves the IgA in the upper respiratory tract and allows the bacterium to survive in that environment. Infection with S. pneumoniae has become an increasing concern over the past two decades because of the rise of penicillin resistance in the community (discussed later). Gram-Negative Bacilli. Common commensal organisms in the gastrointestinal tract, gram-negative bacilli (GNB), can cause pulmonary disease when aspirated into the respiratory tract. Conditions that compromise the cough and epiglottic reßexes, such as alcoholism and diabetes, increase the likelihood of aspiration and infection of the bronchi. The most common pathogens in this group are Pseudomonas aeruginosa and Klebsiella pneumoniae. Pseudomonas is particularly notable because it is extremely difÞcult to treat, owing to its resistance to many antibiotics.

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Relative Virulence of Bacterial Pathogens. Increased presence of bacteria (bacterial load) is strongly associated with the severity of an acute exacerbation, yet the type of bacteria involved also inßuences the severity of airway inßammation. For example, investigators have demonstrated that exacerbations involving P. aeruginosa are associated with more inßammation than those involving H. inßuenzae (Hill AT, 2000; Stockley RA, 2001). In contrast, M. catarrhalis and H. inßuenzae are associated with a similar degree of inßammation, provided the bacterial loads are similar. Researchers are working to determine why different bacteria cause varying degrees of inßammation. This information might help elucidate the pathogenesis of speciÞc infections and thus identify drug-speciÞc targets unique to these pathogens. Other Etiologies of AECB. Causes other than bacterial infection can also precipitate an acute exacerbation, including viral infection, exposure to inhaled toxic substances, and allergies. The common belief is that infection or other triggers, and the consequent inßammatory response elicited in the bronchi, lead to increased airway obstruction (Madison M, 1998). Viruses (including rhinoviruses, respiratory syncytial virus, inßuenza, parainßuenza, and coronavirus) may be involved in up to one-third of AECB episodes (Smith CB, 1980; Seemungal T, 2001). Although patients with COPD do not seem to have increased susceptibility to these viruses, they face more serious consequences, such as more severe and frequent exacerbations and secondary bacterial infections (Madison JM, 1998). Researchers propose that during a viral infection of the LRT, mucociliary clearance is impaired because of loss of ciliated cells and altered mucus ßow. Viral infection also increases epithelial permeability, via inßammatory mediator proteins such as kinins, which escalates the inßammatory process already ongoing in the chronic bronchitic lung. These changes facilitate subsequent bacterial infection or an increase in the number of bacteria that are already colonizing the lower airways. Although the viral infection may be self-limiting, the secondary bacterial infection can perpetuate the exacerbation. Evidence is also emerging that viruses may enhance the inßammatory response to subsequent infection by activating transcription factors. Transcription factors bind to speciÞc consensus sites on DNA and act to regulate (increase or decrease) the rate of transcription of a speciÞc gene (Rahman I, 2002). Transcription-factormediated enhancement of infection and inßammation is a heated area of research in infectious diseases. Pathophysiology Infection associated with AECB is usually localized to the pulmonary mucosa. Most bacteria that infect the bronchial tree either reside as commensal organisms in the nasopharynx (e.g., H. inßuenzae) or act as opportunistic pathogens invading hosts with suppressed immune systems (e.g., P. aeruginosa). Mucosal infections are usually superÞcial, and most bacteria reside in the lumen, associating with mucus or other secretions. Some pathogenic bacteria adhere to the

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epithelial surface, particularly in areas of epithelial damage, while others inÞltrate the mucosa. In recent years, investigators have focused on determining if the presence of bacteria in the LRT adversely affects lung function in patients with CB or COPD. Research also seeks to determine whether the frequency of exacerbations causes progression of the underlying airway disease in the long term, or vice versa (Spencer S, 2004, Wedzicha JA, 2003). A “vicious cycle hypothesis” (Figure 2) has been proposed as the mechanism by which bacteria may affect the LRT (Sethi S, 1999). This hypothesis supposes that the primary initiating damage in CB results from cigarette smoke, childhood respiratory disease, or other factors, with bacterial colonization playing an important but secondary role. After initial damage to the LRT, infection and/or bacterial colonization results in inßammation. The acute inßammatory response to airway infection stimulates mucus hypersecretion, airway-wall edema, and smooth muscle contraction. This inßammation is caused by the presence of cell-associated bacterial antigens (e.g., lipooligosaccharides from H. inßuenzae and M. catarrhalis, capsular material from S. pneumoniae) and the inßammatory response, which is provoked by the secretion of bacterial products (e.g., cilostatin), elastase from recruited polymorphonuclear

FIGURE 2. Schematic diagram of the vicious cycle hypothesis of the role of bacterial infection in chronic obstructive pulmonary disease.

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leukocytes (PMNLs), and the production of cytokines. Chronic inßammation contributes to the damage done to the airway epithelium and to progression of COPD (Campbell GD, 1999). Experts hypothesize that persistence of infection is more likely caused by the severity of the underlying disease and ensuing damage to the lung defenses rather than the virulence of the infecting pathogens (Wilson R, 1998). Thus, infections become more common with disease progression, further aggravating the underlying inßammatory process. The frequency of exacerbations that are bacterial in origin also depends on the predominant underlying pathology. As such, patients with CB are more susceptible to bacterial bronchial infections than are those who have emphysema or asthma (Wilson R, 2000). Because bacteria have an afÞnity for growth in mucus, impairment of mucociliary clearance in the lungs and an increase in mucous production (as is the case in CB) allow bacteria that are inhaled or aspirated into the bronchial tree an opportunity to colonize the mucosa. In patients with mild CB, these infections can resolve spontaneously; however, as the underlying lung disease worsens, the ability to spontaneously resolve infection diminishes. Immunological Hypothesis of Bacterial Re-Infection. Researchers have proposed a model for the recurrence of exacerbations and perpetuation of bacterial infection in patients with CB and COPD. This model assumes that the virulence of the infecting strain and the presence of preexisting protective antibodies are important factors (Sethi S, 2000). Absence—or low levels—of protective antibodies and/or virulent strains predispose an individual to development of an exacerbation. Infection with a low-virulence strain, on the other hand, would lead only to colonization, without inducing an exacerbation. Exacerbation (and possibly colonization) leads to development of antibodies to the infecting strain; these antibodies successfully clear the pathogen from the LRT (with or without antibiotics). Because a substantial proportion of the antibodies that the body mounts against infecting pathogens, particularly H. inßuenzae, are aimed at hypervariable regions of the outer membrane proteins (and therefore strain-speciÞc), these antibodies do not protect the host from alternate strains that might be antigenically different (Sethi S, 2000). The strain-speciÞc antibodies created by the host assist in protecting the lungs against the initial infecting strain, but new strains then replace the previous strain in a repetitive manner, leading to recurrence of exacerbation episodes. This model suggests that important areas requiring further research include identifying pathogen virulence factors and identifying antigens on the surface of bacteria that are shared among several strains and can elicit a protective antibody response. Role of Host Defenses. The bronchi of normal lungs have a well-primed immune system that is ready to mount an inßammatory response to any insult, including infectious pathogens. Impairment of the host defenses in the bronchitic

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airways provides permissive conditions for invading bacteria to establish themselves in the bronchial tree. Cellular host defense mechanisms that can be impaired in patients with CB include the ability to phagocytose (engulf) bacteria by leukocytes (cells involved in host defense), the ability of the leukocytes to exert a bactericidal effect, and the excretion of macrophages and immunoglobulin A (IgA) in the sputum when infection is present (Wilson R, 2000). Repeated injury from high-dose inhalation of atmospheric pollutants or tobacco smoke leads to mucus hypersecretion, a change in the morphology of the airway (involving loss of ciliated cells, an increase in the number of nonciliated goblet cells, and mucosal gland hypertrophy), and the inÞltration of chronic inßammatory cells (lymphocytes, monocytes, and macrophages) into the mucosa. Most experts now believe that an acute bacterial infection of the bronchi results in neutrophil inÞltration into the bronchial lumen; inßammatory mediators then cause subsequent damage to the bronchial mucosa (Chodosh S, 1991). The by-products that the neutrophils release further impair host defenses. In the presence of chronic inßammation owing to an overwhelming number of by-products, these secretions can damage cilia and epithelial cells, stimulate mucus production, impair opsonization, and attract more neutrophils into the airway (Wilson R, 2000). Epithelial cells and other inßammatory cell types (e.g., CD4+, CD8+, macrophages) are also probably essential in the initial inßammatory process leading to the breakdown of lung tissue (Cosio MG, 2002). Many proinßammatory mediators can be measured in human secretions and serve as markers of infection. Also, patients with three or more exacerbations per year have higher baseline sputum cytokine IL-6 and IL-8 levels when stable, suggesting that frequent exacerbations are associated with increased inßammatory airway changes (Bhowmik A, 2000). Understanding the differences in the inßammatory cell types present in chronic airway diseases is a heated area of research and may be predictive of the various pathophysiological mechanisms associated with these diseases. Stimulation and Evasion of the Host Defenses. The abundance of neutrophils in sputum collected from CB or COPD patients during an exacerbation suggests a primary or secondary bacterial infection. Increases in bacterial ßora without increases in neutrophil cell count are more suggestive of a bacterial colonization rather than an acute exacerbation. As such, an increase in cough/sputum production alone in CB or COPD patients is not indicative of an infectious exacerbation. However, assessment of the sputum’s cell populations and bacterial load (through the use of Gram’s stain) can usually identify the infectious agent causing the exacerbation (Madison JM, 1998). Bacterial products themselves serve as potential independent stimuli of inßammation. Bacterial lipopolysaccharide (LPS), commonly found as surface antigen in gram-negative bacteria, is a potent stimulant for a variety of inßammatory mediators and immunoregulatory cytokines from resident and inßammatory cells in the lungs, including tumor necrosis factor alpha (TNFα), prostaglandins, leukotrienes, interleukins, platelet-activating factor, superoxide anion, hydrogen

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peroxide, and nitric oxide. Studies report that the LPS of H. inßuenzae significantly increased the release of IL-6, IL-8, and soluble intracellular adhesion molecule by cultured bronchial epithelial cells in vitro (Wilson R, 2000). Weakened host defenses in the chronic bronchitic airway (e.g., impaired mucociliary clearance) allow bacteria to exploit the local host environment by using mechanisms that evade the remaining host defenses (Wilson R, 1996[a]). Tactics include the following: • • • • • • •

Production of factors that impair mucociliary clearance by paralyzing ciliary beat and stimulating mucus production. Production of enzymes that break down local immunoglobulin. Antigenic heterogeneity to avoid immune surveillance. Growth in bioÞlms. Adherence to epithelium. Survival within epithelial cells. Formation of microcolonies surrounded by a polysaccharide gel.

As a result of immune evasion, bacteria are able to persist in the LRT. Also, the serum and sputum of patients with CB or COPD contain abundant antibodies to antigens of the colonizing bacteria. As such, the antibody response is often in subneutralizing quantities, blocking effective opsonization (the process by which bacteria are altered in such a manner that they are more readily and more efÞciently engulfed by phagocytes) and subsequent clearance of the pathogen from the LRT. When opsonization of bacteria by the appropriate antibodies is successful, the formation of immune complexes stimulates other inßammatory processes that escalate the inßammatory reaction, further destroying the local architecture in the lungs. Clinical Presentation of AECB. Clinical symptoms of a patient with acute exacerbation include increased severity of cough, increased sputum production, and purulent sputum–often associated with chest congestion and chest discomfort. Dyspnea and wheezing are present only when the patient has signiÞcant underlying pulmonary dysfunction. In addition, a patient may complain of malaise, fever, and chills. Patients may only have a few of these symptoms at presentation. Physical examination of a patient with an acute exacerbation may include the following Þndings: coarse rales, wheezes, decreased breath sounds, and tachycardia. The presence of these signs does not help in distinguishing which etiology is responsible for the speciÞc acute exacerbation (Chodosh S, 1992). Complications of AECB. The complications of AECB involve respiratory decompensation and side effects of infection. Patients with AECB can decline into respiratory failure if the increase in the work of breathing cannot be maintained. Such cases require immediate ventilatory support. Repeated AECB can accelerate permanent structural damage in the lung. Further, infection in AECB

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can progress to pneumonia and sepsis. Equally important complications of AECB are the effects the disease has on comorbid conditions. AECB can have a dramatic destabilizing impact on comorbid conditions, such as diabetes, congestive heart failure (CHF), and chronic renal failure, that may otherwise be well controlled. Risk Factors. The main risk factors for AECB include age and pulmonary diseases that increase inßammation and mucus production and impair airway clearance. Age is a risk factor because chronic cough and mucus production tend to increase with advancing age. In addition to the elderly, people at increased risk include those with other types of pulmonary disorders, such as asthma, and people with compromised immune systems. Cigarette smoking is a factor in up to 90% of CB cases, and research suggests smokers already suffering from CB are also more vulnerable to AECBs (Ball P, 2000; Sethi S, 1999). Other known risk factors for CB and AECBs are occupational exposure to dust, pollution (especially sulfur dioxide), and a history of childhood respiratory infection. Comorbidities of Patients with AECB. Patients with AECB and underlying CB often have important comorbidities and may be taking medications that impact medical management and antibiotic selection. They typically have a chronic history of smoking and therefore have subsequent development of obstructive lung disease. Comorbid conditions commonly include coronary artery disease (CAD), CHF, diabetes mellitus, chronic renal failure, and chronic liver disease. AECB may exacerbate other underlying comorbidities such as CHF and result in hospitalization. Treatment of AECB is particularly complicated because of the multiple (and often related) comorbidities. For example, an AECB episode can decompensate patients with otherwise stable CHF or CAD and require them to be hospitalized. Complications and comorbidities associated with AECB have a large impact on overall cost of therapy (the treatment costs associated with AECB are largely due to hospitalization) and on the choice of therapy to minimize drug interactions. The presence of comorbid conditions can increase hospitalization (thereby increasing cost of treatment) and the risk of treatment failure (Niederman MS, 1999). Antibiotic Resistance. The increase in antibiotic-resistant organisms has compromised the empiric use of certain antibiotics in the management of AECB. Studies have shown that resistance in the community of the main causative pathogens in AECB—H. inßuenzae, M. catarrhalis, and S. pneumoniae —has increased signiÞcantly over time (Doern GV, 1995; Brueggemann AB, 2000). Patients with severe lung disease are more likely to harbor pathogens (e.g., P. aeruginosa) that are resistant to Þrst-line antibiotics and therefore may be more likely to fail such therapy. Table 3 shows the prevalence of causative pathogens’ resistance to various antibiotics used commonly to treat AECB. Despite worrisome in vitro data, the clinical impact of antibiotic resistance remains unclear in patients with bronchitis (Gotfried MH, 2000). To date, studies

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ACUTE EXACERBATIONS OF CHRONIC BRONCHITIS

TABLE 3. Prevalence of Resistance to Commonly Prescribed Antibiotics Among H. Influenzae, M. Catarrhalis, and S. Pneumoniae Isolates in the Major Pharmaceutical Markets Prevalence of Resistance (%) United States Haemophilus influenzae Amoxi0.2–4.5 cillin/clavulanate Cefaclor 13 Cefuroxime 1 Ceftriaxone 0 TMP/SMX 9–16 Tetracyclines 1 Azithromycin 0.5 Clarithromycin 0.3–4 Levofloxacin 0 Moraxella catarrhalis Amoxi0 cillin/clavulanate Cefaclor 0.2 Cefuroxime 0 Ceftriaxone 0 TMP/SMX 0.2 Tetracyclines 0 Azithromycin 0 Clarithromycin 0 Fluoroquinolones 0

France

Germany

Italy

Spain

United Kingdom

Japan

0

0

0

0.5

0

0

N.A. 0 0 7–17 N.A. 0.0 5 0

N.A. 0 0 19 N.A. 0.0 5 0

N.A. 0 0 11–22 N.A. 0.0 3 0

N.A. 0 0 35–52 N.A. 0.3 9 0

N.A. 0 0 6–21 N.A. 0.0 5 0

N.A. 0 0 18 N.A. 0.0 7 0

0

0

0

0.5

0

0

N.A. 0 0 0 N.A. 0 0 0

N.A. 0 0 0 N.A. 0 0 0

N.A. 0 0 0 N.A. 0 0 0

N.A. 0 0 0 N.A. 0 0 0

N.A. 0 0 0 N.A. 0 0 0

N.A. 0 0 0 N.A. 0 0 0

Streptococcus pneumoniae Penicillin Intermediatea High-levelb Amoxicillin/clavulanate Intermediate High-level Cefuroxime Ceftriaxone TMP/SMX Tetracyclines Macrolides Levofloxacin

18 16

33 33

7 1

12 5

41 37

6 5

44 10

8 14 27 4 29 >20 23 0.5

20 23 55 5 14 >20 58 0

1.5 0 1 0 4

1.5 3 5 0.5 4–30 30 24 0

22 26 47 8 22 >20 37 0

3 1.5 9 0 2 N.A. 9 0

10 2 37 2 2 >20 66 1.0

9 0.4

a Intermediate resistance = Minimum inhibitory concentration [MIC] 0.12–1.0 mg/L. b High-level resistance = MIC 2 mg/L.

N.A. = Not available. TMP/SMX = Trimethoprim/sulfamethoxazole (co-trimoxazole). Note: Data reflect rounding.

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have not demonstrated clearly that patients infected with resistant strains have worse outcomes than similar patients infected with susceptible strains. However, indirect evidence supporting this concept has been published (Adams SG, 2000). More data are needed to clarify whether outcomes are worse in patients with lung infection caused by resistant organisms. If in vitro resistance is associated with a poor disease prognosis, then the data become even more signiÞcant. Because the emergence of resistance is directly related to antibiotic use and selective pressure, an evidence-based approach to caring for patients today may help ensure that efÞcacious therapy is available in the future. CURRENT THERAPIES Overview Numerous highly effective pharmacological agents are available to treat acute exacerbations of chronic bronchitis (AECB). Table 4 summarizes the general pharmacological agents and classes used to manage AECB. The primary therapies used in AECB treat the causative infection (antibiotics), relieve symptoms (bronchodilators), and treat the underlying inßammation (corticosteroids). Table 5 summarizes the leading antibiotic therapies used to treat the infection. Most AECB cases require empiric antibiotic therapy because of the difÞculty in deÞnitively distinguishing exacerbations of bacterial etiology from those of nonbacterial origin. Empiric therapy is based on likely infecting pathogens, local resistance patterns, severity of the exacerbation, and patient history.

TABLE 4. Pharmacological Management of Underlying Disease During an Acute Exacerbation of Chronic Bronchitis Therapy Bronchodilators (e.g., inhaled beta2 agonists, inhaled anticholinergics, theophyllinea ) Corticosteroids

Mucolytics/antitussives Oxygen therapy NPPV or mechanical ventilation

Comment Administer as soon as possible during an exacerbation. Increase usual dosage. Consider nebulizer for delivery. Oral, inhaled, or intravenous administration. Short courses may provide benefit. Usually given only in hospitalized patients. May help facilitate clearance of airways. Administer if PaO2 is less than 55 mg Hg. Beneficial support strategy for hospitalized patients.

a Potentially toxic; not as effective as inhaled bronchodilators; monitoring is important especially when

coadministered with antibiotics. Hg = Mercury. NPPV = Noninvasive positive-pressure ventilation. PaO2 = Partial pressure of arterial oxygen. Note: When patients come in for treatment of acute exacerbation of chronic bronchitis (AECB), most if not all of the patients will already be on short-acting and long-acting bronchodilators, anticholinergics, and inhaled corticosteroid-containing products to treat their underlying lung condition, which can vary in severity.

454

TABLE 5. Current Therapies Used for Acute Exacerbations of Chronic Bronchitis, 2004 Agent Penicillins Amoxicillin Amoxicillin/clavulanate Piperacillin/tazobactam Ampicillin/sulbactam Ticarcillin/clavulanate Cephalosporins Cefuroxime axetil Cefprozil Cefpodoxime proxetil Cefotaxime Ceftriaxone Cefdinir Carbapenems Imipenem/cilastatin Meropenem Ertapenem Macrolides Erythromycin Clarithromycin Azithromycin Roxithromycin

Company/Brand

Daily Dose

Availability

GlaxoSmithKline’s (GSK’s) Amoxil, generics GSK’s Augmentin, Augmentin ES, Augmentin XR, generics Wyeth/Taisho’s Zosyn/Tazocin Pfizer’s Unasyn, generics GSK’s Timentin

500–1000 mg PO q8h or 875 mg PO q12h 500/125 mg PO q8h or 875/125 mg PO q12h or 2,000/125 mg PO q12h 3.375 mg IV q6h 1,000–3,000 mg IV q6h 3,000/100 mg IV q6h

US, F, G, I, S, UK, J

GSK’s Ceftin/Zinnat/Oracef Bristol-Myers Squibb’s (BMS’s) Cefzil/Procef/Bisoral Pfizer’s Vantin, Sanofi-Aventis’s Orelox, Sankyo’s Banan Abbott/Sanofi-Aventis’ Claforan, generics Roche’s Rocephin, generics Abbott’s Omincef, Fujisawa’s Cefzon

250–500 mg IV q12h 500 mg PO q12h

US, F, G, I, S, UK, J US, I, S, UK

200 mg PO q12h

US, F, G, I, S, UK, J

1–2 g IV q8h

US, F, G, I, S, UK, J

1–2 g IV qd in divided doses q12h 300 mg PO q12h

US, F, G, I, S, UK, J US, J

Merck’s Primaxin AstraZeneca’s Merrem Merck’s Invanz

1–2 g IV q6h 1 g IV q8h 1 g IV qd

US, F, G, I, S, UK, J US, F, G, I, S, UK, J US, G, S, UK

Abbott’s Erythrocin, generics Abbott’s Biaxin/Biaxin XL/Klacid, Taisho’s Clarith Pfizer’s Zithromax Sanofi-Aventis’ Rulid, generics

250 mg PO q6h 250–500 mg PO q12h, or 1,000 mg PO qd 420 mg PO loading, then 250 mg PO qd 150 mg PO bid

US, F, G, I, S, UK, J US, F, G, I, S, UK, J

US, F, G, I, S, UK, J US, F, G, I, S, UK, J US, F, G, I, S, UK, J US, F, I, UK

US, F, G, I, S, UK, J F, G, I, S, UK, J

TABLE 5. (continued) Agent Ketolides Telithromycin Fluoroquinolones Levofloxacin

Moxifloxacin Gatifloxacin Tetracyclines Doxycycline Minocycline Glycopeptides Vancomycin Teicoplanin

Company/Brand

Daily Dose

Availability

Sanofi-Aventis/Fujisawa/Sankyo’s Ketek

800 mg PO qd

US, F, G, I, S, UK, J

Johnson & Johnson’s Levaquin, Sanofi-Aventis’ Tavanic, Daiichi’s Cravit Schering-Plough/Bayer’s Avelox BMS’s Tequin, Kyorin/Dainippon’s Gatiflo

500 mg PO or IV qd

US, F, G, I, S, UK, J

400 mg PO or IV qd 400 mg PO or IV qd

US, F, G, I, S, UK US, G, J

Pfizer’s Vibramycin, generics Wyeth’s Minocin, generics

200 mg PO loading, then 100 mg PO qd 100 mg PO q12h

US, F, G, I, S, UK, J US, F, G, I, S, UK, J

Eli Lilly’s Vancocin, generics Sanofi-Aventis’ Targocid

500 mg IV q6h 400 mg IV loading, then 200 mg IV qd

US, F, G, I, S, UK, J F, G, I, S, UK, J

bid = Twice daily; q4h = Every 4 hours; q6h = Every 6 hours; q8h = Every 8 hours; q12h = Every 12 hours; qd = Once daily. IV = Intravenous formulation. PO = Oral formulation. US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan.

455

456

ACUTE EXACERBATIONS OF CHRONIC BRONCHITIS

This section focuses on the antibiotic therapies used to treat AECB. In addition to the available agents to treat the underlying lung condition and infectious episodes of bronchitis, vaccination against the causative pathogens of AECB has been debated. However, the value of pneumococcal vaccination in patients with chronic bronchitis is not well established; it is not clear whether vaccination prevents AECB in these patients. However, pneumococcal vaccination is safe and can reduce invasive pneumococcal infection, and current recommendations favor vaccinations in all patients with high risk of developing exacerbations (i.e., patients with prior history of underlying chronic pulmonary obstruction). Choice of Antibiotic Therapy. Comparisons of clinical efÞcacy across major classes of antibacterials suggest that many compounds achieve comparable clinical efÞcacy in resolving infection. Indeed, most clinical trials demonstrate clinical equivalence rather than superiority of one agent over another. Clinical trials report both clinical response and bacterial end points achieved by an antibacterial agent. The clinical response suggests an improvement in the symptoms of the disease (e.g., reduction in fever) achieved by the agent. Bacteriological end points indicate pathogen eradication achieved by the agent. The susceptibility of bacterial strains to antibacterials is reported as sensitive, intermediately sensitive/resistant, or resistant based on the minimum inhibitory concentration (MIC) required by the antibacterial to exert its effect. An important factor in antibiotic choice is oral bioavailability. For serious infections, physicians need to ensure that patients achieve a high serum concentration at the site of infection. Antibiotics with poor oral bioavailability have difÞculty achieving this high serum concentration because much of the drug does not cross the gastrointestinal (GI) tract into the bloodstream, and physicians must choose the intravenous (IV) formulation instead. Antibiotics with a high oral bioavailability, however, offer physicians the ability to treat more serious infections in an outpatient setting knowing that these therapies will achieve high serum concentrations. Likewise, for patients who are started on IV antibiotics in the hospital, agents with high bioavailability can facilitate the switch from IV to oral therapy and result in a shorter hospitalization. Physician choice of antibiotic therapy for AECB is driven by the activity, spectrum of coverage, safety, dosing, and drug interactions. Since therapy is typically empiric, without deÞnitive identiÞcation of the pathogen, coverage of the key respiratory pathogens (particularly Haemophilus inßuenzae) is a critical factor in drug selection. Likewise, because patients with AECB often suffer from other comorbidities (particularly cardiopulmonary diseases such as coronary artery disease [CAD] and congestive heart failure [CHF]) and are frequently on numerous other medications, safety and lack of signiÞcant drug interactions are important in the construction of a safe, effective drug regimen. Antibiotic Resistance. Pathogen resistance to an antibiotic reduces the efÞcacy of the agent and can endanger the life of the patient. Resistance is more likely to occur if a patient has been recently treated with an antibiotic (as can be the case in chronic obstructive pulmonary disease [COPD] patients suffering

CURRENT THERAPIES

457

TABLE 6. Emerging Therapies in Development for Acute Exacerbations of Chronic Bronchitis, 2004 Compound

Development Phase

Fluoroquinolones Garenoxacin United States Europe Japan Dihydrofolate reductase inhibitors Iclaprim United States Europe Japan

Marketing Company

PR III III

Schering-Plough Schering-Plough Toyama

— IIa —

— Arpida —

a For skin infections.

R = Registered; S = Suspended; PC = Preclinical.

frequent exacerbations). From a public health perspective, antibacterial resistance can occur after the antibacterials have been used (or overprescribed) in the community for a period of time. Therefore, the efÞcacy of a given antibacterial and the susceptibility of pathogens in the community may change over time as usage patterns change. Indeed, the prevalence of resistance exhibits geographic variation, which researchers believe is due to the variable volume of prescribing. In recent years, antibiotic resistance has become an increasingly important factor inßuencing prescribing choice. During the last 10–15 years, a steady rise in the frequency of antibacterialresistant pathogens, such as β-lactamase-producing H. inßuenzae and Moraxella catarrhalis (pathogens predominantly observed in AECB episodes), strains of penicillin-resistant Streptococcus pneumoniae (Jacobs MR, 2004), and erythromycin-resistant S. pneumoniae, has been observed. This rise has limited the utility of older antibacterials, such as penicillin and early-generation cephalosporins, and spurred the need for new agents. Fortunately, the pharmaceutical industry has developed newer agents over time, with expanded activity against resistant pathogens. However, bacterial drug resistance is an inherent factor in this market and will continue to be the primary driver for innovative new antibacterial drugs. Penicillins Overview. The penicillins comprise several subgroups of agents with a wide range of bacterial coverage and efÞcacy. Each penicillin molecule contains a basic β-lactam structure fused to a Þve-membered ring. Because of their broad spectrum of activity and availability in oral form, the penicillins are commonly used in the treatment of AECB and have become the drugs of choice in treating many common infections.

458

ACUTE EXACERBATIONS OF CHRONIC BRONCHITIS

The penicillins are further divided into the following groups: natural penicillins, aminopenicillins, and the extended-spectrum penicillins. The natural penicillins (e.g., penicillin G) have the narrowest spectrum. They are active only against the gram-positive cocci and, with regard to AECB, active essentially only against S. pneumoniae. The aminopenicillins (such as ampicillin and amoxicillin) are natural penicillin derivatives with an expanded spectrum of activity that includes some gramnegative organisms that do not produce β-lactamases–enzymes that destroys the ring structure of the antibiotic rendering it ineffective—including H. inßuenzae. The aminopenicillins have been in use since the 1970s, and they continue to be valuable therapeutic agents for the treatment of many bacterial infections. These drugs are sometimes used as Þrst-line agents in the treatment of mild-tomoderate AECB when no other risk factors are apparent. However, the usefulness of aminopenicillins is increasingly limited in countries such as France and Spain because of resistance among the common respiratory pathogens but they continue to be used owing to their inexpensiveness. The extended-spectrum penicillins (such as pipericillin and ticarcillin) are semisynthetic and have a broad range of activity including many gram-negative organisms, such as Escherichia coli and K. pneumoniae. These drugs are more active than natural penicillins and aminopenicillins because they are more resistant to inactivation by β-lactamases and/or because they more readily penetrate the outer membranes of gram-negative bacilli. Nonetheless, their rate of bactericidal action and the completeness of this effect can be inconsistent. As a class, the penicillins are among the safest antibiotics. However, one potentially life-threatening adverse effect associated with their use is immediate or delayed hypersensitivity reactions in a small percentage of patients. These reactions can present in a variety of forms, such as mild skin rash, interstitial nephritis, hematological disturbances (e.g., neutropenia), vasculitis, and anaphylactic shock. The overall incidence of hypersensitivity reactions ranges between 0.7% and 10% of patients treated with penicillins. Other more common, yet minor, side effects caused by penicillins are GI disturbances (e.g., nausea, vomiting, diarrhea) (Weiss ME, 1988). A growing concern over the past two decades has been the development of bacterial resistance to the penicillins—particularly in penicillin-resistant S. pneumoniae (PRSP) (Jacobs MR, 2004). Penicillin drug resistance is caused by altered penicillin binding proteins (PBPs) in the cell wall of resistant organisms that result in decreased afÞnity of the antibacterials to their targets. Resistance to penicillins (as well as their dosing schedule and frequency of GI side effects) has resulted in switching to newer-generation drugs such as the ßuoroquinolones, which have better activity against resistant strains of pathogens. In many cases, however, physicians are instead employing higher doses of penicillins and/or using penicillin/penicillinase inhibitors to provide adequate coverage and overcome potential resistance issues. Ticarcillin/clavulanate (GlaxoSmithKline’s Timentin) is an injectable extended-spectrum penicillin/beta-lactamase inhibitor used in severe, complicated

CURRENT THERAPIES

459

hospitalized cases of AECB. For AECB, it is used in similar settings as pipericillin/tazobactam (Wyeth/Taisho’s Zosyn) and will not be discussed in greater detail because of its limited use. Likewise, ampicillin/sulbactam (PÞzer’s Unasyn, generics), an injectable aminopenicillin/beta-lactamase inhibitor that has activity against gram-positive and some gram-negative organisms similar to amoxicillin/clavulanate, will not be discussed further. Mechanism of Action. Penicillins and related β-lactam antibacterials bind to PBPs in bacteria and prevent bacterial cell-wall formation. The PBPs are enzymes that reside on the inner wall of the bacterial cell membrane and serve to maintain cell-wall homeostasis, including structural integrity, cell shape, cell division, capsule synthesis, phage resistance, and regulation of autolysis. The β-lactam ring of penicillins interacts with PBPs and inhibits cell-wall synthesis, thereby killing the cell. Most bacteria contain between four and eight different PBPs; hence antibiotics against the PBPs have to target several different proteins. Organisms that are metabolically inactive and those lacking bacterial cell walls (e.g., Mycoplasma) are not susceptible to β-lactam antibacterials. Amoxicillin. Amoxicillin (GlaxoSmithKline’s Amoxil, generics) (Figure 3) is available in a capsule, chewable tablet, or oral suspension. It is an amino penicillin derivative that is active against gram-positive organisms and some gram-negative strains that do not produce β-lactamases, such as H. inßuenzae, but not K. pneumoniae or P. aeruginosa. Amoxicillin has been commercially available since 1974 and continues to be a valuable therapeutic agent for the treatment of many bacterial infections. As a β-lactam antibiotic, amoxicillin binds to PBPs in bacteria and prevents bacterial cell-wall formation. Amoxicillin is not often used for AECB because of the rising prevalence of β-lactamase-producing H. inßuenzae. In a randomized multicenter trial, 143 subjects with AECB were treated with 250 mg of cefuroxime axetil (GlaxoSmithKline’s Ceftin/Zinnat/Oracef) twice daily and 153 subjects received 250 mg of amoxicillin three times daily. Results showed that 76.2% of subjects treated with cefuroxime axetil were cured or saw clinical improvement compared with 80.4% in the amoxicillin arm (Shah SH, 1994). Side effects associated with amoxicillin are minimal, with some incidence of diarrhea. There are instances of skin rashes, and particularly delayed responses. Amoxicillin is typically well absorbed from the GI tract, has a rapid onset and long duration of action, requires less frequent dosing, and has a favorable H COOH O H HO

C NH2

CH3

N CONH

S H

CH3

H

FIGURE 3. Structure of amoxicillin.

3 H2O

460

ACUTE EXACERBATIONS OF CHRONIC BRONCHITIS

side-effect proÞle. As a generically available product, amoxicillin is also relatively inexpensive. Unfortunately, bacterial resistance to amoxicillin has increased signiÞcantly, both among H. inßuenzae and S. pneumoniae, resulting in a switch toward newer-generation drugs that are active against these pathogens. Amoxicillin/Clavulanate. Amoxicillin/clavulanate (GlaxoSmithKline’s Augmentin, Augmentin ES, Augmentin XR, generics) is a combination of the semisynthetic antibiotic amoxicillin and the β-lactamase inhibitor clavulanic acid. The product is available in immediate-release tablets, extra-strength and extended-release tablets, oral suspension, chewable tablets, and parenteral form (in Europe only). GlaxoSmithKline is attempting to retain sales of its amoxicillin/clavulanate franchise with the branded Augmentin XR and ES formulations, following the market entry of generic competitors to Augmentin in 2002. The drug has a broad spectrum of bactericidal activity against many gram-positive and gram-negative microorganisms and has been a favorite among clinicians because of its efÞcacy, safety, and familiarity. The formulation of amoxicillin and clavulanic acid in the combination drug protects amoxicillin from degradation by β-lactamase enzymes that bacteria might release and effectively extends the antibiotic spectrum of amoxicillin to include many bacteria normally resistant to amoxicillin and other β-lactam antibiotics. The antibiotic component functions by disrupting bacterial cell-wall synthesis. A clinical trial compared the efÞcacy of amoxicillin/clavulanate (250/125 mg three times daily for ten days) with ceftibuten (Schering-Plough’s Cedax, generics), a third-generation cephalosporin, 400 mg once daily for Þve days in 335 adults with AECB (Guest N, 1998). Clinical response was equivalent between the two arms, with cure or improvement in 92.4% of ceftibuten-treated patients and 92.7% of amoxicillin/clavulanate-treated patients. The incidence of adverse events was also comparable, in approximately 15% of patients. Amoxicillin/clavulanate’s efÞcacy, safety, and dosing may account for the longevity of this combination in clinical use. However, in certain deÞned geographical areas, the emergence of S. pneumoniae strains with elevated MICs (which indicate lower susceptibility to the agent) has been observed. To meet the need of treating drug-resistant S. pneumoniae, high-dose amoxicillin/clavulanate formulations have been developed that are active against some resistant bacterial strains. An enhanced dosage form of amoxicillin/clavulanate 2000/125 mg twice daily (GlaxoSmithKline’s Augmentin XR) has been developed for use in adult respiratory tract infection. This enhanced formulation prolongs the time that bacteria are exposed to the antibiotic and promotes coverage of tough-to-treat S. pneumoniae with reduced susceptibility to penicillin, as well as β-lactamaseproducing H. inßuenzae and M. catarrhalis. Amoxicillin/clavulanate is generally well tolerated. The most frequently reported adverse effects as reported in the product insert were diarrhea/loose stools (9%), nausea (3%), skin rashes and urticaria (3%), vomiting (1%), and vaginitis (1%). The overall incidence of side effects, and in particular diarrhea, increased with the higher recommended dose. In clinical practice, diarrhea and

CURRENT THERAPIES

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loose stools are the most frequently reported side effects, although these tolerability issues can be minimized by taking the product with food and rarely result in discontinuation of treatment. Other less frequently reported reactions include abdominal discomfort, ßatulence, and headache. Piperacillin/Tazobactam. Piperacillin/tazobactam (Wyeth/Taisho’s Zosyn) combines the extended-spectrum penicillin piperacillin (Figure 4) with the βlactamase inhibitor tazobactam (Figure 5). Commercially available since 1993, piperacillin/tazobactam is available for parenteral administration only. U.S. patent expiry is expected in 2007. As a β-lactam/β-lactamase inhibitor combination therapy, the tazobactam component protects the antibiotic from degradation by β-lactamase enzymes released by bacteria, thereby extending the effective spectrum of the therapy to treat bacteria resistant to the β-lactam antibiotic component (piperacillin). Like other β-lactam antibiotics, piperacillin/tazobactam achieves its bactericidal effect by interacting with PBPs and disrupting bacterial cell-wall formation. Piperacillin’s broad range of activity (which includes many gram-negative organisms such as P. aeruginosa and K. pneumoniae) is enhanced by the β-lactamase inhibitor tazobactam, effectively extending the spectrum of activity to include H. inßuenzae. Piperacillin/tazobactam is used for severe and/or complicated AECB episodes in hospitalized patients in whom broad (gram-negative) coverage is required. H COONa O H

CH3

N

C

CONH

NHCO

S H

N

O

N

O

CH3

H

C2H5 FIGURE 4. Structure of piperacillin sodium.

FIGURE 5. Structure of tazobactam (R = R1 = H).

462

ACUTE EXACERBATIONS OF CHRONIC BRONCHITIS

A Japanese multicenter comparative study of the use of piperacillin versus piperacillin/tazobactam in chronic respiratory infections demonstrated comparable clinical success between these therapies. The study enrolled a total of 173 patients with chronic RTIs. Eighty-Þve study subjects were treated with 2 grams of piperacillin IV twice daily for 14 days, and 88 subjects were treated with piperacillin/tazobactam (2 grams/500 mg) combination therapy for 14 days. The clinical response was comparable, with 81% and 86% of subjects achieving favorable outcomes, respectively (Oizumi K, 1995). In clinical trials of piperacillin/tazobactam, 90% of the adverse reactions were transient and mild to moderate in severity. The most signiÞcant side effects reported include skin rashes/pruritus, GI complications (including diarrhea, nausea, and vomiting), and allergic reactions (in 1.3%, 0.9%, and 0.5% of treated patients, respectively) (Oizumi K, 1995). Cephalosporins Overview. The cephalosporins contain a basic β-lactam structure fused to a six-membered ring. Drugs in this class differ widely in their spectrum of activity, susceptibility to β-lactamases produced by bacteria, and serum half-life. Cephalosporins are categorized into four generations, with each newer generation representing an improvement in the spectrum of bacterial coverage. First-generation agents have the narrowest spectrum of activity among the cephalosporins. They are most active against staphylococci and streptococci organisms. Most Þrst-generation cephalosporins are oral formulations. Secondgeneration agents have increased activity to cover more gram-negative bacilli, but are usually less active than the Þrst-generation drugs against gram-positive bacteria. Second-generation oral cephalosporins are occasionally used to treat mild episodes of AECB. Third-generation cephalosporins are active against gram-negative organisms. However, their activity against gram-positive organisms is inferior to that of previous generations. These agents are commonly recommended in clinical guidelines for Þrst-line treatment of patients hospitalized with AECB. Compared with second-generation cephalosporins, the third-generation agents have greater stability against β-lactamases and have longer serum half-lives. As a result, they have more-convenient dosing regimens. The third-generation agents may also be used in combination with macrolides, extended-spectrum penicillins, or aminoglycosides to treat severe AECB. Fourth-generation agents have enhanced stability against β-lactamases and provide good coverage of both gram-positive and gram-negative bacteria (particularly P. aeruginosa). However, they are generally reserved for severe, life-threatening infections such as sepsis. As a class, the cephalosporins are generally well tolerated (Gustaferro CA, 1991; Okamoto MP, 1994). Common adverse effects are usually minor; GI disturbances and thrombophlebitis are the most prominent with oral and intravenous (IV) agents, respectively. Disturbances of the GI tract are reported less often with cephalosporins than with the penicillins. Because of their relative safety and broad

CURRENT THERAPIES

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spectrum of activity, cephalosporins are commonly used to treat suspected as well as and proven bacterial infections. Resistance against cephalosporins, as with other β-lactam antibiotics, results from pathogen changes in outer-membrane permeability, stability against βlactamases, and modiÞcation of PBPs. While β-lactamase production by H. inßuenzae or M. catarrhalis limits the use of certain penicillins such as amoxicillin, many cephalosporins are effective in treating infections caused by these β-lactamase-producing bacteria. Resistance to third-generation cephalosporins in gram-negative pathogens is a formidable problem in the hospital setting and is associated with adverse clinical outcomes and increased hospital costs (Cosgrove SE, 2002). Mechanism of Action. As with the penicillins, the β-lactam ring of cephalosporins binds to PBPs in bacteria and prevents bacterial cell-wall formation. By interrupting cell-wall formation, cephalosporins induce cell lysis and death. Cefuroxime Axetil. Cefuroxime axetil (GlaxoSmithKline’s Ceftin/Zinnat/ Oracef) ∗ is a second-generation oral cephalosporin consisting of an esteriÞed pro-drug of cefuroxime; the active antibiotic metabolite cefuroxime is released following cefuroxime axetil absorption from the GI tract. First launched in the United States in 1988, cefuroxime axetil was available as a U.S. generic by 2002. This product is active against a broad spectrum of gram-positive and gramnegative bacteria, including S. pneumoniae, H. inßuenza, K. pneumoniae, and M. catarrhalis. Cefuroxime axetil is indicated for a number of bacterial infections including respiratory tract infections caused by β-lactamase-negative strains of S. pneumoniae, H. inßuenzae, M. catarrhalis, and Haemophilus parainßuenzae. The agent’s efÞcacy in AECB was demonstrated in a randomized multicenter trial comparing the agent with amoxicillin. In the trial, 143 subjects with AECB were treated with 250 mg of cefuroxime axetil twice daily and 153 subjects received 250 mg of amoxicillin three times daily. Results showed that 76.2% of subjects treated with cefuroxime axetil were cured or saw clinical improvement compared with 80.4% in the amoxicillin arm (Shah SH, 1994). In clinical trials of cefuroxime axetil, patients who received 7–10 days of therapy experienced adverse reactions similar in type and frequency to those of other antibiotic agents. Diarrhea (3.7%) and nausea/vomiting (3.0%) were among the most common reactions in trials with adults receiving tablets as reported in product inserts. Cefprozil. Cefprozil (Bristol-Myers Squibb’s Cefzil/Procef/Bisoral) is a secondgeneration oral cephalosporin. The agent has been available in the Untied States since 1992 and is also marketed in Italy, Spain, and the United Kingdom. The product is expected to have patent protection until 2006 in the United States and for a period ranging from 2004 to 2008 in the European markets. Cefprozil is

464

ACUTE EXACERBATIONS OF CHRONIC BRONCHITIS

active against gram-positive and gram-negative bacteria, but not Pseudomonas species. In clinical studies, this cephalosporin was proven active against S. pneumoniae, H. inßuenzae, and M. catarrhalis. In vitro tests have shown cefprozil is also effective against a variety of other bacteria, including streptococci and gram-negative microbes. Cefprozil demonstrated efÞcacy comparable to that of cefaclor (Eli Lilly’s Ceclor, generics) in the treatment of 247 elderly patients with bronchitis. Of 156 subjects who received cefprozil, 84% of those clinically evaluable achieved favorable clinical outcomes, a Þgure comparable to the 82% of patients achieving favorable outcomes in the cefaclor-treated group. In clinical trials for cefprozil, side effects included nausea, diarrhea, vomiting, and abdominal pain (Pelletier LL Jr, 1994). Cefpodoxime Proxetil. Cefpodoxime proxetil (PÞzer’s Vantin, SanoÞAventis’s Orelox, Sankyo’s Banan, generics) is an extended-spectrum, thirdgeneration oral cephalosporin. Sankyo originally developed cefpodoxime proxetil and Þrst launched the cephalosporin in Japan in 1989. In 1992, Upjohn (later Pharmacia) began marketing it in the United States; Aventis marketed the drug in Europe. A generic version of the product (oral suspension and tablets) by Ranbaxy is available in the United States. Cefpodoxime is indicated for the treatment of AECB caused by S. pneumoniae, M. catarrhalis, and H. inßuenzae (non-beta-lactamase-producing strains only). Cefpodoxime proxetil is an esteriÞed pro-drug of cefpodoxime that is stable in the presence of most β-lactamase enzymes, which effectively extends its spectrum of activity against a number of gram-positive and gram-negative bacteria resistant to penicillins and other cephalosporins. However, some extended-spectrum βlactamase enzymes can inactivate cephalosporins. Clinical studies have shown cefpodoxime to be active against S. pneumoniae, E. coli, K. pneumoniae, H. inßuenzae, and M. catarrhalis, and inactive Pseudomonas species. In a multicenter, double-blind, comparative trial of 251 patients with AECB, cefpodoxime proxetil was administered twice daily (at 200 mg dosage) for Þve to ten days to one group of subjects while the other group received 625 mg of amoxicillin/clavulanate three times daily. Clinical efÞcacies for the cefpodoxime proxetil and amoxicillin/clavulanate groups were 97.2% and 94.7%, respectively (Periti P, 1990). Additionally, a dosage of 100 mg or 200 mg of cefpodoxime twice daily demonstrated similar clinical and bacteriological efÞcacy to amoxicillin received in 250 mg doses three times daily. In clinical trials, cefpodoxime is well tolerated; patients reported experiencing nausea (3.3%), vaginal fungal infections (1.0%), vulvovaginal infections (1.3%), abdominal pain (1.2%), and headaches (1.0%), as reported in U.S. prescribing information in product inserts. Cefotaxime. Cefotaxime (Abbott/SanoÞ-Aventis’ Claforan, generics) (Figure 6) is a third-generation, parenteral cephalosporin available for IV or intramuscular (IM) administration. Cefotaxime was Þrst marketed in 1981 in the

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COONa OCH3 O N N

C

CH2OCOCH3

N CONH S H

H2N

H

S

FIGURE 6. Structure of cefotaxime.

United States, where generic versions of the injection are now available. The agent is also marketed by Roche in Japan. Cefotaxime is indicated for the treatment of lower respiratory tract infections caused by S. pneumoniae and other streptococci, E. coli, K. pneumoniae and other Klebsiella species, H. inßuenzae (including ampicillin-resistant strains). Cefotaxime is stable in the presence of many β-lactamase enzymes, which effectively extends its spectrum of activity against a number of gram-positive and gram-negative bacteria that are resistant to penicillins and other cephalosporins. Clinical and laboratory studies have demonstrated the activity of cefotaxime against S. pneumoniae and other streptococci. Gram-negative species susceptible to cefotaxime include E. coli, H. inßuenzae, H. parainßuenzae, K. pneumoniae, and others (SanoÞ-Aventis, 2004[b]). An early clinical trial examined the efÞcacy of cefotaxime in treating patients with severe respiratory infections including AECB and pneumonia. One hundred and ninety patients treated with 2.25–3.0 g of cefotaxime daily for an average of nine days displayed a favorable clinical response in 91% of 184 evaluable cases (Pines A, 1980). Another randomized, double-blind, comparative clinical trial evaluated cefodizime (Fujisawa’s Timecef/SanoÞ-Aventis’s Modivid), a third-generation cephalosporin, and cefotaxime in cases of AECB. One hundred and eighty hospitalized patients were treated twice daily for seven days with 1 g IM injections of either antibiotic. Bacteriological efÞcacy was demonstrated in 98.9% of cefodizime patients and in 95.6% of cefotaxime patients. Cefotaxime is generally well tolerated, but some adverse reactions have been documented, including local inßammation at point of IV administration (4.3%) and GI disturbances (1.4%). Systemic adverse reactions to cefotaxime have been rare. Ceftriaxone. Owing to its once-daily dosing, excellent potency, and spectrum of action, ceftriaxone (Roche’s Rocephin, generics) (Figure 7) is one of the most successful antibiotics on the market. The agent, Þrst launched in 1982, is a semisynthetic, third-generation, parenteral cephalosporin available for IV or IM administration. Ceftriaxone has been available generically in Europe since 2002. In 2000, Cubist Pharmaceuticals announced that it had acquired the rights to oral ceftriaxone and was developing an oral alternative formulation. However, the company discontinued development in 2004 after human clinical trials

466

ACUTE EXACERBATIONS OF CHRONIC BRONCHITIS

FIGURE 7. Structure of ceftriaxone.

showed highly variable bioavailability. Ceftriaxone is indicated for lower respiratory tract infections caused by S. pneumoniae,H. inßuenzae, H. parainßuenzae, and K. pneumoniae. Ceftriaxone has demonstrated a broad spectrum of activity against grampositive and gram-negative bacteria including S. pneumoniae, H. inßuenzae, H. parainßuenzae, K. pneumoniae, M. catarrhalis, and others (Roche Pharma, 2004). The efÞcacy of ceftriaxone in the treatment of lower respiratory tract infections was evaluated in a Phase III clinical study in 118 patients comparing either 1 gm of ceftriaxone or 1 gm cefonicid (GlaxoSmithKline’s Monocid), a secondgeneration cephalosporin, intravenously or intramuscularly, daily for 3 to 11 days (mean was 7 days). Clinical cure or improvement was noted in 93% and 95% of the patients treated with ceftriaxone and cefonicid, respectively. Bacteriologic cure or improvement was noted in 81% and 69% of cases, respectively. The agent is well tolerated in extensive clinical use, but local reactions have been reported (McCabe RE, 1989). Cefdinir. Cefdinir (Abbott’s Omnicef, Fujisawa’s Cefzon) is a third-generation, oral cephalosporin available in capsule and oral suspension forms. Cefdinir was developed and launched by Fujisawa in Japan in 1991 as Cefzon. WarnerLambert’s Parke-Davis (now PÞzer) was the Þrst to license cefdinir for the United States and Western Europe, and began marketing the product in the United States in 1998. Abbott obtained the exclusive marketing rights from Fujisawa and began marketing the drug as Omnicef in 2000. The agent is expected to retain patent protection through 2007 in the United States. Cefdinir is indicated for the treatment of AECB in adults and adolescents caused by penicillinase-susceptible S. pneumoniae and all strains of H. inßuenzae and M. catarrhalis. Cefdinir is stable in the presence of many β-lactamase enzymes, which effectively extends its spectrum of activity against a number of gram-positive and gram-negative bacteria resistant to penicillins and other cephalosporins. Cefdinir has demonstrated activity against S. pneumoniae, H. inßuenzae, K. pneumoniae, M. catarrhalis, and others (Abbott Labs, 2004). In a randomized, dose-comparative Phase III study enrolling 466 subjects with acute bronchitis, treatment with 600 mg of cefdinir once daily was compared with 300 mg of cefdinir twice daily for ten days. Clinical success in assessable

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patients receiving once-daily doses was 91%, and the bacteriologic eradication rate was 92%. In assessable patients receiving twice-daily doses, clinical success was 93%, and the bacteriologic eradication rate was 93% (Sperling MJ, 1996). Another trial compared the two cefdinir dosing regimens with cefuroxime axetil in the treatment of AECB. This double-blind, randomized, multicenter Phase III study enrolled 1,045 subjects and evaluated 589 patient outcomes for efÞcacy after ten days of therapy. Patients who received 600 mg once daily of cefdinir were clinically cured in 81% of cases; those who received cefdinir in 300 mg doses twice daily were cured in 74% of cases; and those who received 250 mg of cefuroxime axetil twice daily were cured in 80% of cases (Van Herwaarden CL, 2000). The Phase III comparative clinical trial showed that cefdinir had a statistically signiÞcant higher rate of adverse reactions (25.9% of 474 patients) than did cephalexin (Lilly’s Keßex, generics), a Þrst-generation cephalosporin (16.1% of 478 patients). In particular, cefdinir causes more GI discomfort (such as diarrhea) than do other cephalosporins. The most frequently reported adverse reactions in U.S. clinical trials with a total of 3,841 adult cefdinir-treated patients were diarrhea (15%), vaginal moniliasis (4% of women), nausea (3%), and headache (2%). Most adverse events were mild and self-limiting in nature with a low (3%) discontinuation rate due to associated adverse events. The most frequently reported adverse reactions in U.S. clinical trials with a total of 1,783 pediatric cefdinir-treated patients were diarrhea (8%), rash (3%), and vomiting (1%) (Tack KJ, 1998). Carbapenems Overview. Carbapenems are penicillin derivatives that have good activity against gram-positive and gram-negative aerobic and anaerobic bacteria. They are highly resistant to β-lactamase and have a very favorable spectrum of activity. A drawback of these agents is that they are available in IV form only. These agents are used in the treatment of severe, complicated AECB when sepsis is of concern. The incidence of adverse reaction is low (1%) and occurs predominantly among the elderly or patients with predisposing central nervous system conditions (Norrby SR, 2000). Carbapenem antibiotics include imipenem (Merck’s Primaxin) meropenem (AstraZeneca’s Merrem), and ertapenem (Merck’s Invanz). This section reviews only imipenem/cilastatin, as this class is generally used in just a small subset of AECB patients. Mechanism of Action. Like the penicillins, carbapenems bind to PBPs in bacteria and prevent bacterial cell-wall formation. By interrupting cell-wall formation, carbapenems induce cell lysis and death. Imipenem/cilastatin. Imipenem/cilastatin (Merck’s Primaxin) is a prototypical carbapenem. Imipenem is coadministered with cilastatin (a peptidase inhibitor) (Figure 8) to prevent its metabolism into nephrotoxic metabolic end products by peptidase enzymes in the body. Coadministration allows extended imipenem dosage

468

ACUTE EXACERBATIONS OF CHRONIC BRONCHITIS

FIGURE 8. Structure of cilastatin.

intervals and causes less toxicity (Norrby SR, 2000). Imipenem/cilastatin is usually reserved for severe AECB in the intensive care setting for patients developing sepsis. In this setting, the agent is often administered with an aminoglycoside. This agent will retain its patent protection through 2009 in the United States, but lost its patent protection in Europe and Japan in 2005. An open-label trial compared imipenem/cilastatin with meropenem (both IV) in 173 hospitalized patients with severe COPD (Hamacher J, 1995). The clinical response at the end of treatment was favorable in both groups, with a cure or improvement achieved in 97.6% of the meropenem patients and in 96.3% of the imipenem/cilastatin patients. Nausea or vomiting was reported more frequently in patients treated with imipenem/cilastatin, whereas an increase in aminotransferases (indicative of liver damage) was reported in the meropenem group. Macrolides Overview. Macrolides inhibit bacterial protein synthesis. They demonstrate excellent activity against atypical organisms (Mycoplasma, Chlamydia, and Legionella species), but their activity against typical pathogens (H. inßuenzae and S. pneumoniae) is variable. Macrolides are indicated for use in AECB and are typically used as Þrst- and second-line agents for this indication. They are available in oral and parenteral formulations. The advanced-generation macrolides have serum half-lives that are much longer than earlier-generation agents, allowing for once-daily dosing. In addition, the newer macrolides demonstrate better activity against S. pneumoniae and excellent distribution into respiratory tissues. Side effects, particularly of the early macrolides, are associated primarily with the GI tract (e.g., abdominal cramps, nausea, vomiting). Hypersensitivity reactions are rare. The advanced-generation macrolides have a lower incidence of side effects than the older agents. The prevalence of macrolide resistance has increased dramatically over the past two decades. Geographically, it is highest in the Asia/PaciÞc region with 73% resistance in Japan and 81.5% in Hong Kong. In Europe, France (47%) and Italy (42%) have a high prevalence of resistance, while 19–34% of pneumococci are macrolide resistant in the United States (Jacobs MR 1999). Mechanism of Action. Macrolides act by binding to the 23S ribosomal RNA (rRNA) in the 50S subunit of the ribosome. Binding to the 23S rRNA inhibits

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the translocation of RNA during protein synthesis and blocks bacterial protein synthesis. Erythromycin. Erythromycin (Lilly/Shionogi’s Ilosone, Abbott’s Erythrocin, generics) (Figure 9) is the class standard macrolide that has been available since the 1950s. The agent has been widely utilized for respiratory tract infections, but the next-generation macrolides, which provide more-convenient dosing and expanded spectrum, have replaced this agent for more-severe respiratory infections such as community acquired pneumonia (CAP) and AECB. The agent is available in both oral and parenteral formulations. Erythromycin acts by binding to the 50S ribosomal subunit of susceptible microorganisms, resulting in inhibition of protein synthesis. Erythromycin is active against a range of gram-positive, gram-negative, and atypical organisms. However, many strains of H. inßuenzae and S. pneumoniae are resistant to erythromycin. Early studies of erythromycin demonstrated its efÞcacy in respiratory tract infections. A double-blind, randomized Phase III clinical trial compared the safety and efÞcacy of dirithromycin (Lilly’s Dynabec) (500 mg once daily) and erythromycin (250 mg orally four times daily) in the treatment of either acute bacterial bronchitis or acute bacterial exacerbations of chronic bronchitis (Gaillat J, 1993). The trial included 1,222 patients (529 with acute bronchitis and 693 with acute exacerbations of chronic bronchitis). Of this number, 135 patients with acute bronchitis and 202 patients with acute exacerbations of chronic bronchitis were evaluable. In acute exacerbations of chronic bronchitis, the drugs were also effective with 98.7% and 95.0% cure or improvement at 10–14 days post-therapy for dirithromycin and erythromycin, respectively. Pathogen eradication rates were 75.3% in both treatment groups. There were no statistically signiÞcant differences in clinical and bacteriological results between treatments

O H

CH3 H

CH3

CH3 OH

H HO

CH3 H

O HO

H

CH3

O N(CH3)2

H

O H3C H

CH3

O H CH2CH3 HO

O O

CH3 OCH3

CH3 FIGURE 9. Structure of erythromycin.

OH

470

ACUTE EXACERBATIONS OF CHRONIC BRONCHITIS

in patients with acute bronchitis or acute exacerbations of chronic bronchitis. Of the 1,222 patients included, no signiÞcant differences were observed in the number of patients reporting adverse events. There were 9 early discontinuations due to adverse events in the dirithromycin group and 14 in the erythromycin group. Erythromycin is associated with GI side effects in about 20% of patients, causing physicians to shift to the newer macrolides. Clarithromycin. Clarithromycin (Abbott’s Biaxin, Biaxin XL/Klacid, Taisho’s Clarith) (Figure 10) is an advanced-generation macrolide commonly used for multiple respiratory tract infections, including CAP, AECB, otitis media, and sinusitus. This product is available in tablets, extended-release tablets, and granules. Clarithromycin acts by binding to the 50S ribosomal subunit of susceptible microorganisms, resulting in inhibition of protein synthesis. Its spectrum of activity includes a range of aerobic and anaerobic gram-positive and gram-negative microorganisms; similar to other macrolides, clarithromycin is highly active against atypical pathogens such as Mycoplasma pneumoniae. Clarithromycin is potent against staphylococci and streptococci and has modest activity against H. inßuenzae. Several trials have evaluated the efÞcacy of clarithromycin in AECB. In one multicenter, randomized, investigator-blinded, randomized Phase III study, 287 subjects with AECB were treated with either clarithromycin extended-release (two 500 mg tablets once daily for seven days) or amoxicillin/clavulanate (one 875 mg tablet twice daily for ten days) (Anzueto A, 2001). The clinical efÞcacy was comparable between the two groups, with the clinical cure rate of 85% for clarithromycin compared with 87% for the amoxicillin/clavulanate group. Clarithromycin achieved a bacteriological cure rate of 92% compared with 89% for the amoxicillin/clavulanate group. Amoxicillin/clavulanate had a higher discontinuation rate related to side effects (6%) compared with clarithromycin (1%). Adverse events generally occurred with a similar frequency in the two treatment groups.

FIGURE 10. Structure of clarithromycin.

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Azithromycin. Azithromycin (PÞzer’s Zithromax/Zitromax) (Figure 11) is widely used for the treatment of respiratory tract infections, including AECB. Azithromycin is available in both oral and IV formulations. The agent is recognized for its broad efÞcacy, dosing advantages, and favorable side-effect proÞle. Azithromycin has become the leading macrolide in the United States owing to its high safety, long half-life (allowing for a short treatment course), and excellent promotion by PÞzer. Azithromycin acts by binding to the 50S ribosomal subunit of susceptible microorganisms and interferes with microbial protein synthesis. It demonstrates in vitro activity against a wide range of bacteria, including gram-positive bacteria such as S. pneumoniae and gram-negative bacteria such as H. inßuenzae. Azithromycin demonstrates cross resistance with erythromycin-resistant grampositive strains and most strains of methicillin-resistant staphylococci. In a randomized, double-blind, controlled Phase III clinical trial in AECB, azithromycin (500 mg once daily for three days) was compared with clarithromycin (500 mg twice daily for ten days) (PÞzer, 2005). The primary end point of this trial was the clinical cure rate measured at days 21–24. For the 304 patients analyzed at the day 21–24 visit, the clinical cure rate for three days of azithromycin was 85% (125/147) compared with 82% (129/157) for ten days of clarithromycin. In the safety analysis of this study, the incidences of treatment-related adverse events, primarily GI, were comparable between treatment arms. Azithromycin is rapidly and extensively taken up by leukocytes, which deliver the drug to the site of infection, where the drug maintains high sustained concentrations. In addition, azithromycin has a long half-life (68 hours) allowing for its convenient Þve-day dosing regimen. Azithromycin also offers excellent tolerability, with a low incidence of adverse events. This drug compares favorably with amoxicillin/clavulanate and other competitive agents from a side-effect and tolerability standpoint. The most common side effects are diarrhea/loose stools (4–5%), nausea (3%), and abdominal pain (2–3%); the overall discontinuation rate is 0.7%.

FIGURE 11. Structure of azithromycin.

472

ACUTE EXACERBATIONS OF CHRONIC BRONCHITIS

Ketolides Ketolides are a new class of macrolide derivatives designed speciÞcally to combat macrolide-resistant respiratory tract pathogens. The ketolides are semisynthetic derivatives of the macrolide erythromycin A, with a keto group replacing the lcladinose group at position 3 of the macrolactone ring. The ketolides exhibit good activity against gram-positive and some gram-negative organisms, and have excellent activity against drug-resistant S. pneumoniae, including macrolideresistant strains (Poehlsgaard J, 2002). Ketolides display excellent pharmacokinetics, allowing once-daily dose administration and extensive tissue distribution relative to serum. The available ketolide, telithromycin, is well tolerated with side effects primarily being GI symptoms (diarrhea, nausea, and headache). Mechanism of Action. Ketolides, similar to the macrolides, act by binding to the 23S ribosomal RNA (rRNA) in the 50S subunit of the ribosome. Binding to the 23S rRNA inhibits the translocation of RNA during protein synthesis and blocks bacterial protein synthesis. Ketolides and macrolides have essentially the same binding sites, which are located on a portion of the rRNA and on ribosomal proteins. However, ketolides bind to ribosomes with higher afÞnity than macrolides. This difference could contribute to ketolide’s activity against macrolide-resistant strains (Poehlsgaard J, 2002). Telithromycin. Telithromycin (SanoÞ-Aventis/Fujisawa/Sankyo’s Ketek) (Figure 12), an oral ketolide, is the Þrst member of this new class to be approved for clinical use. Telithromycin was Þrst approved in Europe in 2001; Japanese approval was attained in 2003. In the United States, telithromycin’s approval was delayed for approximately two years. The FDA originally issued an approvable letter for telithromycin in 2001, and requested additional data to support the Þling. To support U.S. approval, which Þnally occurred in April 2004, SanoÞ-Aventis conducted the largest-ever comparative antibiotic trial, enrolling over 24,000 subjects. The product was launched in August 2004. It is expected to retain patent protection through 2015 in the United States and Japan, and through 2016 in Europe. Telithromycin is active against a broad range of pathogens, including S. pneumoniae (including multidrug-resistant strains), H. inßuenzae, M. catarrhalis, Chlamydia pneumoniae, and M. pneumoniae. Against isolates of S. pneumoniae, telithromycin has been shown to demonstrate concentration-dependent bactericidal activity in vitro. Telithromycin’s advantage over currently available macrolides rests in its enhanced activity against erythromycin-resistant S. pneumoniae. Antibacterial activity against other respiratory pathogens, such as H. inßuenzae and atypical bacteria, is similar to that of available advanced-generation macrolides, except that telithromycin appears to have a lower propensity to induce certain types of resistance (inducible methylase gene expression) (Buxbaum A, 2003). Telithromycin’s efÞcacy in AECB was demonstrated in three randomized, double-blind, controlled studies. The agent’s clinical cure rate was comparable to

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FIGURE 12. Structure of telithromycin.

those of cefuroxime axetil, clarithromycin, and amoxicillin/clavulanate, ranging between 85% and 86%. A 5-day course of telithromycin versus cefuroxime axetil 500 mg twice a day (ten-day therapy) was compared, resulting in efÞcacy rates of 86.4% and 83.1% for telithromycin and cefuroxime, respectively. A Þve-day course of therapy for telithromycin was compared with amoxicillin/clavulanic acid (500/125 mg three times a day for ten days), resulting in efÞcacy rates of 86.1% and 82.1% for telithromycin and amoxicillin/clavulanic acid, respectively. A Þve-day course of therapy of telithromycin versus clarithromycin (500 mg twice a day for ten days) resulted in efÞcacy rates of 85.8% for telithromycin compared with 89.2% for clarithromycin (Aubier M, 2000; DeAbate C, 2000). Telithromycin’s side effects include nausea and diarrhea, which, in clinical trials, occurred at rates comparable to comparator antibiotics. Visual disturbances (namely blurred vision and/or difÞculty focusing) have also occurred in approximately 1% of patients. In the large comparative trial, telithromycin demonstrated comparable efÞcacy to amoxicillin/clavulanate in terms of hospitalization, need for new antimicrobial medication for the primary infection or complication, and time lost from work (SanoÞ-Aventis, 2004[a]). Fluoroquinolones Overview. Fluoroquinolones are broad-spectrum antibacterials that have experienced an upsurge in use in recent years. Because of their broad-spectrum activity, high efÞcacy, favorable dosing, and availability in oral and IV form, these agents are indicated for a range of bacterial infections, including respiratory, GI, and urinary tract infections. Earlier generations of ßuoroquinolones (e.g., oßoxacin) had limited activity against some respiratory pathogens, such as S. pneumoniae.

474

ACUTE EXACERBATIONS OF CHRONIC BRONCHITIS

However, more recent ßuoroquinolone agents (so-called third-generation agents or the “respiratory ßuoroquinolones”) are active against a broad spectrum of gram-positive and gram-negative bacteria, including atypical organisms. Therefore, they are highly effective in RTIs. An important factor inßuencing the use of recently launched ßuoroquinolones is that several agents from this class were withdrawn from the market owing to toxicity (e.g., trovaßoxacin [PÞzer’s Trovan], grepaßoxacin [GlaxoSmithKline’s Raxar]). Fluoroquinolones, as a class, are associated with alterations in normal cardiac conduction (i.e., prolongation of the QT interval), which can cause cardiac arrhythmias in vulnerable individuals. The degree of QT prolongation varies for each agent. Among these, sparßoxacin (Mylan’s Zagam) and grepaßoxacin have been associated with the longest QT prolongation (Ball P, 2000). Both of these agents have now been removed from the U.S. market. Grepaßoxacin use resulted in seven associated cardiac-related deaths. To date, resistance to the ßuoroquinolones by respiratory pathogens has been extremely rare (Hoban DJ, 2001). However, increasing ßuoroquinolone resistance in S. pneumoniae isolates has been reported, and clinical failures with levoßoxacin have been noted in Canada (Odland BA, 1999; Chen DK, 1999). Fluoroquinolone-resistant isolates are more common among persons older than 65, who have the highest level of ßuoroquinolone use. Prior exposure to ßuoroquinolones is one of the major risk factors for the acquisition of levoßoxacin-resistant S. pneumoniae (Ho PL, 2001). Emerging resistance to older ßuoroquinolones, such as ciproßoxacin, has also been demonstrated worldwide, ranging from 3% to 12% (Linares, 1999). Fluoroquinolone resistance often involves alteration of their targets, the topoisomerases. Mechanism of Action. Fluoroquinolones interact with two related yet distinct targets within the bacterial cell: DNA gyrase and topoisomerase IV. DNA gyrase and topoisomerase IV are vital enzymes involved in bacterial DNA replication. These agents are potent inhibitors of nucleic acid synthesis. The exact nature of the interaction of quinolones with their target enzymes is not completely understood. However, this interaction blocks the progression of DNA replication, leading to strand breaks and rapid cell death. The inhibition of DNA gyrase is bactericidal, whereas inhibition of topoisomerase IV is primarily bacteriostatic. Levofloxacin. Levoßoxacin (Johnson & Johnson’s Levaquin, SanoÞ-Aventis’ Tavanic, Daiichi’s Cravit) (Figure 13) is approved for the treatment of AECB and several other bacterial infections (e.g., pneumonia, urinary tract infection, skin infections). Levoßoxacin was Þrst launched in Japan in 1993 and has been widely available since 1998. The agent is expected to lose patent protection in the United States and Japan in 2010 and in Europe in 2011. The drug is a broad-spectrum antibacterial and is available in both oral and IV formulations. Levoßoxacin is the levo enantiomer of oßoxacin (Ortho McNeil’s Floxin, generics). The agent’s mechanism of action involves inhibition of bacterial topoisomerase IV and DNA gyrase, which are enzymes required for DNA replication,

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FIGURE 13. Structure of levofloxacin, the (S)-enantiomer.

transcription, repair, and recombination. Levoßoxacin has in vitro activity against a wide range of gram-negative and gram-positive microorganisms. A randomized, double-blind, multicenter Phase III study among adult patients with AECB compared the efÞcacy and safety of two different doses of levoßoxacin with cefuroxime axetil (Shah PM, 1999). A total of 832 patients were randomized to receive oral levoßoxacin (250 mg or 500 mg once daily) or oral cefuroxime axetil (250 mg twice daily) for 7–10 days. The primary efÞcacy analysis was based on the clinical response in patients with bacteriologically conÞrmed AECB, determined 5–14 days after the end of therapy. Both doses of levoßoxacin were at least as effective as cefuroxime axetil and were active against the main pathogens of clinical relevance (H. inßuenzae, S. pneumoniae, and M. catarrhalis). The cure rates in the treated population were 70% for levoßoxacin 250 mg, 70% for levoßoxacin 500 mg, and 61% for cefuroxime axetil. All three treatment regimens were equally well tolerated. Moxifloxacin. Moxißoxacin (Bayer’s Avelox) (Figure 14) is a ßuoroquinolone used for the treatment of respiratory tract infections including AECB. In December 1999, the FDA approved the oral formulation of moxißoxacin, and in December 2001, the agency approved the parenteral formulation of this product for the same indications. In Germany, oral moxißoxacin received regulatory approval in September 1999 for the treatment of community-acquired infections, including AECB. The parenteral formulation was approved in May 2002. Oral moxißoxacin was recently launched in Spain and Italy and is awaiting regulatory approval in other European markets. In Japan, moxißoxacin is in Phase III and Phase I trials for the oral and IV formulations, respectively. This agent will retain patent protection through 2014 in the United States and through 2009 in Europe and Japan. Moxißoxacin displays excellent activity against a broad spectrum of bacteria, including penicillin- and macrolide-resistant S. pneumoniae. It is highly active against the following: • •

Common RTI pathogens such as S. pneumoniae, H. inßuenzae, and M. catarrhalis. RTI pathogens with reduced susceptibility to conventional agents such as penicillin and macrolide-resistant strains of S. pneumoniae.

476

ACUTE EXACERBATIONS OF CHRONIC BRONCHITIS

O

O

F OH H

N

N

HN

O H

CH3

FIGURE 14. Structure of moxifloxacin.

• •

Unusual community RTI pathogens such as Staphylococcus aureus and K. pneumoniae. Atypical microorganisms, such as Mycoplasma, Legionella, and Chlamydia.

Moxißoxacin offers superior activity against S. pneumoniae compared with that of current quinolones (e.g., ciproßoxacin, levoßoxacin) and is extremely effective in treating other respiratory pathogens. A multi-center, multinational, randomized, double-blind Phase III study compared moxißoxacin with amoxicillin in patients experiencing AECB (Wilson R, 2004). During a 12-month monitoring period, 730 patients with AECB were randomized to receive moxißoxacin (400 mg, once daily, for Þve days) or a standard treatment regimen consisting of amoxicillin (500 mg, three times daily, for seven days) or clarithromycin (500 mg, twice daily, for seven days) or the cephalosporin cefuroxime axetil (250 mg, twice daily, for seven days). The primary end point was clinical success (resolution and improvement) at 7–10 days post treatment. Patients were also monitored after nine months to assess long-term outcomes, including time until their next exacerbation. Moxißoxacin demonstrated a signiÞcantly higher clinical cure rate (70.9% versus 62.8% in the comparator arm) and a signiÞcantly higher bacteriological response rate (92% versus 81% in the comparator arm). A follow-up study showed that for patients who received moxißoxacin, the average interval between episodes of AECB was approximately two weeks longer than for those who received standard therapy. Moxißoxacin is not associated with phototoxicity, but it has been shown to alter the QT interval above baseline (Culley CM, 2001). The frequency and severity of adverse reactions were equal to those experienced among the comparators (clarithromycin and cefuroxime axetil) and included nausea (8%), diarrhea (6%), and dizziness (3%). Gatifloxacin. Gatißoxacin (Bristol-Myers Squibb’s Tequin, Kyorin/Dainippon’s Gatißo) (Figure 15) was approved in the United States in December 1999 in both oral and IV formulations. The oral drug was launched in Japan in June 2002, where Kyorin is comarketing the drug with Dainippon Pharmaceutical. Gatißoxacin has a broad spectrum of activity similar to that observed in other thirdgeneration ßuoroquinolones (Blondeau JM, 2000). The agent is well absorbed

CURRENT THERAPIES

O

477

O

F OH CH3 HN

N

N O CH3

FIGURE 15. Structure of gatifloxacin.

following oral administration (with close to 100% bioavailability), and its pharmacodynamic and pharmacokinetic properties (e.g., high volume of distribution, long elimination half-life) allow for once-daily administration (Nakashima M, 1995). A multi-center, prospective, randomized, double-blind, Phase III study in 527 adult outpatients with AECB compared therapy with either 400 mg of gatißoxacin once daily for 5 or 7 days, or 500 mg clarithromycin twice daily for 10 days (Gotfried MH, 2001). Researchers followed up patients in these trial groups for 21–28 days after treatment ended. Overall clinical efÞcacy (including efÞcacy at the last follow-up) in all three treatment groups was 87%, proving clinical equivalence not only between gatißoxacin and clarithromycin but also between gatißoxacin Þve-day therapy and gatißoxacin seven-day therapy. Overall microbiological eradication rates (excluding patients with resistant pathogens) were 98%, 94%, and 98% in the gatißoxacin Þve-day, seven-day, and clarithromycin groups, respectively. All S. pneumoniae and H. inßuenzae isolates identiÞed before study treatment were susceptible to gatißoxacin, and no pathogens susceptible to gatißoxacin before treatment developed resistance during therapy. The researchers did not report the overall rate of drug-related adverse events in the study but listed the most common adverse events as diarrhea (7% in gatißoxacin Þve-day therapy, 6% in gatißoxacin seven-day therapy, and 6% in clarithromycin ten-day therapy groups) and nausea (5%, 6%, and 6%, respectively) (Gotfried MH, 2001). Gatißoxacin’s product labeling warns against giving this agent to patients with long QT syndrome. It should not be given to those receiving type IA or III antiarrhythmics and should be used with caution if the patient is receiving other drugs that prolong the QT interval (Bristol-Myers Squibb, 2003). In postmarketing surveillance, serious disturbances of glucose homeostasis in diabetic patients being treated with gatißoxacin have been reported. The relevance of this phenomenon and its effect on the drug’s use are not clear at this time. Tetracyclines Overview. Tetracyclines are the prototypical broad-spectrum antibiotic and are occasionally used as Þrst-line agents in some markets (e.g., Germany) for the treatment of mild AECB, particularly when cost, penicillin hypersensitivity, and β-lactam resistance are of concern. The widespread use of tetracyclines has

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ACUTE EXACERBATIONS OF CHRONIC BRONCHITIS

resulted in a steady increase in the prevalence of resistance to these agents. Therefore, empiric use of tetracyclines is usually restricted to regions where resistance levels remain low or when other appropriate antibiotics are contraindicated. Although some tetracyclines have very low activity against S. pneumoniae, doxycycline maintains good antipneumococcal activity and is the tetracycline most commonly used for the treatment of AECB. Minocycline (Wyeth’s Minocin, generics) is another widely available tetracycline also used for RTIs. However, the agent will not be discussed further owing to its limited use in AECB. All tetracyclines have important adverse reactions with respect to bones and teeth (i.e., bone deposition and permanent teeth staining), and they are contraindicated during pregnancy and for children under age eight. Adverse events most commonly reported include stomach upset, diarrhea, nausea, headache, and vomiting. Phototoxicity, dizziness, and vertigo may also occur. Mechanism of Action. Tetracyclines bind to receptors on the 30S subunit of bacterial ribosomes and prevent the attachment of transfer RNA (tRNA) to the ribosomal complex. Binding to the 30S subunit effectively prevents the addition of new amino acids to the growing polypeptide chain, halting the process of protein synthesis. Tetracyclines are usually bacteriostatic, but, in some organisms, their effect can be bactericidal at high doses. Doxycycline. Doxycycline (PÞzer’s Vibramycin, generics) (Figure 16) is a broad-spectrum antibiotic derived from oxytetracycline that has been used to treat both gram-negative and gram-positive bacterial infections since the 1960s. It is currently available in oral suspension, capsule, syrup, and IV formulations and has an improved pharmacokinetic proÞle that includes higher oral bioavailability, greater distribution volume, and longer serum half-life compared with earlier tetracyclines. Doxycycline is the prototypical broad-spectrum antibiotic. Its antimicrobial activity is attributed to the inhibition of protein synthesis by binding to the 30S subunit of the bacterial ribosome and subsequently preventing the addition of new amino acids to the growing polypeptide chain. This mechanism of action is reversible, distorting the subunit in such a way that the anticodons of charged tRNAs cannot properly align with the codons of the mRNA. Doxycycline is approved for the treatment of infections caused by H. inßuenzae, and the agent is active against S. pneumoniae. In a multicenter Phase III

FIGURE 16. Structure of doxycycline.

CURRENT THERAPIES

479

trial conducted in seven European countries, doxycycline demonstrated rapid improvement, usually by the third to Þfth day in 87% of a total 1,747 patients with infections of the respiratory tract. Patients ranged in age from 6 years to older than 80, and approximately 50% of these study subjects were diagnosed with acute bronchitis and AECB. Minimal GI side effects were observed, causing only 4 patients to withdraw from the study (Pestel M, 1975). Adverse events most commonly reported with use of doxycycline include stomach upset, diarrhea, nausea, headache, and vomiting. Phototoxicity, dizziness, and vertigo can also occur with doxycycline use. Glycopeptides Overview. Glycopeptides are used for the treatment of severe or life-threatening infections that are caused by gram-positive organisms such as streptococci and staphylococci. In AECB, they are used rarely. Physicians typically reserve them for severe AECB episodes in which multidrug-resistant organisms are suspected or in which the patient has complications, such as bacteremia or meningitis. They are also used as second-line agents in patients who cannot receive or who have failed to respond to penicillins and cephalosporins. Glycopeptide therapy is relatively well tolerated. The most severe adverse reactions are toxicities associated with the inner ear and kidneys, but they occur only infrequently. Hypersensitivity, of which skin rash is the most common allergic reaction, is reported in 5–10% of patients. Other possible side effects of glycopeptides include GI disturbances, muscle pain, blood disorders (e.g., eosinophilia), and disturbances in liver enzymes. Mechanism of Action. Glycopeptides inhibit bacterial cell-wall biosynthesis by blocking glycopeptide polymerization. This effect produces immediate inhibition of cell-wall synthesis and secondary damage to the cytoplasmic membrane. Glycopeptides also alter the permeability of the cytoplasmic membrane and directly inhibit RNA synthesis. However, the impact these effects have on bacterial viability is largely unknown. These agents have a concentration-independent bactericidal action resulting primarily from inhibition of cell-wall synthesis; they are considered slowly bactericidal against staphylococci. Vancomycin. Vancomycin (Lilly’s Vancocin, generics) (Figure 17) is a narrowspectrum, bactericidal glycopeptide antibiotic. Vancomycin was developed in the 1950s and has been available worldwide since the 1960s; it is available in oral and IV formulations. Because this agent has been available for an extensive period of time, no highquality clinical studies pertaining to AECB have been conducted in recent years. Thus, this section does not cite speciÞc efÞcacy results from clinical studies for vancomycin. Physicians use the injectable glycopeptides extensively to treat systemic infections because these compounds are active against gram-positive bacteria such as

480

ACUTE EXACERBATIONS OF CHRONIC BRONCHITIS

H3N

HO

Me

OH

HO Me

O

O

O

O Cl O

O H

HO

OH

Cl H O O H NH −O

HN H

O

O H

H N

Me

N N

H

O H

O

H

N H

H

+

NH2 H

O

2C

H2N

OH HO

OH FIGURE 17. Structure of vancomycin.

S. pneumoniae. Vancomycin has a relatively long duration of action and therefore can be administered every 12 hours. Toxicity to the inner ear has occurred in patients receiving vancomycin and can be transient or permanent. Most cases have been reported in patients receiving high IV doses or in those with a history of hearing complications. Vancomycin is also associated with kidney toxicity, and patients are screened for a history of renal complications. Renal failure has been reported in patients receiving high doses of IV vancomycin, and physicians exercise particular caution when treating patients concomitantly with another aminoglycoside. Vancomycin can also cause infusion-related ßushing (“red man syndrome”) in some patients. Resistance to vancomycin can occur through a modiÞcation of the components of the bacterial cell wall. Vancomycin binds to a component of the cell wall that when modiÞed or altered results in reduced binding of the drug to the bacteria. However, no cross-resistance between vancomycin and other antibacterial agents has been observed in gram-positive microbes. Teicoplanin. Teicoplanin (SanoÞ-Aventis’ Targocid) is a narrow-spectrum, bactericidal glycopeptide antibiotic similar in structure to vancomycin. It was Þrst launched in France and Italy in 1988 and by 1997 was available in most of Europe. It was launched in Japan in 1997, where Aventis has partnered with Fujisawa for distribution and co-promotion, but has never been available in the United States, where development was suspended in 1999.

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Teicoplanin is available in parenteral formulations and has a signiÞcantly longer duration of action than vancomycin, thus allowing once-daily administration. Unlike vancomycin, teicoplanin can be given by IM as well as by IV injection. Considered a valuable alternative to vancomycin, teicoplanin is used as empirical therapy in immunocompromised patients with hematological malignancies. The effectiveness of teicoplanin has been documented in several indications including lower respiratory infections. An open multicenter Phase III trial in Europe to evaluate the efÞcacy of teicoplanin enrolled a total of 1,431 hospitalized patients. Most patients had S. aureus infections, 19.6% of which were methicillinresistant. Teicoplanin monotherapy was used in 1,037 cases and in combination with other antibacterials in 394 cases. An overall favorable clinical success was seen in 91.7% of the 1,333 evaluable cases (Lewis P, 1988). The most common adverse reactions shared with glycopeptides generally include inner ear and kidney toxicities. Teicoplanin has fewer propensities than does vancomycin to cause toxicities such as inner ear and kidney toxicity, or infusion-related ßushing (“red man syndrome”). Hypersensitivity reactions are also a concern of physicians when considering this therapy. EMERGING THERAPIES Despite the large patient populations with respiratory tract infections (RTIs), in recent years, many biopharmaceutical companies have shifted development effort away from RTIs to pursue other disease areas with perceived higher unmet need (e.g., resistant hospital-acquired infections). Likewise, many companies have shifted away from antibacterial drug development in favor of chronic disease markets perceived to have higher return on investment. The reasons behind the move away from antibiotics include a perceived high level of competition, relatively low unmet need, and relatively high development costs and large clinical trials required for approval of community-based antibiotics. As a result, the pipeline of emerging antibacterial agents, particularly for RTIs, has dwindled. Virtually all of the products approved over the past 10–15 years have been “next-generation” agents within current therapeutic classes (particularly the macrolides and ßuoroquinolones). The “next-generation” macrolides and ßuoroquinolones have provided advantages such as higher activity against respiratory pathogens, broader spectrum of activity, and more-convenient dosing. However, reßecting the decreased interest as well as the scientiÞc challenge, no novel classes have been introduced in the past two decades–with the exception of linezolid (PÞzer’s Zyvox), which is primarily used for complicated gram-positive hospital infections. The peptidyl deformylase inhibitors (PDF inhibitors) represent the Þrst truly novel antibacterial class for RTIs. Future development options in acute exacerbations of chronic bronchitis (AECB) can be considered in two main areas: continued improvement upon the current generation of antibiotics, and development of novel classes of antibiotics active against emerging drug-resistant pathogens (e.g., future ßuoroquinoloneresistant bacteria). In the Þrst respect, the opportunity for improvement has

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ACUTE EXACERBATIONS OF CHRONIC BRONCHITIS

become increasingly narrow, owing to the availability of multiple highly effective, once-daily, and relatively safe and well tolerated agents. The second area of improvement, novel mechanisms and classes against resistant infections, is likely to become more important in the more distant future (beyond Þve years) because there are currently multiple second- and third-line options. Within the RTI markets, AECB is an important segment because of its high incidence, acute concern for antibacterial resistance (because these patients have frequent recurrences and antibiotic courses), and high Þnancial cost associated with hospitalization. Fluoroquinolones Overview. The ßuoroquinolones have proven to be one of the most commercially and clinically successful antibacterial classes for the treatment of AECB. Their broad spectrum and high potency against respiratory pathogens (which improved with the more recent launches such as moxißoxacin and gatißoxacin), safety, and convenient once-daily dosing have earned them a favorable position in the minds of clinicians. One of their main advantages in AECB has been their high H. inßuenzae activity compared with other agents such as the macrolides. The few concerns associated with this class of agents include their potential to induce abnormal cardiac conductions (i.e., prolongation of the QT interval on electrocardiograms) and liver toxicity (particularly since the withdrawal of PÞzer’s trovaßoxacin). Emerging ßuoroquinolones in development provide potential advances over currently marketed agents mainly in terms of higher potency against respiratory pathogens. Mechanism of Action. Fluoroquinolones act by inhibiting bacterial topoisomerase IV and DNA gyrase, which are enzymes required for DNA replication, transcription, repair, and recombination. The exact nature of the interaction of ßuoroquinolones with their target enzymes is not completely understood; however, this interaction blocks the progression of DNA replication, leading to strand breaks and rapid cell death. These agents are potent inhibitors of nucleic acid synthesis. Gemifloxacin. Gemißoxacin (LG Life Sciences [a spin-off of LG Chemical]/Oscient’s [formerly Genesoft/Genome Therapeutics] Factive) is the newest ßuoroquinolone to enter the U.S. market. The oral product was launched in September 2004 for the treatment of mild to moderate AECB and communityacquired pneumonia (CAP). An IV form is also in development. The drug was formerly being developed by SmithKline Beecham (now GlaxoSmithKline [GSK]). SmithKline Beecham submitted a new drug application (NDA) in December 1999 for the treatment of bacterial respiratory and urinary tract infections, including CAP, chronic bronchitis, and acute sinusitis. However, the FDA issued a nonapprovable letter in 2000, citing insufÞcient safety data about gemißoxacin’s effect on QT interval, liver toxicity, and rash in premenopausal women. Additional studies were conducted to support the application; however,

EMERGING THERAPIES

483

GSK returned commercialization rights to LG Life Sciences in 2002. Subsequently, Genesoft Pharmaceuticals, a small, privately held company, acquired commercialization rights for the product in North America and Europe. In 2004, Genesoft merged with Genome Therapeutics to form Oscient Pharmaceuticals. Gemißoxacin, as a member of the ßuoroquinolone class, acts by inhibiting bacterial topoisomerase IV and DNA gyrase. These enzymes are required for DNA replication, transcription, repair, and recombination. In vitro, gemißoxacin has broad-spectrum activity against gram-positive, gram-negative, and atypical pathogens including S. pneumoniae, H. inßuenzae, Moraxella catarrhalis, Legionella pneumophila, Mycoplasma pneumoniae, and Chlamydia pneumoniae. Studies have shown that the agent has the highest in vitro potency against S. pneumoniae, compared with ciproßoxacin (Schering-Plough’s Cipro, generics), levoßoxacin (Johnson & Johnson’s Levaquin), gatißoxacin (Bristol-Myers Squibb’s [BMS’s] Tequin), and moxißoxacin (Bayer’s Avelox), and may be active against strains of S. pneumoniae resistant to these agents (Boswell FJ, 2002, Koeth LM, 2002). Gemißoxacin has been shown to be highly effective for the treatment of AECB in several large clinical trials. In a randomized, double-blinded, multicenter study, gemißoxacin (320 mg once daily for Þve days) was compared with clarithromycin (500 mg twice daily for seven days) in 712 subjects with AECB (Wilson R, 2002). The clinical success rates as well as the bacterial success rates of the two arms were equivalent (85.4% versus 84.6% and 86.7% versus 73.1%, respectively). A subset of patients was followed up for an additional 26 weeks to monitor the frequency of AECB recurrence. SigniÞcantly more patients receiving gemißoxacin than clarithromycin remained free of AECB recurrences (71.0% versus 58.5%, respectively). In a second randomized, open-label study in adult hospitalized AECB patients, a 5-day course of gemißoxacin was compared with sequential IV ceftriaxone followed by oral cefuroxime axetil treatment (up to 10 days) (Wilson R, 2003). In 274 treated patients, the clinical success rates on an intent-to-treat basis at 21–28 days follow up were 82.6% for gemißoxacin compared with 72.1% for the ceftriaxone/cefuroxime arm. Both therapies were well tolerated. The study showed that gemißoxacin-treated patients were discharged from the hospital two days earlier, a statistically shorter length of stay. A third randomized, double-blind clinical trial compared gemißoxacin (320 mg once daily for Þve days) to amoxicillin/clavulanate (500/125 mg three times daily for seven days) in AECB patients. Six hundred patients were evaluated. The two arms demonstrated equivalent clinical success rates (93.6% versus 93.2%), and gemißoxacin showed a superior, but not statistically signiÞcant, bacterial success rate (90.9% compared with 79.5%, respectively). Both therapies were well tolerated (File TM, 2001). Finally, a randomized, double-blinded study examined a Þve-day course of gemißoxacin compared with a seven-day course of levoßoxacin (Sethi S, 2004). Three hundred and sixty subjects with AECB were enrolled. The clinical success rate at follow-up at two to three weeks was 85.2% with gemißoxacin and

484

ACUTE EXACERBATIONS OF CHRONIC BRONCHITIS

78.1% with levoßoxacin. At long-term follow-up (four to Þve weeks), the clinical success rates were 83.7% with gemißoxacin and 78.4% with levoßoxacin. Safety analysis from clinical studies conducted with gemißoxacin that included almost 7,000 patients treated with the drug indicates that the adverse event rate of the agent is similar to comparator treatment groups (Ball P, 2004)[b]. The most common minor treatment-related side effects were diarrhea, rash, and nausea, with the mild rash occurring at a more common frequency than comparator antibiotics (2.8% versus 0.6%, respectively. Gemißoxacin may differentiate itself based on its potency (it has one of the highest potencies against respiratory pathogens) and its convenient Þve-day dosing schedule. However, gemißoxacin would be the third respiratory ßuoroquinolone on the market to compete with levoßoxacin (following BMS’s gatißoxacin and Bayer’s moxißoxacin), the current market leader. Therefore, it will face signiÞcant competition both in differentiating itself to primary care physicians and in gaining favorable formulary status. Gemifloxacin. Gemißoxacin (LG Life Sciences [a spin-off of LG Chemical]/Oscient’s [formerly Genesoft/Genome Therapeutics] Factive) is the newest ßuoroquinolone to enter the U.S. market. The oral product was launched in September 2004 for the treatment of mild to moderate AECB and communityacquired pneumonia (CAP). An IV form is also in development. The drug was formerly being developed by SmithKline Beecham (now GlaxoSmithKline [GSK]). SmithKline Beecham submitted a new drug application (NDA) in December 1999 for the treatment of bacterial respiratory and urinary tract infections, including CAP, chronic bronchitis, and acute sinusitis. However, the FDA issued a nonapprovable letter in 2000, citing insufÞcient safety data about gemißoxacin’s effect on QT interval, liver toxicity, and rash in premenopausal women. Additional studies were conducted to support the application; however, GSK returned commercialization rights to LG Life Sciences in 2002. Subsequently, Genesoft Pharmaceuticals, a small, privately held company, acquired commercialization rights for the product in North America and Europe. In 2004, Genesoft merged with Genome Therapeutics to form Oscient Pharmaceuticals. Gemißoxacin, as a member of the ßuoroquinolone class, acts by inhibiting bacterial topoisomerase IV and DNA gyrase. These enzymes are required for DNA replication, transcription, repair, and recombination. In vitro, gemißoxacin has broad-spectrum activity against gram-positive, gram-negative, and atypical pathogens including S. pneumoniae, H. inßuenzae, Moraxella catarrhalis, Legionella pneumophila, Mycoplasma pneumoniae, and Chlamydia pneumoniae. Studies have shown that the agent has the highest in vitro potency against S. pneumoniae, compared with ciproßoxacin (Schering-Plough’s Cipro, generics), levoßoxacin (Johnson & Johnson’s Levaquin), gatißoxacin (Bristol-Myers Squibb’s [BMS’s] Tequin), and moxißoxacin (Bayer’s Avelox), and may be active against strains of S. pneumoniae resistant to these agents (Boswell FJ, 2002, Koeth LM, 2002). Gemißoxacin has been shown to be highly effective for the treatment of AECB in several large clinical trials. In a randomized, double-blinded, multicenter study,

EMERGING THERAPIES

485

gemißoxacin (320 mg once daily for Þve days) was compared with clarithromycin (500 mg twice daily for seven days) in 712 subjects with AECB (Wilson R, 2002). The clinical success rates as well as the bacterial success rates of the two arms were equivalent (85.4% versus 84.6% and 86.7% versus 73.1%, respectively). A subset of patients was followed up for an additional 26 weeks to monitor the frequency of AECB recurrence. SigniÞcantly more patients receiving gemißoxacin than clarithromycin remained free of AECB recurrences (71.0% versus 58.5%, respectively). In a second randomized, open-label study in adult hospitalized AECB patients, a 5-day course of gemißoxacin was compared with sequential IV ceftriaxone followed by oral cefuroxime axetil treatment (up to 10 days) (Wilson R, 2003). In 274 treated patients, the clinical success rates on an intent-to-treat basis at 21–28 days follow up were 82.6% for gemißoxacin compared with 72.1% for the ceftriaxone/cefuroxime arm. Both therapies were well tolerated. The study showed that gemißoxacin-treated patients were discharged from the hospital two days earlier, a statistically shorter length of stay. A third randomized, double-blind clinical trial compared gemißoxacin (320 mg once daily for Þve days) to amoxicillin/clavulanate (500/125 mg three times daily for seven days) in AECB patients. Six hundred patients were evaluated. The two arms demonstrated equivalent clinical success rates (93.6% versus 93.2%), and gemißoxacin showed a superior, but not statistically signiÞcant, bacterial success rate (90.9% compared with 79.5%, respectively). Both therapies were well tolerated (File TM, 2001). Finally, a randomized, double-blinded study examined a Þve-day course of gemißoxacin compared with a seven-day course of levoßoxacin (Sethi S, 2004). Three hundred and sixty subjects with AECB were enrolled. The clinical success rate at follow-up at two to three weeks was 85.2% with gemißoxacin and 78.1% with levoßoxacin. At long-term follow-up (four to Þve weeks), the clinical success rates were 83.7% with gemißoxacin and 78.4% with levoßoxacin. Safety analysis from clinical studies conducted with gemißoxacin that included almost 7,000 patients treated with the drug indicates that the adverse event rate of the agent is similar to comparator treatment groups (Ball P, 2004)[b]. The most common minor treatment-related side effects were diarrhea, rash, and nausea, with the mild rash occurring at a more common frequency than comparator antibiotics (2.8% versus 0.6%, respectively. Gemißoxacin may differentiate itself based on its potency (it has one of the highest potencies against respiratory pathogens) and its convenient Þve-day dosing schedule. However, gemißoxacin would be the third respiratory ßuoroquinolone on the market to compete with levoßoxacin (following BMS’s gatißoxacin and Bayer’s moxißoxacin), the current market leader. Therefore, it will face signiÞcant competition both in differentiating itself to primary care physicians and in gaining favorable formulary status. Garenoxacin. Garenoxacin (Schering-Plough/Toyama’s T-3811, des-F(6)quinolone garenoxacin) is a potent, oral, broad-spectrum quinolone that is preregistered in the United States. Garenoxacin has a broad spectrum of activity, similar

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ACUTE EXACERBATIONS OF CHRONIC BRONCHITIS

to that displayed by the other new ßuoroquinolones (Bassetti M, 2002; Christiansen KJ, 2004; Takahata M, 1999; Gordon KA, 2002). Garenoxacin shows the highest antibacterial activity against S. pneumoniae when compared with ciproßoxacin, levoßoxacin, gatißoxacin, and moxißoxacin, and it is more potent than ciproßoxacin, levoßoxacin, and moxißoxacin against anaerobes (Bassetti M, 2002; Credito KL, 2001[b]). It is also highly potent against ciproßoxacin-resistant strains of S. pneumoniae and Staphylococcus aureus, and like gatißoxacin, it is associated with a lower risk of promoting resistance development (Boswell FJ, 2001; Entenza JM, 2001; Hartman-Neumann S, 2001). Garenoxacin has favorable pharmacokinetics and is expected to be administered as an oral or injectable once-daily formulation. A Phase II trial comparing the safety and tolerability of garenoxacin (dosed orally at 400 mg once daily) with placebo for 28 days in 45 healthy volunteers showed the drug to be both safe and well tolerated, with no serious adverse effects (Stewart C, 2001). Similarly, Phase II clinical trial results for the IV formulation showed garenoxacin to be safe and well tolerated, with no serious adverse effects. No clinically signiÞcant effects on liver function and QT interval have been reported (Gajjar D, 2001). In a randomized, double-blind, placebo-controlled, Phase III study, treatment with garenoxacin (400 mg/day for either Þve or ten days) in 301 patients with AECB was clinically efÞcacious. Clinical response rates, determined 28 days after completion of therapy, were 90% and 93% for the Þve- and ten-day treatment groups, respectively (Fogarty C, 2001). The same study showed that both the Þveand ten-day treatment regimens were well tolerated; nausea (4%) and diarrhea (4%) were the most commonly reported adverse events. Garenoxacin is a promising new ßuoroquinolone with potent activity against key respiratory pathogens. In AECB, the agent provides the potential advantage of retained activity against resistant pathogens. The product has the potential to be the fourth respiratory ßuoroquinolone on the market (following gatißoxacin, moxißoxacin, and gemißoxacin). Garenoxacin’s most differentiating feature may be its low chondrotoxic potential and, therefore, its potential for use in pediatric patients. (Although obviously not the focus of the AECB market, this feature is important in the product’s overall market potential.) Unlike other ßuoroquinolones, which are not approved in children because of toxicity in joints and cartilage, toxicological studies of garenoxacin’s effect on immature and juvenile joint cartilage have shown that this drug does not induce cartilage defects or signiÞcant arthropathy in immature rats or dogs (Kappel EM, 2002; Kastner M, 2004). Sitafloxacin. Daiichi’s injectable ßuoroquinolone sitaßoxacin is undergoing Phase III trials in Japan against S. pneumoniae infections. Daiichi Þled a marketing application for the oral formulation in Japan, which was subsequently withdrawn because of insufÞcient data (Investor Conference, April 28, 2004, www.net-ir.ne.jp/setumei/e45050404ebb/mms/start.html). Phase III trials in Japan were reinitiated for respiratory and other infections. The agent is in Phase II development in the United States and Europe for serious nosocomial infections,

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including hospital-acquired pneumonia, due to drug-resistant bacteria. Daiichi is also developing an injectable formulation. Sitaßoxacin exhibits activity similar to that of gemißoxacin against grampositive bacteria, and it has activity equivalent to or better than that of most ßuoroquinolones against gram-negative bacteria and other respiratory tract pathogens (Zhanel GG, 2002). SigniÞcantly, sitaßoxacin is more potent than other ßuoroquinolones against methicillin-resistant S. aureus (MRSA). It is equivalent to ciproßoxacin and better than most ßuoroquinolones against Pseudomonas aeruginosa, and more potent than ciproßoxacin against Klebsiella pneumoniae. It displays the highest activity of all ßuoroquinolones against anaerobes (Niki Y, 1999). A Phase II, randomized, open-label, multi-center study assessing the safety, tolerability, and efÞcacy of sitaßoxacin (400 mg once daily) compared with imipenem (imipenem/cilastatin, 500 mg three times daily) showed that sitaßoxacin is safe and effective in the treatment of hospitalized patients with pneumonia. Researchers noted a clinical cure in 32 (out of 35) subjects receiving sitaßoxacin and mild-to-moderate drug-related adverse events in 5 subjects. The most common adverse events reported were injection-site reactions and GI effects (Feldman C, 2001). In vitro activity of sitaßoxacin and other new ßuoroquinolones was compared with a number of control drugs in the same class (such as ciproßoxacin and levoßoxacin) by reference dilution tests against 2,156 recent U.S. clinical isolates of S. pneumoniae, H. inßuenzae, and Moraxella catarrhalis. All the ßuoroquinolones demonstrated excellent in vitro activity against these three pathogens, and sitaßoxacin exhibited one of the highest potencies, at 0.5 µg/mL (Deshpande LM, 2000). All the isolates of H. inßuenzae and M. catarrhalis were inhibited by the investigational and comparator ßuoroquinolones at ≤0.5 µg/mL, irrespective of their beta-lactamase-producing abilities. In a second study, sitaßoxacin demonstrated the highest in vitro potency against H. inßuenzae compared with gemißoxacin, garenoxacin, levoßoxacin, and moxißoxacin (Biedenbach DJ, 2003). Cephalosporins Overview. Cephalosporins have traditionally been one of the most commonly used antibacterial classes for the treatment of respiratory tract infections. In recent years, drug development within this class has been limited owing to competitiveness, the lack of novel candidates, and high development costs relative to return. Cephalosporins differ widely in their spectrum of activity, susceptibility to βlactamases produced by bacteria, and serum half-life. These agents are highly successful agents for the treatment of RTI; however, they need to be combined with a macrolide for coverage of atypical organisms (e.g., such as Mycoplasma). Dong-A pharmaceuticals in South Korea is developing DA-7101, indicated for the potential treatment of respiratory and urinary tract infections. It is currently reported to be in Phase III trials in South Korea. Because of the lack of available data on its development status, DA-7101 will not be discussed in further detail here.

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Mechanism of Action. Cephalosporins, like the penicillins, bind to penicillinbinding proteins (PBPs) via their β-lactam ring structure and prevent bacterial cell-wall formation. By interrupting cell-wall formation, cephalosporins induce cell lysis and death. Ceftizoxime Alapivoxil. Ceftizoxime alapivoxil is an oral formulation of the marketed third-generation cephalosporin ceftizoxime (Fujisawa’s CeÞzox) that was being developed in Japan by Asahi Kasei. In August 2002, Asahi Þled an NDA in Japan. However, Asahi Kasei reported in February 2004 that the preregistration Þled in Japan was withdrawn because of insufÞcient data (Public Relations OfÞce, Asahi Kasei, July 2004). The compound was not in development in the United States or Europe. Ceftizoxime alapivoxil is a broad-spectrum antibacterial with activity against both gram-positive and gram-negative organisms. It acts by preventing bacterial cell-wall formation. Phase II clinical trials have been completed in which ceftizoxime alapivoxil 100 mg or 200 mg twice daily was orally administered to a total of 1,595 patients. High efÞcacy rates were observed in patients with chronic bronchitis (88.7%), bronchiectasis (91.2%), and pneumonia (88.2%). In general, clinical efÞcacy by dose was 86.5% for 100 mg and 84.1% for 200 mg, administered twice daily. The optimum dose was presumed to be 100 mg twice daily (Asahi Chemical Industry, Company Communication, March 17, 2000). The limited data available suggest that this orally active pro-drug of ceftizoxime may be a promising antibacterial agent. However, there are many highly effective oral cephalosporins (e.g., cefuroxime axetil). For ceftizoxime alapivoxil to compete in this segment, it would have to demonstrate excellent bioavailability, better dosing than existing cephalosporins, activity against resistant gram-positive strains, minimal side effects, and cost-effectiveness. Ketolides Overview. Ketolides are a new class of antibacterials specially designed to combat respiratory tract pathogens that have acquired resistance to macrolides. The ketolides are derivatives of the macrolide erythromycin A. Telithromycin (SanoÞAventis’s Ketek) is the Þrst and only member of this new class to be approved for clinical use. The ketolides exhibit good activity against gram-positive and some gram-negative organisms, but their main advantage is their excellent activity against erythromycin-resistant S. pneumoniae. Resistance to existing macrolide antibiotics creates an important commercial opportunity for entry of the “next-generation” macrolide antibiotics. Macrolides are currently a preferred class of antibacterials for Þrst-line treatment of RTIs because of their spectrum of activity and ease of use. However, as resistance to these agents increase, the ketolides could replace them in the RTI market. Mechanism of Action. Ketolides exert their antibacterial activity by interacting with the bacterial ribosome and inhibiting its ability to make new proteins.

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Ketolides bind to a portion of the rRNA on ribosomal proteins. Ketolides bind to a region of the ribosomes with higher afÞnity than macrolides. This binding could contribute to ketolide’s activity against macrolide-resistant strains (Poehlsgaard J, 2002). Cethromycin. Abbott and Taisho were developing the ketolide cethromycin (ABT-773), the lead compound from a series of macrolide antibacterials, for the potential treatment of bacterial infections and respiratory infections in particular. Abbott was reportedly looking for a licensing partner for cethromycin in the Unites States and Europe. In Japan, Abbott continued to support the development of ABT-773 by partner Taisho. However, in April 2004, Taisho announced it had suspended its collaboration with Abbott because of Taisho’s changing development strategy and priority in its prescription pharmaceutical business (Taisho Pharmaceutical, press release, April 22, 2004). Abbott has released no further information on a licensing partner to continue development of cethromycin. In vitro studies of cethromycin have found that the compound demonstrates very good activity against respiratory tract pathogens acquired in the community, including penicillin- and macrolide-resistant S. pneumoniae (Casellas J, 2002; Credito KL, 2001[a]; Dubois J, 2001; Shortridge VD, 2002). Cethromycin’s in vitro efÞcacy against macrolide-resistant S. pneumoniae is superior to that of telithromycin, and it has demonstrated bactericidal activity against S. pneumoniae and H. inßuenzae in experimental models for pneumonia in rodents (Michelow IC, 2001; Mitten M, 2001). Pharmacokinetic information on cethromycin is still emerging. A Phase I clinical trial involving 48 healthy male volunteers suggests that, for a single oral dose (100–1,200 mg), adverse events were mild to moderate in severity, with GI disturbances being the most commonly reported events. Streptogramins Overview. Streptogramins are produced by streptomycetes and are classiÞed as A or B compounds, based on their mechanism of action as outlined below. As a class, streptogramin A and B compounds are bacteriostatic when used separately. However, when used in combination, they can act in synergy to become bactericidal. They are active primarily against gram-positive bacteria (Allington DR, 2001). Streptogramin combinations are bactericidal (concentration-independent) against streptococci and staphylococci and are bacteriostatic against enterococci. While streptogramins are expected to be used in treating serious nosocomial infections caused by antibiotic-resistant, gram-positive bacteria, these agents also have a potential role for the treatment of severe cases of AECB in the hospital setting. In particular, as the prevalence of multidrug-resistant gram-positive bacteria increases, the streptogramins could play an important role as second-line agents. Quinupristin/dalfopristin (King Pharmaceuticals/SanoÞ-Aventis/Fujisawa’s Synercid), the Þrst globally available streptogramin, has proved effective in treating patients suffering from severe, multidrug-resistant bacterial infections.

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Mechanism of Action. Streptogramins enter bacterial cells by diffusion and bind to different sites on the 50S ribosomal subunit to form a stable drugribosome-drug ternary complex resulting in an irreversible inhibition of protein synthesis. Streptogramin A inhibits peptide chain elongation, and streptogramin B inhibits peptide bond formation. The synergistic bactericidal effect of the combination appears to result from conformational changes brought about in the peptidyl transferase center. XRP-2868. SanoÞ-Aventis is developing XRP-2868∗ , an oral streptogramin combination consisting of 30% RPR-202868 (a type B streptogramin) and 70% RPR-132552 (a type A streptogramin) for the potential treatment of AECB and other RTIs. In February 2004, XRP-2868 was in Phase I studies in the United States (SanoÞ-Aventis Web site, February 5, 2004). In September 2003, preclinical data from a study of the synergistic activity of RPR-202868 and RPR-132552 against S. aureus, S. pneumoniae, and H. inßuenzae were presented at the 43rd ICAAC meeting. The compounds showed a synergistic bacteriostatic effect against S. aureus. Against S. pneumoniae, the compounds showed a marked synergistic bactericidal effect. RPR-132552 was bactericidal against H. inßuenzae alone at concentrations over 2 mg/L, while RPR-202868 showed minimal activity (Bryskier AJ, 2003). In another study, the in vitro activity of RPR-202868 was compared with that of other antibacterial agents against S. pneumoniae and H. inßuenzae (Pankuch GA, 2003). Tested in 261 S. pneumoniae strains, XRP-2868 demonstrated potency similar to that of pristinamycin (SanoÞ-Aventis’s Pysotacine) and potency that is one to two times superior to that of quinupristin/dalfopristin. XRP-2868 also demonstrated superior potency against PRSP compared with azithromycin and clarithromycin. In addition, XRP-2868 demonstrated higher potency against H. inßuenzae when compared with azithromycin, clarithromycin, erythromycin, and pristinamycin. Peptidyl Deformylase (PDF) Inhibitors Overview. The dramatic rise in S. pneumoniae resistance to penicillin, macrolides, and other antimicrobials is concerning. While the ßuoroquinolones currently enjoy a low level of bacterial resistance in the community, experts expect resistance against this class to increase as they are more widely used, thereby limiting future antibacterial options. Consequently, there is considerable interest in the development of novel antimicrobials, such as peptide deformylase (PDF) inhibitors, that have the potential to be active against pathogens resistant to currently available antimicrobials. PDF is an enzyme involved in bacterial protein synthesis, and this target is highly conserved across bacterial species. Therefore, PDF inhibitors can potentially provide broad-spectrum activity. In addition, there appears to be little interaction with the human protein synthesis component; thus, the inhibitors are likely to have favorable selectivity to bacteria (and therefore safety in humans). PDF inhibitors are at an early stage of development but have generated much interest due to the need for novel, innovative antibacterials. Results from further

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trials will be needed to determine their efÞcacy and probable use in the future. Several companies, including Roche, are working on early-stage development of PDF inhibitors. Because of minimal available clinical trial information, Roche’s compounds are not covered in detail. Mechanism of Action. PDF is an essential bacterial metalloenzyme required for protein synthesis and thus represents a good target for antibacterial therapy. PDF inhibitors act on the PDF enzyme, disrupting protein maturation and inhibiting protein synthesis. Protein synthesis in bacteria, under normal conditions, is initiated by the amino acid formyl-methionyl-tRNA. Consequently, all nascent polypeptides are synthesized with N-formyl-methionine at the N terminus. The formyl group is removed by PDF during elongation of the polypeptide chain. Deformylation is also a prerequisite for protein maturation (Apfel CM, 2001). PDF inhibitors act on the enzyme PDF to prevent deformylation of the growing peptide chain and thus inhibit protein synthesis. BB-81384. Oscient Pharmaceuticals (formerly Genesoft/Genome Therapeutics), under license from Vernalis (formerly British Biotech), was developing a lead compound from a series of IV PDF inhibitors for the potential treatment of bacterial infections, and RTIs in particular. The agent was being explored as an IV therapy for potential application in AECB. However, in March 2004, Genome Therapeutics stated this Þrst-generation compound did not have the ideal spectrum of activity against common respiratory pathogens and that the company was exploring second-generation orally available PDF inhibitors with a greater potential to target the broader antibiotic market. Oscient Pharmaceuticals is currently investigating several preclinical oral PDF inhibitors for community-based RTIs. Several compounds have been identiÞed as having attractive antibacterial properties, including good activity against H. inßuenzae. BB-81384, a novel PDF inhibitor with good activity against S. pneumoniae in vitro, is the Þrst compound of this class to be proÞled for oral pharmacokinetics and has demonstrated oral anti-pneumococcal efÞcacy in mice (Gross M, 2004). Pharmacokinetic studies in mice showed that the agent has good oral bioavailability. BB-81384 was a potent and selective inhibitor of PDF with an IC50 approximately 10 nM against most S. pneumoniae pathogens. In models of lung and thigh infection, BB-81384 reduced the bacterial load. Dihydrofolate Reductase Inhibitors Overview. The dihydrofolate reductase (DHFR) inhibitors target bacterial DHFR, an enzyme involved in the folic acid pathway, thereby disrupting bacterial replication. DHFR inhibitors target both gram-positive and gram negativebacteria and hence could serve as broad-spectrum antibacterials. Trimethoprim/sulfamethoxazole (Roche’s Bactrim, Monarch’s Septra, generics), the most common folic acid biosynthesis inhibitor, was used in the past to treat RTIs such as mild pneumonia and sinusitis. However, the emergence of S. pneumoniae resistant to trimethoprim/sulfamethoxazole, concerns regarding sulfa hypersensitivity, and poor activity against atypical RTI organisms has limited its use. Newer drugs

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in this class are being designed that overcome current mechanisms of resistance and provide a better spectrum of coverage against RTI pathogens. These agents may become increasingly important over the next 10–20 years as resistance to current drug classes builds. Mechanism of Action. The folic acid pathway inhibitors act by blocking the action of the bacterial DHFR enzyme—an enzyme essential for the synthesis of thymine nucleotides. By inhibiting the synthesis of thymine nucleotides, DNA and RNA synthesis is prevented, thereby killing the bacteria. More speciÞcally, in thymine synthesis, folate donates a methyl group to uracil to make thymine. In the process, the folate molecule is oxidized and must be reduced by DHFR to allow another cycle of synthesis. If DHFR is inhibited, the absence of reduced folate causes a precipitous drop in the levels of thymine nucleotides, causing uracil to be erroneously incorporated in its place in growing DNA strands. Replication is eventually disrupted by this process, killing the bacteria (Quinlivan EP, 2000; Schneider P, 2003). Bacterial DHFR is distinct enough from human DHFR that drugs targeting the bacterial DHFR protein are generally not toxic to human cells. Iclaprim. Arpida licensed all commercialization rights to the DHFR inhibitor iclaprim (AR-100) from Roche. The drug has the potential to be used against serious antibiotic-resistant infections, including RTIs. The agent has completed Phase II trials in Europe in treatment of skin infections and was shown to be well tolerated and to have high clinical and microbiological response rates in these infections. While no information is available regarding its development for AECB, iclaprim’s spectrum of activity makes it a potential emerging candidate for the treatment of this indication.Previous clinical trials have evaluated an IV formulation of iclaprim, but Arpida is working to develop an oral formulation as well. Iclaprim targets a range of bacterial pathogens, including MRSA and vancomycin-resistant S. aureus (VRSA). Results from in vitro studies suggest that the potency of iclaprim against C. pneumoniae is similar to that of leading antibacterials used in RTIs, such as azithromycin and levoßoxacin (Kohlhoff SA, 2004). While the agent is being developed for skin infections, it demonstrated good activity against gram-positive and gram-negative pathogens involved in RTIs such as S. pneumoniae and H. inßuenzae as well as intracellular pathogens such as Chlamydia. Therefore, it has the potential to be developed for AECB. Availability in both oral and IV forms could give the agent an advantage in the hospital setting and for oral step-down therapy.

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Sethi S, et al. New strains of bacteria and exacerbations of chronic obstructive pulmonary disease. New England Journal of Medicine. 2002;347(7):465–471. Shah PM, et al. Levoßoxacin versus cefuroxime axetil in the treatment of acute exacerbation of chronic bronchitis: results of a randomized, double-blind study. Journal of Antimicrobial Chemotherapy. 1999;43(4):529–539. Shah SH, et al. Cefuroxime axetil in the treatment of bronchitis: comparison with amoxycillin in a multicentre study in general practice patients. British Journal of Clinical Practice. 1994;48(4):185–189. Shortridge VD, et al. Comparison of in vitro activities of ABT-773 and telithromycin against macrolide-susceptible and -resistant Streptococci and Staphylococci. Antimicrobial Agents and Chemotherapy. 2002;46:783–786. Siafakas NM, et al. ERS consensus statement. Optimal assessment and management of chronic obstructive pulmonary disease (COPD). European Respiratory Journal. 1995;8:1398–1420. Smith CB, et. al. Association of viral and Mycoplasma pneumoniae infections with acute respiratory illness in patients with chronic obstructive pulmonary diseases. American Review of Respiratory Diseases. 1980;121:225–232. Societe de Pneumologie de Langue Francaise (SPLF). Recommendations for the management of COPD of the French-Language Pneumological Society. Revue des Maladies Respiratoires. 1997;(suppl 2). Sohy C, et al. Acute exacerbation of chronic obstructive pulmonary disease and antibiotics: what studies are still needed? European Respiratory Journal. 2002;19:966–975. Soler M, et al. Bronchial microbial patterns in severe exacerbations of chronic obstructive pulmonary disease (COPD) requiring mechanical ventilation. American Journal of Respiratory and Critical Care Medicine. 1998;157:1498–1505. Song JH, et al. Spread of drug-resistant Streptococcus pneumoniae in Asian countries: Asian Network for Surveillance of Resistant Pathogens (ANSORP) study. Clinical Infectious Diseases. 1999;28:1206–1211. Soriano JB, et al. The proportional Venn diagram of obstructive lung disease: two approximations from the United States and the United Kingdom. Chest. 2003;124(2):474–481. Spencer S, et al. Impact of preventing exacerbations on deterioration of health status in COPD. European Respiratory Journal. 2004;23(5):698–702. Sperling MJ, Puopolo et al. EfÞcacy and safety of cefdinir in the treatment of patients with acute bronchitis. Clinical Therapeutics. 1996;18(4):626–634. Sprangler SK, et al. Time-kill studies on susceptibility of nine penicillin-susceptible and -resistant pneumococci to cefditoren compared with nine other b-lactams. Journal of Antimicrobial Chemotherapy. 1997;39:141–148. Stass H, et al. Pharmacokinetics and elimination of moxißoxacin after oral and intravenous administration in man. Journal of Antimicrobial Chemotherapy. 1999;43(suppl B):83–90. Stein GE, Havlichek DH. Sparßoxacin: potential clinical and economic impact in the treatment of respiratory infections. Pharmacotherapy. 1997;17:1139–1147. Stewart C, et al. Safety, tolerability and pharmacokinetics of BMS-284756, a novel desF(6)- quinolone, following 28 days of once daily oral dosing in healthy subjects. Presented at the 41st Interscience Conference on Antimicrobial Agents and Chemotherapy (ICAAC); December 2001; Chicago, IL. Abstract A-46.

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Community-Acquired Pneumonia

ETIOLOGY AND PATHOPHYSIOLOGY Introduction Pneumonia is deÞned as inßammation of the cells within the lung that results in the consolidation and Þlling of alveolar spaces with exudates. Unlike bronchitis, which typically involves the major airways, pneumonia affects the pulmonary parenchyma (i.e., the lung tissue). Clinically, however, many patients often have combined pathologies. Pneumonia has a variety of causes but is most commonly infectious in nature; occasionally, it is caused by chemicals and trauma. Pneumonia is often classiÞed into more speciÞc categories based on the type of infectious agent that is causing disease (bacterial versus viral or fungal pneumonia) or the setting where the infectious agent was acquired, whether in the hospital (nosocomial) or in the ambulatory setting (community-acquired). Here, the focus is on the bacterial causes of community-acquired pneumonia (CAP). (Pneumonia acquired in the nursing home setting is not generally classiÞed as CAP and is not discussed here.) In the International ClassiÞcation of Diseases, 9th Revision (ICD-9), the codes related to pneumonia are 480–486. In the ICD Tenth Revision (ICD-10), the corresponding codes are J12-J18. Etiology The upper respiratory tract is colonized by several bacterial species that make up the normal ßora. Although these commensal organisms can cause disease in certain situations (e.g., in immunocompromised persons), they generally serve a protective function by limiting the growth of other pathogenic bacteria in the Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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respiratory tract. In the lower respiratory tract (LRT), macrophages that can clear foreign particles reside within the alveoli and serve as the Þnal defense against infection. Pneumonia occurs when pathogenic organisms establish an infection in the LRT as a consequence of depressed host response and/or invasion of the LRT by highly virulent organisms. Classification of Pneumonia. CAP can be classiÞed according to the region of the lung that is affected or the spectrum of symptoms a patient presents. Both classiÞcations indicate the level of disease severity and direct clinicians to the type of therapy to employ. Three forms of pneumonia are typically classiÞed according to the region of the lung involved: • •

•

Bronchopneumonia, which usually involves consolidated areas of acute inßammation of the small airway alveoli. Lobar pneumonia, a more extensive disease than bronchopneumonia in which consolidation occurs in large portions of a lobe or in an entire lobe. This form of CAP can be quite severe and can have an impact on the future function of the affected tissue. Interstitial pneumonia, characterized by inßammation in the interstitium, the soft tissue between the alveoli.

Likewise, pneumonia can be classiÞed clinically as “typical” or “atypical,” according to the presenting symptoms. The classical clinical presentation of typical pneumonia includes fever, cough with sputum production, pleuritic pain (pain on inhalation), and chills. Atypical pneumonia is associated with any number of systemic symptoms, including myalgia (muscle aches), and headache. With either type of pneumonia, confusion and personality changes attributed to hypoxia (below normal levels of oxygen in the blood and tissues) can occur in the elderly (Martin RE, 1991). Often, patients with atypical pneumonia may have only mild respiratory symptoms (Monsieur I, 1997). Pneumonia due to infection with so-called atypical pathogens can present as typical pneumonia, and atypical pneumonia can be caused by bacterial pathogens thought to cause only typical pneumonia. Presentation of disease may depend not only on the pathogen causing the disease but also on the host. Common Etiologic Pathogens. Many bacteria have been shown to cause CAP, but researchers and clinicians identify several bacterial species as the most common causes of the disease. However, epidemiological studies investigating the etiology of the disease show that in the majority of cases, pathogen identiÞcation is not readily attainable. Pathogens are classiÞed as typical or atypical based on the spectrum of symptoms associated with them (described in the preceding section). In most studies, Þve pathogens (discussed in the following sections) have been found to account for approximately 90% of all CAP (American Thoracic Society, 2001). Streptococcus pneumoniae is the bacterium most commonly implicated in CAP, followed by other frequently isolated, typical etiologic agents

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such as Haemophilus inßuenzae and Staphylococcus aureus (Mandell LA, 2003). In recent years, however, atypical respiratory pathogens have been observed with increasing frequency in CAP patients (Marrie TJ, 1996); the pathogens most often implicated are Mycoplasma pneumoniae, Chlamydia pneumoniae, and Legionella species. Other pathogens implicated in CAP include the respiratory viruses (inßuenza A and B, parainßuenza viruses, and respiratory syncytial virus) and anaerobes, which tend to occur in alcoholic patients. Less common etiologies include Chlamydia psittaci and Coxiella burnetii (Lieberman D, 1999). Atypical pathogens tend to be a more common cause of CAP in younger patients and are generally associated with a less severe pneumonia. However, they can affect all age-groups and cause severe disease (especially in the immunosuppressed) as well as mild-to-moderate illness (Mandell LA, 2000). The pathogens discussed in this section are some of the most commonly reported causes of bacterial CAP in the major pharmaceutical markets. Table 1 summarizes the prevalence of each of these pathogens in CAP and the types of pneumonia they cause. The bacterial pathogens are classiÞed as “gram-positive” or “gram-negative” based on their response to a staining technique called Gram’s stain. Gram’s stain distinguishes bacteria based on differences in physiological and structural properties of the cell wall, speciÞcally its ability to retain the crystal violet dye used in the staining procedure. Bacteria that retain the stain are classiÞed as gram-positive; bacteria that do not retain the stain are classiÞed as gram-negative. The distinction between gram-positive and gram-negative bacteria is important because gram-positive bacteria are generally more sensitive than gram-negative TABLE 1. Characteristics and Frequency of Select Pathogens in Community-Acquired Pneumonia

Pathogen Streptococcus pneumoniae Haemophilus influenzae Moraxella catarrhalis Mycoplasma pneumoniae Chlamydia pneumoniae Legionella pneumophila Klebsiella pneumoniae Staphylococcus aureus

Type of Pneumonia Caused

Type of Symptoms Usually Present

Gram Stain Characteristic

Frequencya (%)

Gram-positive cocci

8–46

Lobar pneumonia

Typical

Gram-negative coccobacilli Gram-negative cocci Does not stain well with Gram-stain Gram-negative coccobacilli Gram-negative bacilli Gram-negative bacilli Gram-positive cocci

2–12

Typical

0–2

Lobar or bronchopneumonia Bronchopneumonia

Typical

2–29

Bronchopneumonia

Atypical

6–18

Interstitial pneumonia Lobar or bronchopneumonia Lobar pneumonia

Atypical

1–16 0–8 0–3

Lobar or bronchopneumonia

Atypical Typical Typical

a Frequency indicates the percentage of patients diagnosed with community-acquired pneumonia (CAP) caused by the individual pathogen in a series of epidemiological studies.

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bacteria to growth inhibition by antibiotics that target cell-wall synthesis and to attack by phagocytosis. The following sections highlight some of the more important aspects of disease associated with each pathogen (Marrie TJ, 1998). Streptococcus Pneumoniae. S. pneumoniae is the most commonly reported cause of bacterial CAP. Pneumonia caused by this gram-positive bacterium, also known as pneumococcal pneumonia, is often lobar in nature (see “ClassiÞcation of Pneumonia”) and most commonly presents as typical pneumonia. S. pneumoniae is encased in a polysaccharide capsule that resists phagocytosis by macrophages; therefore, the antibody-dependent killing mechanisms (i.e., the complement cascade, a series of proteins that can destroy bacteria) are important in the immune system’s defense against this organism. S. pneumoniae also contains the enzyme IgA1 protease, which cleaves the IgA in the upper respiratory tract, thereby allowing the bacterium to survive in that environment. This ability accounts for the fact that S. pneumoniae commonly resides in the upper respiratory tract. It is important to note that more than 90 serotypes of S. pneumoniae have been identiÞed (based on the polysaccharide capsule); therefore, several serotypes must be included in vaccine candidates to confer protection against pneumococcal pneumonia. Currently marketed vaccines include 23 serotypes of pneumococcus, covering more than 80% of types that cause invasive disease (Konradsen HB, 2002). Haemophilus Influenzae. H. inßuenzae, a gram-negative bacterium, can cause either lobar or bronchopneumonia (see “ClassiÞcation of Pneumonia”) and is usually associated with typical pneumonia symptoms. This pneumonia is often marked by the sudden onset of pleuritic chest pain, possibly preceded by rhinitis. In rare cases, necrosis of the lung tissue with eventual abscess formation can occur with this type of pneumonia and result in scarring. H. inßuenzae is a commensal organism in the upper respiratory tract; it tends to cause infection most commonly when it invades the lower respiratory tract of patients with underlying respiratory illness, such as chronic obstructive pulmonary disease (COPD). Since the 1970s, H. inßuenzae resistance to β-lactam antibiotics such as amoxicillin and ampicillin has been on the rise. H. inßuenzae resistance is mediated primarily through production of the enzyme β-lactamase, which inactivates certain β-lactam antimicrobials, including amoxicillin and ampicillin. In the United States, 30–40% of H. inßuenzae isolates produce β-lactamase (Pfaller MA, 2001; Gordon, KA, 2003; Hoban DJ, 2001). Risk factors for the development of amoxicillin-resistant infection with H. inßuenzae include recent hospitalization and outpatient prescription of β-lactam antibiotics within the previous three months (Seaton RA, 2000). Mycoplasma Pneumoniae. Pneumonia resulting from M. pneumoniae is usually a bronchopneumonia that presents atypically; nonspeciÞc symptoms such as headache, myalgia, and malaise occur in up to 70% of patients. Pneumonia with rash can occur with M. pneumoniae. The onset of this pneumonia is gradual, with a prominent cough. Mycoplasma species have a three-layered cell membrane but

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no true cell wall; for that reason, they do not retain color well with Gram’s stain (although the bacteria will show gram-negative staining). Furthermore, antibiotics targeted at the bacterial cell wall (e.g., β-lactams) are not effective against these bacteria. Like S. pneumoniae, M. pneumoniae is frequently carried in the nasopharynx of healthy persons. Chlamydia Pneumoniae. C. pneumoniae is an intracellular, gram-negative pathogen that typically causes an interstitial pneumonia (see “ClassiÞcation of Pneumonia”). It presents as an atypical pneumonia, and symptoms are frequently mild and nonspeciÞc, such as fever and cough. Unlike many other bacterial species, C. pneumoniae does not contain a peptidoglycan layer within its cell wall, which is the target for the β-lactam antibiotics. Therefore, these drugs are ineffective against this organism. Legionella Pneumophila. Researchers have identiÞed more than 40 species in the Legionellaceae family and more than 60 serogroups. Several species can cause disease, but L. pneumophila is the most common cause of pneumonia. The infection is also known as Legionnaire’s disease because it was Þrst recognized during an outbreak among attendees of an American Legion convention. This gram-negative bacterium can cause either lobar pneumonia or bronchopneumonia and is usually associated with atypical symptoms. Upper respiratory tract symptoms are not very common; typically, fever and minimal cough are present. In 25–50% of patients, L. pneumophila pneumonia is accompanied by gastrointestinal (GI) symptoms such as anorexia, nausea, and vomiting. Pneumonia caused by L. pneumophila can be particularly severe compared with pneumonia caused by the other common pathogens: some outbreaks have a mortality rate exceeding 60%. L. pneumophila is spread by inhalation of infected, water-borne aerosols and is frequently associated with contaminated water systems, such as air conditioners and cooling towers. Because it is an intracellular pathogen that replicates within macrophages and monocytes, the cell-mediated immune response is particularly important in clearing this infection. Gram-Negative Bacilli. Gram-negative bacilli (GNB), common commensal organisms in the GI tract, can cause pulmonary disease when aspirated into the respiratory tract. Conditions that compromise the cough and gag reßexes, such as alcoholism and diabetes, increase the likelihood of aspiration and are risk factors for development of GNB pneumonia. The most common pneumonia-causing pathogen in this group is Klebsiella pneumoniae, which causes lobar pneumonia that is frequently characterized by chest pain, sudden fever, and bloody sputum. Disease is often severe and aggressive, marked by necrosis and abscess formation and signiÞcant consolidation on X ray. Pseudomonas aeruginosa, also a GNB, rarely causes CAP but is notable because it is often difÞcult to treat, owing to its resistance to many antibiotics. Staphylococcus Aureus. CAP caused by S. aureus is uncommon (in contrast to hospital-acquired, or nosocomial, pneumonia), but failure to suspect S. aureus infection and delays in appropriate therapy can be fatal. Although 15–30% of adults are nasal carriers of this gram-positive bacterium, resultant CAP is rare

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and presumably requires an underlying host defect. Indeed, CAP associated with S. aureus typically occurs as a secondary infection to inßuenza or viral pneumonia or in patients with focal infections that spread through the bloodstream (e.g., secondary to endocarditis [infection of the heart valves]). Staphylococcal pneumonia is often associated with pleural effusions, and lung abscesses develop in 25% of patients. Typical symptoms include sudden onset of pleuritic chest pain, fever, and cough. This bacterium is often associated with severe pneumonia and a high incidence of complications, especially in patients with compromised immune systems (e.g., neonates, the elderly). Viruses. Although bacteria cause the vast majority of adult cases of CAP, pneumonia in infancy and childhood is frequently viral in origin. In fact, in patients younger than Þve years old, the majority of cases of CAP are viral. The most common etiologic agents are respiratory syncytial virus (RSV), parainßuenza virus, inßuenza virus, and adenovirus. Children frequently present with upper respiratory symptoms such as rhinorrhea (runny nose) or pharyngitis (sore throat) prior to developing a cough. Because it is very difÞcult to differentiate viral from bacterial infections, physicians treat most infections with antibacterial agents. Pathophysiology Host defenses such as anatomic, functional, and mechanical barriers serve to protect the intact bronchial tree from invading organisms. Some key factors in the pathophysiology of CAP are alterations in host defense mechanisms, invasion by virulent microorganisms, and the quantity (i.e., inoculum) of the invading microbes. CAP is usually acquired by inhalation or aspiration of pulmonary pathogenic organisms into a lung segment or lobe. Much less commonly, CAP may result from a secondary bacteremia from a distant source—for example, CAP secondary to Escherichia coli urinary tract infection and/or bacteremia. Defense of the Respiratory Tract. Several host defense mechanisms combine to create a generally sterile environment within the lung (Figure 1). Protection of the lung from infection begins with nonspeciÞc defense mechanisms in the upper respiratory tract. Large inhaled particles are frequently caught in the nasal hair. Particles that pass through the nasal hair may be Þltered out in the turbinate bafßes (curved bones within the nasal cavity), which catch large particles and prevent them from moving lower in the respiratory tract. Mucus lines the entire respiratory tract and traps potential pathogens, which can then be expelled by various means, including the cough reßex. The ciliated cells of the trachea can transport particles trapped in mucus from below the larynx upward until they can be swallowed for destruction in the GI tract. The epiglottis prevents ßuids and food from entering the respiratory tract during swallowing, thereby preventing the descent of pathogens into the lower respiratory tract. The salivary glands protect the respiratory tract as well by secreting several nonspeciÞc bactericidal proteins, including immunoglobulin A (IgA) and lysozyme (Fish D, 2001). Despite the many mechanisms protecting the lower respiratory tract, pathogens can inÞltrate the lung and establish infection. Bacteria typically reach the lung

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FIGURE 1. Diagram of the respiratory tract and select immune defenses

when infected aerosols or aspirated secretions from the upper respiratory tract are inhaled or, much less commonly, through the bloodstream from another site of infection. Aspiration is the most common route of infection that occurs in the respiratory tract. Normally, the host defense mechanism clears out any aspirated foreign microorganisms from the lower respiratory tract. An infection usually occurs when one component of the defense mechanism is not functioning properly. This malfunction results in microbial colonization of the upper respiratory tract (Schmitt SK, 2004). Once infection is established, immune responses develop, leading to a suppurative (pus-forming) reaction. In pneumonia, the alveoli, or airspaces of the lung, Þll with exudates as a result of the immune response to inßammation (discussed later). The pathology that develops in the lung during the course of pneumonia is an important clinical manifestation that, to some extent, helps to differentiate possible causative pathogens. Cellular immunity and humoral immunity also play important roles and can help protect the body from bacterial organisms.

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The Immune Response. The alveolar macrophages secrete numerous cytokines, which recruit neutrophils to the site of infection. Neutrophils are the primary cells responsible for the destruction of bacterial pathogens in the lung; they are the abundant immune-cell type in the majority of infectious pneumonias. The antibody response to infection, typically involving the development of IgM during the initial acute infection, followed by IgG, is one of the most important pathogen-speciÞc responses and plays an important role in diagnostic evaluation. The complement cascade is often mediated by the development of antibodies and is important in the destruction of pathogens. For intracellular pathogens (e.g., Legionella pneumophila), which replicate within the alveolar macrophages and are not exposed to neutrophils in the extracellular environment during replication, the cell-mediated immune system (which targets intracellular pathogens) is responsible for destroying infected cells. Both CD4+ T cells and CD8+ T cells are involved in this type of response, but the exact nature of this response is unclear. Clinical Presentation of Disease. As previously discussed, the classical clinical presentation of “typical” pneumonia includes fever, cough with sputum production, pleuritic pain, and chills, while “atypical” pneumonia is associated with any number of systemic symptoms, including myalgia (muscle aches) and headache. Confusion and personality changes attributed to hypoxia (belownormal levels of oxygen in gases, arterial blood, and tissues) can also occur in elderly patients (Martin RE, 1991). Patients with atypical pneumonia may often have only mild respiratory symptoms (Monsieur I, 1997). In addition, pneumonia due to infection with so-called atypical pathogens can present as typical pneumonia, and atypical pneumonia can be caused by bacterial pathogens thought to cause only typical pneumonia. Presentation of disease may depend not only on the pathogen causing the disease but also on the host. Elderly patients with pneumonia, for example, can present with very mild respiratory symptoms because the cough reßex is diminished. Patients may also appear sunken and dehydrated because of their inability to eat and drink, which results from an inability to breathe properly. Therefore, the clinical presentation of pneumonia is often of little diagnostic signiÞcance in terms of differentiating among the numerous possible bacterial pathogens that can cause disease. Complications of Pneumonia. Common, signiÞcant, and life-threatening complications associated with CAP are sepsis (a syndrome resulting from bacteremia, characterized by marked hypotension progressing to multiple organ failure), septic shock, respiratory failure, pulmonary embolism, congestive heart failure, cardiac arrhythmias, myocardial infarctions, renal insufÞciency, stroke or transient ischemic attack, and invasive suppurative infections such as empyema (purulent ßuid in pleural tissue) and brain abscess. The progression to severe pneumonia can result in respiratory failure requiring admission to the intensive care unit (ICU). The combination of bacterial replication and immune response can lead to severe tissue destruction, which can result

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in the accumulation of scar tissue and, consequently, permanently diminished lung function. Diffuse damage within the alveoli can prompt acute respiratory distress syndrome (ARDS), which is often fatal. One of the most serious complications of pneumonia is the progression to bacteremia (the presence of bacteria in the blood) because pathogens that have spread to the blood can cause sepsis and eventually death. Bacteremic pneumonia occurs in approximately 10–20% of pneumonia cases caused by S. pneumoniae and is associated with a two- to Þvefold increase in mortality rate (Fine MJ, 1996). Mortality from pneumonia usually results from the complications of sepsis. Although current treatment is very effective for CAP, the mortality rate remains high primarily because once sepsis develops, patients frequently become refractory to treatment. The mortality for bacterial CAP ranges from 1% for outpatients to 35% or greater for hospitalized patients (for example, in the ICU). An estimated 90% of deaths resulting from pneumonia occur in people aged 65 or older (Kaplan V, 2003). Risk Factors. Pulmonary illnesses can cause structural damage within the respiratory tract that diminishes the efÞcacy of the natural defense mechanisms described previously and can change the normal ßora in the respiratory tract such that more virulent pathogens can ßourish. Several risk factors are associated with increasing susceptibility to pneumonia. Smoking increases the risk of acquiring pneumonia by destroying both the ciliated cells and alveoli. Similarly, viral infections can damage the ciliated epithelium and alveoli; therefore, prior and ongoing viral respiratory infections can predispose a person to bacterial pneumonia. Toxic fumes, industrial smoke, and other air pollutants may also damage cilia function. Certain comorbidities increase the likelihood of acquiring pneumonia by compromising the respiratory tract defense systems that prevent and Þght infection. In addition to leaving hosts more susceptible to development of disease, these comorbidities are associated with progression to more severe disease. Alcoholics are at risk for developing pneumonia because while intoxicated, they have a decreased gag reßex, thus increasing the chance of aspi rating pathogens. Alcohol also weakens the immune response by restricting the mobilization of neutrophils. Consequently, CAP in the chronic alcoholic is characterized by a more severe clinical course and higher mortality rate than CAP in nonalcoholics (Chen MZ, 2001). One of the subpopulations most susceptible to pneumonia is the elderly; their decreased cough reßex and weaker immune system can delay and disrupt the response to infection (Masters PA, 1998). Similarly, immunocompromised people, such as AIDS patients and patients without a spleen, frequently contract pneumonia. Because of their diminished ability to Þght infections, these patients are susceptible to bacterial pathogens that would not cause disease in immunocompetent patients. Patients with COPD commonly have infectious exacerbations that place them at risk for developing pneumonia.

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COMMUNITY-ACQUIRED PNEUMONIA

TABLE 2. Mechanisms of Resistance Against Select Antibacterial Classes, 2004 Drug Class Penicillins, carbapenems, and cephalosporins (β-lactams)

Macrolides and ketolides

Fluoroquinolones

Tetracyclines

Glycopeptides

Mechanism Expression of β-lactamases (which hydrolyze β-lactam ring) is found in both gram-negative and gram-positive bacteria, and at least 340 β-lactamases have been described thus far. Sophisticated restructuring of the penicillin-binding proteins (PBPs) can result in penicillin resistance. Modifications in outer-membrane permeability can limit efficacy of the β-lactams. Alterations in the ribosomal binding site may be plasmid- or chromosomally mediated. Enzymatic modification of 50S ribosome target. Alterations in antibiotic transport (overexpression of efflux pumps) that expel antibiotics out of the bacterium. Production of enzymes that inactivate the antibiotic (e.g., hydrolysis by esterases) limits their efficacy. Decreased permeability limits the intracellular concentration. Mutations in genes coding for DNA gyrase and/or DNA topoisomerase IV decrease susceptibility. Overexpression of efflux pumps expels fluoroquinolones from the bacterium. Resistance to the tetracyclines can develop rapidly via the selection of isolates that lack permeability to the drug. Changes in the bacterial cell membrane that prevent accumulation of the drug within the cell (efflux). Production of enzymes that protect the ribosome complex limits efficacy of tetracyclines. Modification of the D-alanyl-D-alanine components of the cell wall into D-alanyl-D-lactate components decreases susceptibility to glycopeptides.

In children, the major risk factor for developing pneumonia is prior, usually viral infection of the upper respiratory tract. The tremendous increase in secretions, an increase in the density of colonizing bacteria, and impairment of the mucociliary apparatus lead to aspiration and pneumonia. Antimicrobial Resistance. The prevalence in the community of bacterial resistance to common antibiotic classes has risen dramatically over the past two to three decades. Pathogens can develop antibiotic resistance through many mechanisms. These mechanisms include production of enzymes that degrade the antibiotic (e.g., β-lactamases), modiÞcation of drug targets (e.g., modiÞed penicillin-binding proteins and ribosomes that limit the activity of penicillins and macrolides, respectively), and expression of membrane proteins that pump the antibiotic out of the bacteria. Table 2 presents the mechanisms by which bacteria become resistant to the various classes of available antibiotics. Alterations in or mutations of the bacterial chromosomes were the Þrst identiÞed mechanisms of resistance, but resistance is more frequently acquired through transfer of genetic material from one bacterium to another. One of the most worrisome mechanisms of resistance involves the ability of bacteria to exclude antimicrobial agents from the cell. Some

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gram-negative and gram-positive pathogens have an efßux pump that can remove drugs fast enough to protect the bacteria from the drug and its effects. Resistance to an antibiotic may also result from a change or alteration in the target site at which the drug acts. Some strains of bacteria alter the protein composition of their outer cell membrane to prevent a drug from reaching its target site. Metabolic bypass is another common way in which bacteria can become resistant to antibiotics; in this case, the antibiotic interferes with the metabolic pathway by blocking or interfering with a critical step. Some resistant bacteria produce enzymes that cleave or alter the molecular structure of an antibiotic, thereby rendering it ineffective. In vitro susceptibility testing is the basis for classifying bacterial strains as either resistant or sensitive. Most organisms tested in the microbiology laboratory are reported to be either sensitive or resistant on the basis of breakpoints determined by the National Committee for Clinical Laboratory Standards. Susceptibility of strains is reported as sensitive, intermediately sensitive/resistant, or resistant based on the minimal inhibitory concentration (MIC) required by the antibacterial to exert its effect. The clinical relevance of decreased antibiotic susceptibility is controversial. Trials have demonstrated conßicting results with respect to impact on outcome of infection with a resistant pathogen (Rothermel CD, 2004; Song JH, 2004). In many regions, high-dose amoxicillin (3 grams per day) is commonly used to overcome community levels of resistance. Excessive and routine use of antibiotics has contributed to the increase in antibiotic resistance by selectively encouraging the survival and grow of resistant strains of bacteria. Hospitals and surgeons may promote antibiotic resistance through the routine and sometimes overextended use of prophylactic antibiotics with surgery. Patients contribute to antibiotic resistance when they do not take the drugs according to directions. An insufÞcient dose of antibiotic usually fails to kill all the disease-causing bacteria. The survivors are the most resistant strains, which may later cause infectious illnesses that are difÞcult to treat. CURRENT THERAPIES Overview Many highly effective agents are available for the treatment of bacterial community-acquired pneumonia (CAP) and other community-acquire respiratory tract infections (RTIs). Currently marketed antibiotics for CAP demonstrate similar efÞcacy rates in clinical trials, and these agents have generally achieved clinical symptom resolution in 85–95% of trial participants. The drugs differ mainly in their spectrum of activity (the organisms against which the agent is effective), side-effect proÞles, dosing schedules, and dose form availability.Broad-spectrum antibiotics are effective against a wide range of pathogens; narrow-spectrum antibiotics tend to be efÞcacious against a narrower range of bacterial species, generally either gram-positive or gram-negative organisms (see Table 1 for characteristics and frequency of select pathogens involved in CAP). Table 3 summarizes the leading antibiotic therapies currently available to treat CAP; Table 4 summarizes the mechanisms of action of the major classes of antibiotics.

520

TABLE 3. Current Therapies Used for Community-Acquired Pneumonia, 2004 Agent Penicillins Amoxicillin

Company/Brand GSK’s Amoxil, generics

US, F, G, I, S, UK, J

GSK’s Ceftin/Zinnat/Oracef BMS’s Cefzil/Procef/Bisoral Pfizer’s Vantin, Sanofi-Aventis’s Orelox, Sankyo’s Banan Abbott/Aventis’ Claforan, generics Roche’s Rocephin, generics Abbott’s Omnicef, Fujisawa’s Cefzon

250–500 mg PO bid 500 mg PO bid 200 mg PO bid

US, F, G, I, S, UK, J US, I, S, UK US, F, G, I, S, UK, J

1–2 g IV tid

US, F, G, I, S, UK, J

1–2 g IV qd 300 mg PO bid

US, F, G, I, S, UK, J US, J

Merck’s Primaxin

1–2 g IV q6h

US, F, G, I, S, UK, J

Abbott’s Erythrocin, Eli Lilly/Shionogi’s Ilosone, generics Abbott’s Biaxin/Biaxin XL/Klacid, Taisho’s Clarith Pfizer’s Zithromax/Zitromax

250 mg PO qid

US, F, G, I, S, UK, J

250–500 mg PO bid, or 1,000 mg PO qd 500 mg PO loading, then 250 mg PO qd

US, F, G, I, S, UK, J

GSK’s Augmentin, Augmentin ES, Augmentin XR, generics

Piperacillin/tazobactam

Wyeth/Taisho’s Zosyn/Tazocin

Cefotaxime Ceftriaxone Cefdinir Carbapenems Imipenem/cilastatin Macrolides Erythromycin

Clarithromycin Azithromycin

Availability

500–1,000 mg PO tid or 875 mg PO bid 500/125 mg PO tid or 875/125 mg PO bid or 2,000/125 mg PO bid 3.375 mg IV q6h

Amoxicillin/clavulanate

Cephalosporins Cefuroxime axetil Cefprozil Cefpodoxime proxetil

Daily Dose

US, F, G, I, S, UK, J US, F, G, I, S, UK, J

US, F, G, I, S, UK, J

TABLE 3. (continued) Agent Ketolides Telithromycin

Company/Brand

Daily Dose

Availability

SanofiAventis/Fujisawa/Sankyo’s Ketek

800 mg PO qd

US, F, G, I, S, UK, J

Johnson & Johnson’s Levaquin, Sanofi-Aventis’s Tavanic, Daiichi’s Cravit Bayer Schering-Plough’s Avelox BMS’s Tequin, Kyorin/Dainippon’s Gatiflo

500–750 mg PO or IV qd

US, F, G, I, S, UK, J

400 mg PO or IV qd

US, F, G, I, S, UK

400 mg PO or IV qd

US, G, J

Pfizer’s Vibramycin, generics

200 mg PO loading, then 100 mg PO qd

US, F, G, I, S, UK, J

Schering-Plough’s Garamycin, generics

3–5 mg/kg IV daily in divided doses tid

US, F, G, I, S, UK, J

Glycopeptides Vancomycin Teicoplanin

Lilly’s Vancocin, generics Sanofi-Aventis’s Targocid

500 mg IV q6h 400 mg IV loading, then 200 mg IV qd

US, F, G, I, S, UK, J F, G, I, S, UK, J

Oxazolidinones Linezolid

Pfizer’s Zyvox

600 mg PO or IV bid

US, F, G, I, S, UK, J

Fluoroquinolones Levofloxacin

Moxifloxacin Gatifloxacin Tetracyclines Doxycycline Aminoglycosides Gentamicin

IV = Intravenously; PO = Orally; qid = Four times daily; bid = Twice daily; tid = Three times daily; qd = Once daily. US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan. BMS = Bristol-Myers Squibb; GSK = GlaxoSmithKline.

521

522

COMMUNITY-ACQUIRED PNEUMONIA

In general, public health ofÞcials and thought leaders have promoted the use of narrow-spectrum agents whenever possible to limit the development of antibiotic resistance. However, broad-spectrum agents are more commonly used because therapy is largely empiric and physicians tend to be more comfortable with broader coverage. Several types of measures are used to assess the efÞcacy of antibiotics in vitro and in clinical trials. In vitro, the minimum inhibitory concentration (MIC) is the minimum concentration of antibiotic required to inhibit growth of a bacterium (also used to measure decreased susceptibility to antibiotics). In clinical trials, the clinical response rate, clinical success rate, or clinical cure rate refers to improvement in subjective symptoms and/or objective measures such as chest X ray. Bacterial eradication rate refers to the elimination of pathogens in sputum and/or in respiratory tract samples. Choice of Antibiotic Therapy. The choice of antibiotic therapy for CAP is based on factors such as suspected etiologic pathogen, age and comorbidities, treatment setting (community or hospital), ability to comply with oral therapy, and severity of disease. Likewise, spectrum of coverage, side-effect proÞle, safety, dosing, drug interactions, and cost have an important inßuence on drug selection. Pharmacokinetics (e.g., absorption, distribution and tissue penetration, metabolism, and elimination) and pharmacodynamics (e.g., mechanism of action, rate at which a bactericidal or bacteriostatic effect occurs) have a direct bearing on clinical efÞcacy and can be important factors in differentiating an antibacterial agent in the minds of clinicians. Because many CAP patients must be hospitalized, an important product attribute is availability in oral and intravenous (IV) forms. Ideally, products should have high oral bioavailability so that doses are interchangeable between the two forms. Products with high bioavailability allow physicians to prescribe oral doses conÞdently because they can be sure of achieving high serum concentrations. Agents with high bioavailability and IV and oral formulations are also beneÞcial in the hospital setting because they can accelerate IV-to-oral transition and hospital discharge. In recent years, the rising prevalence of drug resistance in the community has begun to inßuence prescribing choice by heightening concerns about antibiotic overuse and misuse (discussed further on). Likewise, health care costs play an increasing role in physician choice of antibiotic. An antibiotic’s ability to kill a pathogen (bactericidal activity) or to inhibit its growth (bacteriostatic activity) typically depends on the organism involved. In the presence of a fully functioning immune response, a drug will help to cure the infection by inhibiting the growth of the infecting organism. When host defenses are lacking (i.e., in immunocompromised patients such as AIDS or cancer patients), or when the infecting organism is protected from the immune response (e.g., by a polysaccharide capsule), curing the infection depends almost entirely on the drug. In such cases, therefore, bactericidal activity is required. Antibiotic Resistance. Antimicrobial resistance in Streptococcus pneumoniae and other CAP pathogens has progressed at an alarming rate. Approximately one-third of pneumococci exhibit reduced susceptibility to penicillin (i.e., higher

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TABLE 4. Mechanism of Action of Drugs Commonly Used in the Treatment of Community-Acquired Pneumonia Drug Class

Mechanism of Action

Penicillins and cephalosporins (β-lactams)

• β-lactams bind to penicillin-binding proteins (PBPs), preventing bacterial cell-wall formation. • Structurally different β-lactams target different PBPs; many broad-spectrum β-lactams bind to more than one PBP, enhancing overall potency. • Different β-lactams cause a variety of biochemical, morphological, and antibacterial effects within the same bacterial species. • Unrelated species of bacteria respond differently to the same β-lactam; some are both killed and lysed, and others respond with a reversible inhibition of growth. • The mechanism by which cell lysis and death occurs is complex; PBPs are involved in numerous roles, including maintenance of structural integrity, determination of cell shape, cell division, induction of capsule synthesis, phage resistance, and regulation of autolysis. • β-lactams may be bacteriostatic or bactericidal in effect. • Organisms that are metabolically inactive and those lacking bacterial cell walls (e.g., Mycoplasma) are not susceptible to β-lactams.

Macrolides and ketolides

• Macrolides and ketolides act by binding to ribosomal RNA (rRNA) in the 50S subunit of the ribosome. • These drugs are bacteriostatic but can be bactericidal in highly susceptible bacteria or at high concentrations.

Fluoroquinolones

• Fluoroquinolones interact with two related yet distinct targets within the bacterial cell: DNA gyrase and topoisomerase IV. • DNA gyrase and topoisomerase IV are vital enzymes involved in bacterial DNA replication. • These agents are potent inhibitors of nucleic acid synthesis. • The exact nature of the interaction of quinolones with their target enzymes is not completely understood; however, this interaction blocks the progression of DNA replication, leading to strand breaks and rapid cell death. • The inhibition of DNA gyrase is bactericidal, whereas inhibition of topoisomerase IV is primarily bacteriostatic.

524

COMMUNITY-ACQUIRED PNEUMONIA

TABLE 4. (continued) Drug Class

Mechanism of Action

Tetracyclines

• Tetracyclines bind to receptors on the 30S subunit of bacterial ribosomes and prevent the attachment of transfer RNA (tRNA) to the ribosomal complex. • Binding to the 30S subunit effectively prevents the addition of new amino acids to the growing polypeptide chain, halting the process of protein synthesis. • Tetracyclines are usually bacteriostatic, but in some organisms, their effect at high doses can be bactericidal.

Aminoglycosides

• Aminoglycosides impair protein synthesis by binding to receptors on the 30S subunit of the bacterial ribosome. • Binding to the 30S subunit interferes with the initiation of polypeptide formation, causes the production of defective proteins (misreading of the messenger RNA template), and disrupts ribosome function. • The effect of aminoglycosides is bactericidal. • Precise mechanisms by which aminoglycosides exert a bactericidal effect are not entirely understood.

Glycopeptides

• Glycopeptides inhibit bacterial cell-wall biosynthesis by blocking glycopeptide polymerization. They also alter bacterial cell-membrane permeability and RNA synthesis. • Glycopeptides produce immediate inhibition of cell-wall synthesis and secondary damage to the cytoplasmic membrane. • Magnesium, manganese, calcium, and ferrous ions reduce the degree of adsorption of vancomycin to the cell wall, but the in vivo importance of this interaction is unknown. • Bactericidal action results primarily from inhibition of cell-wall synthesis.

Oxazolidinones

• Oxazolidinones bind to a site on the bacterial 23S ribosomal RNA of the 50S subunit, preventing the formation of a functional 70S initiation complex, which is an essential component of the bacterial translation process. • The result is inhibition of the initiation of protein translation. No direct action on DNA or RNA synthesis has been observed.

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TABLE 5. Prevalence of Resistance to Commonly Prescribed Antibiotics in S. pneumoniae and H. influenzae Isolates in the Major Pharmaceutical Markets, 2004 Prevalence of Resistance (%) United States Streptococcus pneumoniae Penicillin Intermediatea High-levelb Amoxicillin/clavulanate Intermediate High-level Cefuroxime Ceftriaxone TMP/SMX Tetracyclines Macrolides Levofloxacin Haemophilus influenzae β-lactamase production Amoxicillin/clavulanate Cefaclor Cefuroxime Ceftriaxone TMP/SMX Tetracyclines Azithromycin Clarithromycin Levofloxacin

France Germany

18 16

33 33

8 14 27 4 29 > 20 23 0.5 31–42 0.2–4.5 13 1 0 9–16 1 0.5 0.3–4 0

7 1

Italy

United Spain Kingdom Japan

12 5

41 37

6 5

44 10

20 23 55 5 14 > 20 47–58 0

1.5 1.5 0 3 1 5 0 0.5 4 4–30 N.A. 30 9 24–42 0.4 0

22 26 47 8 22 > 20 37 0

3 1.5 9 0 2 NR 9 0

10 2 37 2 2 > 20 66–78 1.0

20–28 0 N.A. 0 0 7–17 N.A. 0 5 0

6 2–8 28–32 0 0 0.5 N.A. N.A. N.A. 0 0 0 0 0 0 19 11–22 35–52 N.A. N.A. N.A. 0 0 0.3 5 3 9 0 0 0

14–18 0 N.A. 0 0 6–21 N.A. 0 5 0

15 0 N.A. 0 0 18 N.A. 0 7 0

a High-level resistance (minimum inhibitory concentration [MIC] = 2 mg/L). b Intermediate susceptibility (MIC = 0.12–1.0 mg/L).

N.A. = Not available. TMP/SMX = Trimethoprim/sulfamethoxazole (co-trimoxazole). Note: Prevalence numbers have been rounded.

MICs) that also confers reduced susceptibility to other agents in the β-lactam class of antibiotics. Macrolide resistance levels range between 23% and 30% in the United States, and much higher levels are observed in some other markets (Brown SD, 2004; Farrell DJ, 2004; Doern GV, 2004). Table 5 shows the prevalence of resistance to various antibiotics in S. pneumoniae and Haemophilus inßuenzae. Pathogen resistance to an antibiotic can reduce the clinical efÞcacy of the agent and potentially endanger the life of the patient. It is more likely to occur if a patient has had recent multiple courses of antibiotic therapy (e.g., patients who have had recurrent CAP). Historically, antibiotic resistance has had an important impact on the antibacterial market, both by spurring the uptake of novel classes of agents and by causing public health groups to promote less use of antibiotics. However, the clinical impact of drug resistance is controversial, and conßicting studies have shown different relationships between infection with a

526

COMMUNITY-ACQUIRED PNEUMONIA

resistant pathogen and clinical outcome (Rothermel CD, 2004; Song JH, 2004). In speciÞc segments of the population—such as high-risk patients with severe CAP and/or multiple comorbidities—concern about antibacterial resistance plays a much bigger role in drug selection. Rising levels of bacterial resistance have inßuenced treatment guidelines and resulted in broad public health attempts to limit antibiotic usage. Over time, antibacterial resistance to a drug can increase in the community setting if an agent is overprescribed. The prevalence of resistance has been related to the volume of prescribing, and overprescription has been proposed as one reason for the geographic variation in antibacterial resistance. To prevent further increases in resistance, public health groups and thought leaders have promoted use of antibiotics with low-resistance potential or narrowspectrum therapy to treat respiratory infections (Cunha BA, 2002). Clinical guidelines for respiratory diseases also commonly advise physicians to tailor empiric therapy to local susceptibility patterns. Penicillins Overview. The penicillins comprise several subgroups of agents with a wide range of bacterial coverage and efÞcacy. Each penicillin molecule contains a basic β-lactam structure fused to a Þve-membered ring. Because of their broad spectrum of activity and availability in oral form, the penicillins are commonly used in the treatment of CAP and have become the drugs of choice in treating many common infections. The penicillins are divided into the following groups: natural penicillins, aminopenicillins, and the extended-spectrum penicillins. The natural penicillins (e.g., penicillin G) have the narrowest spectrum. They are active only against the gram-positive cocci; in CAP, they are active essentially against S. pneumoniae only. The aminopenicillins (such as ampicillin and amoxicillin) are natural penicillin derivatives that expand the spectrum of activity to include some gram-negative organisms, including H. inßuenzae strains that do not produce β-lactamases (enzymes that destroy the ring structure of the antibiotic, thereby rendering it ineffective), but not Klebsiella pneumoniae or Pseudomonas aeruginosa. The aminopenicillins have been used since the 1970s, and they continue to be valuable therapeutic agents for the treatment of many bacterial infections. These drugs are sometimes used as Þrst-line agents in the treatment of mildto-moderate CAP if no risk factors are apparent. However, the usefulness of aminopenicillins is increasingly limited in countries such as France and Spain because of resistance among the common respiratory pathogens. The extended-spectrum penicillins (such as piperacillin and ticarcillin) are also semisynthetic penicillins and have a broad range of activity, including many gram-negative organisms such as Escherichia coli and K. pneumoniae. These drugs are more active than natural penicillins and aminopenicillins because they are more resistant to inactivation by β-lactamase and/or because they more readily penetrate the outer membranes of gram-negative bacilli. Nonetheless, their rate of bactericidal action and the completeness of this effect can be inconsistent.

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As a class, the penicillins are among the safest antibiotics. However, one potentially life-threatening adverse effect associated with their use is immediate or delayed hypersensitivity reactions. These reactions can present in a variety of forms, ranging from mild skin rash to anaphylactic shock. The overall incidence of hypersensitivity reactions ranges between 0.7% and 10% of patients treated with penicillins (Weiss ME, 1988). Other more common, yet minor, side effects caused by penicillins are GI disturbances (e.g., nausea, vomiting, diarrhea). A growing concern over the past two decades has been the development of bacterial resistance to the penicillins—particularly penicillin-resistant S. pneumoniae (PRSP). Penicillin drug resistance is caused by production of β-lactamase enzymes and/or changes in penicillin-binding proteins (PBPs) in the cell wall of resistant organisms that result in decreased afÞnity of the antibacterials to their targets. Because of resistance to penicillins (as well as their dosing schedules and frequency of GI side effects), physicians have been switching to newergeneration drugs, such as the ßuoroquinolones, that enable shorter courses of therapy and once-daily dosing and have better activity against resistant pathogens. In some cases, however, physicians are using higher doses of penicillins and/or penicillin/penicillinase inhibitors to overcome resistance. Ticarcillin/clavulanate (GlaxoSmithKline’s Timentin) is an injectable, extended-spectrum penicillin/β-lactamase inhibitor used in severe, hospitalized CAP. This agent and piperacillin/tazobactam (Wyeth/Taisho’s Zosyn) are used in similar settings; ticarcillin/clavulanate will not be discussed in greater detail. Likewise, ampicillin/sulbactam (PÞzer’s Unasyn), an injectable aminopenicillin/βlactamase inhibitor, will not be discussed further; this agent’s activity against gram-positive and some gram-negative organisms is similar to that of amoxicillin/clavulanate. Mechanism of Action. Table 4 lists the mechanisms of action of various antibacterials. Penicillins and related β-lactam antibacterials bind to PBPs in bacteria and prevent bacterial cell-wall formation. The PBPs are enzymes that reside on the inner wall of the bacterial cell membrane and help to maintain the cell wall homeostasis, including structural integrity, cell shape, cell division, capsule synthesis, phage resistance, and regulation of autolysis. The β-lactam ring of penicillins interacts with PBPs and inhibits cell-wall synthesis, thereby killing the cell. Organisms that are metabolically inactive or lack bacterial cell walls (e.g., Mycoplasma) are not susceptible to β-lactam antibacterials. Amoxicillin. Amoxicillin (GlaxoSmithKline’s Amoxil, generics) (Figure 2) is av ailable in capsule, tablet, and oral suspension formulations. This H COOH O H HO

C NH2

CH3

N CONH

S H

CH3

H

FIGURE 2. Structure of amoxicillin

3 H2O

528

COMMUNITY-ACQUIRED PNEUMONIA

aminopenicillin is active against gram-positive organisms and some gramnegative organisms, such as H. inßuenzae strains that do not produce βlactamases, but not against K. pneumoniae or P. aeruginosa. Amoxicillin has been commercially available since 1974 and continues to be a valuable therapeutic agent for the treatment of many bacterial infections. It is used as a Þrst-line agent in the treatment of mild-to-moderate CAP if self-resolution is likely and no other risk factors are apparent. As a β-lactam antibiotic, amoxicillin binds to PBPs in bacteria and prevents bacterial cell-wall formation. A randomized, double-blind clinical trial compared the efÞcacy of amoxicillin with that of telithromycin (Hagberg L, 2002). Four hundred and four adults were randomized to treatment with 1,000 mg amoxicillin three times daily or to 800 mg of telithromycin once daily for ten days. At 17- to 24-day follow-up, the clinical cure rate for telithromycin was 94.6%, comparable to amoxicillin’s 90.1% cure rate. Side effects associated with amoxicillin are generally minimal, with some incidence of diarrhea, instances of skin rashes, and, particularly, delayed responses. Hypersensitivity to penicillins is an important adverse event and occurs in about 10% of the population (Solensky R, 2003). Amoxicillin is typically well absorbed from the GI tract, has a rapid onset and long duration of action, requires less frequent dosing, and has a favorable side-effect proÞle. As a generically available product, amoxicillin is also relatively inexpensive. At one time, amoxicillin was highly effective against the S. pneumoniae. Unfortunately, bacterial resistance to amoxicillin has increased signiÞcantly in S. pneumoniae and in H. inßuenzae. Consequently, physicians have turned to newer generations of drugs that have a lower resistance potential and are active against these pathogens. Amoxicillin/Clavulanate. Amoxicillin/clavulanate (GlaxoSmithKline’s Augmentin, Augmentin ES, Augmentin XR, generics) is a combination of amoxicillin and the β-lactamase inhibitor clavulanate. The product is available in immediate-release tablets, extra-strength and extended-release tablets, powder for oral suspension, chewable tablets, and parenteral form (in Europe only). GlaxoSmithKline (GSK) is attempting to retain sales of its amoxicillin/clavulanate franchise with the branded Augmentin XR and ES formulations; generics competition began to erode sales in the United States in 2002. The drug has a broad spectrum of bactericidal activity against many gram-positive and gram-negative microorganisms and has been a favorite of clinicians thanks to its efÞcacy, safety, and familiarity. The formulation of amoxicillin and clavulanic acid in the combination drug protects amoxicillin from degradation by β-lactamase enzymes that bacteria might release and effectively extends the antibiotic spectrum of amoxicillin to include many bacteria normally resistant to amoxicillin and other β-lactam antibiotics. The antibiotic component functions by disrupting bacterial cell-wall synthesis. At least three randomized, controlled, double-blind, comparative clinical studies and one noncomparative study were conducted in adults with CAP (GlaxoSmithKline, 2004 [a,b]). In the comparative studies, 582 patients

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received amoxicillin/clavulanate at a dose of 2,000 mg/125 mg orally every 12 hours for 7 or 10 days; the combined clinical success rate ranged from 86.3% to 94.7% in patients who received amoxicillin/clavulanate. In the noncomparative study to assess both clinical and bacteriologic efÞcacy, 1,122 patients received amoxicillin/clavulanate 2,000 mg/125 mg orally every 12 hours for 7 days; the clinical success rate was 85.6%. Amoxicillin/clavulanate is generally well tolerated. The most frequently reported adverse effects were diarrhea/loose stools (9%), nausea (3%), skin rashes and urticaria (3%), vomiting (1%), and vaginitis (1%). The overall incidence of side effects, particularly diarrhea, increased with the higher recommended dose. In clinical practice, diarrhea and loose stools are the most frequently reported side effects, but these tolerability problems can be minimized by taking the product with food and rarely result in discontinuation of treatment. Other, less frequently reported reactions include abdominal discomfort, ßatulence, and headache. The low propensity of amoxicillin/clavulanate to select resistance mutations and a favorable pharmokinetic proÞle predictive of high bacteriologic efÞcacy may account for the longevity of this combination in clinical use. However, in certain deÞned geographical areas, the emergence of S. pneumoniae strains with elevated MICs (i.e., lower susceptibility to the agent) has been observed. To treat drug-resistant S. pneumoniae, high-dose amoxicillin/clavulanate formulations have been developed that are active against some resistant bacterial strains. An enhanced tablet dosage form of amoxicillin/clavulanate (2,000/125 mg twice daily [GSK’s Augmentin XR]) has been developed for use in adult respiratory tract infection. This enhanced formulation prolongs the time that bacteria are exposed to the antibiotic and promotes coverage of tough-to-treat S. pneumoniae with reduced susceptibility to amoxicillin, β-lactamase-producing H. inßuenzae, and Moraxella catarrhalis. Piperacillin/Tazobactam. Piperacillin/tazobactam (Wyeth/Taisho’s Zosyn Tazocin) combines the extended-spectrum penicillin piperacillin (Figure 3) with the βlactamase inhibitor tazobactam (Figure 4). Piperacillin/tazobactam, commercially available since 1993, is available for parenteral administration only. U.S. patent expiry of piperacillin/tazobactam is expected in 2007. In this β-lactam/β-lactamase inhibitor combination therapy, the latter protects the antibiotic from degradation by β-lactamase enzymes released by bacteria, thereby extending the effective spectrum of the therapy to treat bacteria resistant to the β-lactam antibiotic treatment alone. Like other β-lactam therapies, piperacillin/tazobactam achieves its bactericidal effect by interacting with PBPs and disrupting bacterial cell-wall formation. Piperacillin’s broad range of activity (which includes many gram-negative organisms such as P. aeruginosa and K. pneumoniae) is enhanced by the β-lactamase inhibitor tazobactam, which effectively extends the spectrum of activity to include H. inßuenzae. Piperacillin/tazobactam is used for severe and/or complicated CAP episodes in hospitalized patients in whom broad (gram-negative) coverage is required.

530

COMMUNITY-ACQUIRED PNEUMONIA

H COONa O H

CH3

N

C

CONH

NHCO

S H

N

O

N

O

CH3

H

C2H5

FIGURE 3. Structure of piperacillin

FIGURE 4. Structure of tazobactam (R = R1 = H)

An open-label, randomized, multicenter study compared piperacillin/ tazobactam with amoxicillin/clavulanate for the treatment of severe pneumonia in 84 patients. Investigators reported a 90% favorable clinical response rate with piperacillin/tazobactam and 84% with amoxicillin/clavulanate (Speich R, 1998). In a second randomized, open-label, controlled study, piperacillin/tazobactam was signiÞcantly more effective than ticarcillin/clavulanic acid in terms of clinical and microbiological outcome in patients with CAP. For at least 5 days, 177 patients received piperacillin/tazobactam (3 g/375 mg), and 122 received ticarcillin/clavulanate(3 g/100 mg), every 6 hours by IV infusion. Favorable bacteriologic response rates at follow-up were 91% and 67%, respectively (Shlaes DM, 1994). In clinical trials of piperacillin/tazobactam, 90% of the adverse reactions were transient and mild to moderate in severity. The most signiÞcant side effects reported include skin rashes/pruritus, GI complications (including diarrhea, nausea, and vomiting), and allergic reactions in 1.3%, 0.9%, and 0.5% of treated patients, respectively. Cephalosporins Overview. The cephalosporins contain a basic β-lactam structure fused to a six-membered ring. Drugs in this class differ widely in their spectrum of activity, susceptibility to β-lactamases produced by bacteria, and serum half-life. Cephalosporins are categorized into four generations, with each newer generation representing an improvement in the spectrum of bacterial coverage. The most important agents in this class are the second- and third-generation agents, which are commonly used in hospitalized patients in combination with a macrolide. One

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of the limitations of the cephalosporins is lack of activity against the atypical pathogens, such as Chlamydia pneumoniae and Mycoplasma pneumoniae. First-generation cephalosporins have the narrowest spectrum of activity and are most active against staphylococci and streptococci. Most Þrst-generation cephalosporins are oral formulations. For CAP, use of Þrst-generation cephalosporins is limited to treatment of mild episodes, usually in patients with a history of delayed-reaction penicillin hypersensitivity (increased sensitivity to penicillin that does not appear until 24–48 hours after exposure), or cases where the identity of the infecting pathogen is known. Second-generation agents have increased activity to cover more gram-negative bacilli, but they are usually less active than the Þrst-generation drugs against gram-positive bacteria. The oral second-generation cephalosporins are used empirically to treat mild episodes of CAP. Comparable in activity to the oral secondgeneration cephalosporins, the parenteral second-generation cephalosporins are typically prescribed for patients who are unresponsive to Þrst-line therapies or unable to take oral formulations. Third-generation cephalosporins are active against gram-negative organisms, but their activity against gram-positive organisms is inferior to that of previous generations. These agents are the cephalosporins most commonly recommended in clinical guidelines for Þrst-line treatment of patients hospitalized with CAP. Compared with second-generation cephalosporins, the third-generation agents are more potent against gram-negative bacteria, have greater stability against β-lactamases, and have longer serum half-lives. As a result, they have more convenient dosing regimens. The third-generation agents may also be used in combination with extended-spectrum penicillins or aminoglycosides to treat severe CAP. Fourth-generation agents have enhanced stability against β-lactamases and provide good coverage of both gram-positive and gram-negative bacteria. Fourthgeneration cephalosporins have pharmacokinetic properties similar to those of the third-generation cephalosporins. They are reserved for severe infections such as sepsis given their efÞcacy against problem pathogens such as Pseudomonas species. As a class, the cephalosporins are generally well tolerated (Gustaferro CA, 1991; Lepper H, 1995; Okamoto MP, 1994). Common adverse effects are usually minor; GI disturbances and thrombophlebitis (with IV agents) are the most prominent. Disturbances of the GI tract are reported less often with cephalosporins than with the penicillins. Cephalosporins have declined in use for outpatient CAP as a result of their poor activity against atypical organisms and the rise of the macrolides and ßuoroquinolones. However, second- and third-generation cephalosporins have an important role in treatment of hospitalized patients. Resistance against cephalosporins, as with other β-lactam antibiotics, results from pathogen changes in outer-membrane permeability, stability against βlactamases, and modiÞcation of PBPs. While β-lactamase production by H. inßuenzae or M. catarrhalis limits the use of certain penicillins such as amoxicillin, many cephalosporins are effective in treating infections caused by these

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COMMUNITY-ACQUIRED PNEUMONIA

β-lactamase-producing bacteria. Resistance to third-generation cephalosporins in gram-negative pathogens is a formidable problem in the hospital setting and is associated with adverse clinical outcomes and increased hospital costs (Cosgrove SE, 2002). Mechanism of Action. Cephalosporins, like the penicillins, bind to PBPs in bacteria and prevent bacterial cell-wall formation. By interrupting cell-wall formation, cephalosporins induce cell lysis and death. The cephalosporins contain a β-lactam ring that interacts with the PBPs. Cefuroxime Axetil. Cefuroxime axetil (GSK’s Ceftin/Zinnat/Oracef) *is a second-generation cephalosporin consisting of an esteriÞed pro-drug of cefuroxime created for oral formulation; the active antibiotic metabolite cefuroxime is released after cefuroxime axetil absorption from the GI tract. First launched in the United States in 1988, cefuroxime axetil was available as a generic in the United States by 2002. Cefuroxime is active against a broad spectrum of gram-positive and gram-negative bacteria, including Staphylococcus aureus, Streptococcus pneumoniae, Streptococcus pyogenes, E. coli, H. inßuenzae, K. pneumoniae, and M. catarrhalis. Cefuroxime axetil is indicated for a number of bacterial infections, including RTIs caused by β-lactamase-negative strains of S. pneumoniae, H. inßuenzae, M. catarrhalis, and Haemophilus parainßuenzae. A multicenter trial of 185 patients hospitalized with CAP compared the efÞcacy of two dosing regimens of cefuroxime axetil (250 mg twice daily and 500 mg twice daily) with cefaclor (500 mg three times daily). The 250 mg dose group had a 90% favorable clinical response (58% cured, 32% clinically improved); the 500 mg cefuroxime axetil group had a 98% favorable clinical response (94% cured, 4% clinically improved). The third group of patients receiving cefaclor had a 97% favorable clinical response (88% cured, 9% clinically improved) (Yangco BG, 1990). In clinical trials for cefuroxime axetil, patients who received seven to ten days of therapy experienced adverse reactions similar in type and frequency to adverse reactions associated with other antibiotic agents (GlaxoSmithKline, 2003). Diarrhea (3.7%) and nausea/vomiting (3.0%) were among the most common reactions in trials with adults receiving tablets, while pediatric patients receiving the oral formulation experienced diarrhea (8.6%), dislike of taste (5.0%), diaper rash (3.6%), and nausea/vomiting (2.6%). Cefprozil. Cefprozil (Bristol-Myers Squibb’s [BMS’s] Cefzil/Procef/Bisoral) is a second-generation oral cephalosporin. The agent has been available in the United States since 1992 and has been marketed in a few European countries (Italy, Spain, and the United Kingdom). The agent is expected to have patent protection until 2006 in the United States and between 2004 and 2008 in the European markets. Cefprozil is active against gram-positive and gramnegative bacteria, but not Pseudomonas species. In clinical studies, cefprozil proved active against S. aureus, S. pneumoniae, S. pyogenes, H. inßuenzae, and M. catarrhalis. Although cefprozil is not speciÞcally indicated for CAP (the agent is approved for other RTIs, including acute exacerbations of chronic bronchitis [AECB],

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pharyngitis, otitis media, and acute sinusitis), one comparative clinical trial used cefprozil and cefaclor in the treatment of pneumonia. Of 90 patients who received 500 mg of cefprozil twice per day, 74 (82%) showed a clinical response; of 67 patients who received 500 mg of cefaclor every eight hours, 53 (79%) demonstrated a clinical response (Ball P, 1994). In clinical trials for cefprozil, side effects included nausea, diarrhea, vomiting, and abdominal pain. Cefpodoxime Proxetil. Cefpodoxime proxetil (PÞzer’s Vantin, SanoÞ-Aventis’ Orelox, Sankyo’s Banan) is an extended-spectrum, third-generation, oral cephalosporin. Sankyo originally developed cefpodoxime proxetil and Þrst launched the cephalosporin in Japan in 1989. It was marketed in the United States by Upjohn (later Pharmacia) in 1992; Aventis marketed the agent in Europe. A generic version of cefpodoxime developed by Ranbaxy has been approved in the United States. Cefpodoxime is indicated for the treatment of CAP caused by S. pneumoniae and H. inßuenzae (including β-lactamase-producing strains). Cefpodoxime proxetil is an esteriÞed pro-drug of cefpodoxime created for oral formulation. It is stable in the presence of most β-lactamase enzymes, effectively extending its spectrum of activity against several gram-positive and gram-negative bacteria that are resistant to penicillins and other cephalosporins. However, some extended-spectrum β-lactamase enzymes can inactivate cephalosporins. Clinical studies have shown cefpodoxime to be active against S. pneumoniae, S. aureus, H. inßuenzae, E. coli, K. pneumoniae, and M. catarrhalis and inactive against enterococci and Pseudomonas species. In a comparative trial in more than 200 patients with CAP, 200 mg of cefpodoxime proxetil, administered twice daily for 5 to 10 days, was compared with 500 mg of amoxicillin administered three times daily. The study established that the clinical and bacteriologic efÞcacy of the two antibiotics were comparable (Geddes AM, 1991). In clinical trials of cefpodoxime, 7.0% of treated patients experienced diarrhea, 3.3% reported nausea, 1.0% reported vaginal fungal infections, and 1.2% experienced abdominal pain (Pharmacia & Upjohn, 2003). The frequency of side effects is similar to frequency associated with other oral cephalosporins. Cefotaxime. Cefotaxime (Abbott/SanoÞ-Aventis’s Claforan, generics) (Figure 5) is a third-generation, parenteral cephalosporin available for intravenous (IV) or intramuscular (IM) administration. Cefotaxime was Þrst marketed in 1981 in the United States, where generic versions of the injection are now available. The cephalosporin is also marketed by Roche in Japan. COONa OCH3 O N N

C

CH2OCOCH3

N CONH S H

H2N

H

S

FIGURE 5. Structure of cefotaxime.

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COMMUNITY-ACQUIRED PNEUMONIA

Cefotaxime is stable in the presence of many β-lactamase enzymes, effectively extending its spectrum of activity against a number of gram-positive and gram-negative bacteria resistant to penicillins and other cephalosporins. Clinical and laboratory studies have demonstrated the activity of cefotaxime against S. aureus, S. pneumoniae, and other streptococci. Gram-negative species susceptible to cefotaxime include E. coli, H. inßuenzae, H. parainßuenzae, K. pneumoniae, and others. It is also active against P. aeruginosa. Cefotaxime is indicated for treatment of lower respiratory tract infections caused by S. pneumoniae, S. pyogenes, and other streptococci (but not enterococci), S. aureus, E. coli, K. pneumoniae and other Klebsiella species, H. inßuenzae (including ampicillin-resistant strains), H. parainßuenzae, and Pseudomonas species (including P. aeruginosa). A clinical trial examined the efÞcacy of cefotaxime in treating patients with severe respiratory infections, including pneumonia. One hundred and ninety patients were treated with 2.25–3.0 g of cefotaxime daily for an average of 9 days and displayed a favorable clinical response in 91% of 184 evaluable cases (Pines A, 1980). Cefotaxime is generally well tolerated, but some adverse reactions have been documented, including local inßammation at point of IV administration (4.3%) and GI disturbances (1.4%). Systemic adverse reactions to cefotaxime have been rare. The frequency of side effects is similar to frequency associated with other parenteral cephalosporins. Ceftriaxone. Ceftriaxone (Roche’s Rocephin, generics) (Figure 6) is one of the most successful antibiotics on the market. The agent is a third-generation parenteral cephalosporin available for IV or IM administration that was Þrst launched in 1982. Ceftriaxone has been available generically in Europe since 2002. In 2000, Cubist Pharmaceuticals announced that it had acquired the rights to oral ceftriaxone and was developing an alternate oral formulation. However, the company discontinued development in 2004 after human clinical trials showed highly variable bioavailability of the drug when administered orally. Ceftriaxone has demonstrated a broad spectrum of activity against grampositive and gram-negative bacteria, including S. aureus, S. pneumoniae, S. pyogenes, and other streptococci, E. coli, H. inßuenzae, H. parainßuenzae, K. pneumoniae, M. catarrhalis, and others. Ceftriaxone is indicated for lower respiratory tract infections caused by S. aureus, S. pneumoniae, H. inßuenzae, H. parainßuenzae, K. pneumoniae, and

FIGURE 6. Structure of ceftriaxone

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E. coli. The agent is one of the most widely used parenteral antibiotics for respiratory tract infections. The efÞcacy of ceftriaxone in the treatment of lower respiratory tract infections was evaluated in a comparative clinical study with cefonicid; 118 patients received 1 g of either ceftriaxone or cefonicid, IV or IM, daily for 3 to 11 days (mean was seven days). Clinical cure or improvement was noted in 93% of patients treated with ceftriaxone and 95% of patients treated with cefonicid. Bacteriologic cure or improvement was noted in 81% and 69% of cases, respectively (McCabe RE, 1989). The agent is well tolerated in extensive clinical use. Local reactions have been reported following parenteral administration of antibiotics, including ceftriaxone. Pain, induration, and tenderness were reported in about 1% of cases. The frequency of side effects is comparable to frequency associated with other parenteral cephalosporins. Cefdinir. Cefdinir (Abbott’s Omnicef, Fujisawa’s Cefzon) is a third-generation, oral cephalosporin available as a capsule and as an oral suspension. Cefdinir was developed and launched by Fujisawa in Japan in 1991 as Cefzon. WarnerLambert’s Parke-Davis (now PÞzer) was the Þrst to license cefdinir for the United States and Western Europe and began marketing cefdinir in the United States in 1998, but Abbott obtained the exclusive marketing rights from Fujisawa and began marketing the drug as Omnicef in 2000. The agent is expected to retain patent protection through 2007 in the United States. Cefdinir is indicated for treatment of adults and adolescents who have CAP caused by penicillinasesusceptible S. pneumoniae and all strains of H. inßuenzae, H. parainßuenzae, and M. catarrhalis. Cefdinir is stable in the presence of many β-lactamase enzymes, effectively extending its spectrum of activity against a number of gram-positive and gramnegative bacteria resistant to penicillins and other cephalosporins. Cefdinir has demonstrated clinical and bacteriologic activity against S. aureus, Staphylococcus epidermitis (in vitro), S. pneumoniae, S. pyogenes, E. coli (in vitro), H. inßuenzae, H. parainßuenzae, K. pneumoniae (in vitro), M. catarrhalis, and others. Cefdinir is not active against Pseudomonas species. A double-blind, controlled, Phase III clinical study enrolled a total of 690 patients in the United States to compare the efÞcacy of cefdinir and cefaclor for treatment of CAP in adults and adolescents. One group of 347 patients received 300 mg of cefdinir twice daily for ten days; another group of 343 patients received 500 mg of cefaclor three times daily for ten days. Clinical cure or improvement rates for cefdinir and cefaclor were similar: 89% (166/187) and 86% (160/186), respectively; bacteriologic eradication rates were 92% (238/260) and 93% (245/264), respectively (Drehobl M, 1997). A Phase III comparative clinical trial showed that cefdinir had a statistically signiÞcant higher rate of adverse reactions (25.9%) than cephalexin (16.1%) (Tack KJ, 1998). In particular, cefdinir causes more GI discomfort (such as diarrhea) than other cephalosporins. The most frequently reported adverse reactions in U.S. clinical trials with a total of 3,841 cefdinir-treated adult patients were diarrhea (15%), vaginal moniliasis (4% of women), nausea (3%), and headache (2%). Most

536

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adverse events were mild and self-limiting in nature, with a low discontinuation rate (3%) due to associated adverse events. The most frequently reported adverse reactions in U.S. clinical trials with a total of 1,783 cefdinir-treated pediatric patients were diarrhea (8%), rash (3%), and vomiting (1%). Carbapenems Overview. Carbapenems are penicillin derivatives that have good activity against gram-positive and gram-negative aerobic and anaerobic bacteria. They are highly resistant to β-lactamase and have a very favorable spectrum of activity. A drawback of these agents is that they are available in IV form only. These agents are used in the treatment of severe CAP when sepsis is a concern. The incidence of this adverse reaction is low (1%) and occurs predominantly in the elderly or in patients with predisposing CNS conditions (Norrby SR, 2000). Carbapenem antibiotics include imipenem (Merck’s Primaxin), meropenem (AstraZeneca’s Merrem), and ertapenem (Merck’s Invanz). This section reviews only imipenem/cilastatin because this class is generally used only in a small subset of CAP patients. Mechanism of Action. Carbapenems, like the penicillins, bind to PBPs in bacteria and prevent bacterial cell-wall formation. By interrupting cell-wall formation, carbapenems induce cell lysis and death. Imipenem/Cilastatin. Merck’s imipenem/cilastatin (Primaxin) is a prototypical carbapenem. Imipenem is coadministered with cilastatin (a peptidase inhibitor) (Figure 7) to prevent its metabolism into nephrotoxic products by peptidase enzymes in the body. Coadministration allows extended imipenem dosage intervals and causes less toxicity (Norrby SR, 2000). Imipenem/cilastatin is usually reserved for severe CAP in the intensive care setting for patients who develop sepsis. In this setting, the agent is often administered with an aminoglycoside. This agent is expected to retain patent protection through 2009 in the United States. A randomized trial compared imipenem/cilastatin with the third-generation cephalosporin ceftazidime (both IV) in 60 hospitalized patients with moderate to severe CAP (Ho A, 1997). The response rate in both arms was 70%, recorded as improvement in sputum, purulence, cough, and dyspnea scores. Macrolides Overview. Macrolides are a class of drugs that inhibit bacterial protein synthesis. They demonstrate excellent activity against atypical organisms (Mycoplasma, Chlamydia, and Legionella species), but have variable activity against typical

FIGURE 7. Structure of cilastatin

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pathogens (S. pneumoniae and H. inßuenzae). Macrolides are indicated for use in mild-to-moderate CAP and are typically used as Þrst- and second-line agents for this indication. Likewise, they are used in combination with penicillins or cephalosporins for severe CAP to provide atypical pathogen coverage. Although resistance levels to the macrolides may differ among the various agents in this class, cross-resistance is assumed for gram-positive organisms. The macrolides are available in oral and parenteral formulations. Compared with erythromycin, the prototypical macrolide, the advanced-generation macrolides have much longer serum half-lives, excellent distribution into respiratory tissues, and improved tolerability. Roxithromycin (SanoÞ-Aventis’s Rulid, generics) is a macrolide launched extensively in Europe; its use has been in general decline and it will not be discussed further. Side effects, particularly of the early macrolides, are associated primarily with the GI tract (e.g., abdominal cramps, nausea, vomiting). Hypersensitivity reactions are rare. In comparison, the advanced-generation macrolides have a lower incidence of side effects. The prevalence of macrolide resistance in S. pneumoniae has increased dramatically over the past two decades. Resistance is highest in the Asia/PaciÞc region: 78% resistance in Japan, 72% in Hong Kong (Inoue M, 2004). In Europe, France (47%) and Italy (42%) have a high prevalence of resistance. In the United States, 23–30% of pneumococci are macrolide-resistant (Brown SD, 2004; Doern GV, 2004; Farrell DJ, 2004; Felmingham D, 2002; Jacobs MR, 2003; Jacobs MR, 1999). Mechanism of Action. Macrolides act by binding to the 23S ribosomal RNA (rRNA) in the 50S subunit of the ribosome. Binding to the 23S rRNA inhibits the translocation of RNA during protein synthesis and blocks bacterial protein synthesis. Macrolide drugs are generally bacteriostatic but can be bactericidal in highly susceptible bacteria or at high concentrations. Erythromycin. Erythromycin (Abbott’s Erythrocin, Eli Lilly/Shionogi’s Ilosone, generics) (Figure 8) is the class-standard macrolide and has been available O H

CH3 H

CH3

CH3 OH

H HO

CH3 H

O HO

H

CH3

H

O H3C H

CH3

O H CH2CH3 HO

O N(CH3)2

OH

O O

CH3 OCH3

CH3

FIGURE 8. Structure of erythromycin

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COMMUNITY-ACQUIRED PNEUMONIA

since the 1950s. The agent has been widely utilized for RTIs, but the nextgeneration macrolides, which provide more convenient dosing, better tolerability, and expanded spectrum, have replaced this agent for more severe respiratory infections such as CAP. The agent is available in oral and parenteral formulations. Erythromycin acts by binding to the 50S ribosomal subunit of susceptible microorganisms, thereby inhibiting protein synthesis. This agent is active against a range of gram-positive and atypical organisms and a limited number of gramnegative organisms. However, many strains of H. inßuenzae and S. pneumoniae are resistant to erythromycin. Early studies of erythromycin demonstrated its efÞcacy in RTIs. In a randomized, double-blind, Phase III trial, 591 patients with CAP received either a single daily dose of dirithromycin (500 mg) or erythromycin (250 mg) four times daily. Clinical response rates were similar in both treatment groups: the clinical and bacteriologic response rates for erythromycin-treated patients were 92.1% and 90.3%, respectively, and 94.5% and 93.0% respectively for dirithromycin-treated patients. Notably, the agent is associated with GI discomfort in about 20% of patients, an adverse effect that has, in part, caused physicians to shift to the newer macrolides. Clarithromycin. Clarithromycin (Abbott’s Biaxin, Biaxin XL/Klacid, Taisho’s Clarith) (Figure 9) is an advanced-generation macrolide commonly used for multiple RTIs, including CAP, AECB, otitis media, and sinusitus. The agent is available in tablets, extended-release tablets, and granules. Clarithromycin acts by binding to the 50S ribosomal subunit of susceptible microorganisms, resulting in inhibition of protein synthesis. Its spectrum of activity includes a range of aerobic and anaerobic gram-positive and gram-negative microorganisms, and like other macrolides, clarithromycin is highly active against atypical pathogens such as M. pneumoniae. This drug is more potent against staphylococci and streptococci and is perceived to have modest activity against H. inßuenzae. The efÞcacy of clarithromycin has been established in multiple clinical studies. In one Phase III study, 280 patients with CAP were randomized to receive either clarithromycin (250 mg) twice daily or erythromycin stearate (500 mg) four times

FIGURE 9. Structure of clarithromycin

CURRENT THERAPIES

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FIGURE 10. Structure of azithromycin

daily for 14 days (Anderson G, 1991). There was no statistically signiÞcant difference between the two groups in terms of clinical cure (52% for clarithromycin, 40% for erythromycin); clinical success (clinical cure and improvement; 89% for clarithromycin, 98% for erythromycin); or radiological response (90% for both groups). On an intention-to-treat basis, clarithromycin showed better success: the clinical cure rate after two weeks of treatment was 45% in patients who received clarithromycin compared with 25% in the erythromycin group. Adverse effects, mainly gastrointestinal, caused discontinuation of treatment in 4% patients in the clarithromycin-treated group, compared with 19% of patients treated with erythromycin. Azithromycin. Azithromycin (PÞzer’s Zithromax/Zitromax) (Figure 10) is widely used for treatment of RTIs, including CAP, AECB, otitis media and pharyngitis. The agent was launched in the early 1990s. Azithromycin can also be used to treat skin infections and some sexually transmitted diseases in adults. In children, azithromycin is used to treat ear infections and pharyngitis/tonsillitis. This agent is available in oral and IV formulations. Azithromycin is recognized for its broad efÞcacy, dosing advantages, favorable side-effect proÞle, and goodtasting liquid formulation for children. It has become the leading macrolide in the United States, owing primarily to its safety, long-half life (which allows an abbreviated Þve-day treatment course), and excellent promotion by PÞzer. Azithromycin acts by binding to the 50S ribosomal subunit of susceptible microorganisms and interfering with microbial protein synthesis. It demonstrates activity in vitro against a wide range of bacteria, including gram-positive bacteria such as S. aureus, S. pneumoniae, and other streptococci, and gram-negative bacteria such as H. inßuenzae and H. parainßuenzae. Azithromycin demonstrates cross-resistance with erythromycin-resistant, gram-positive strains and most strains of methicillin-resistant staphylococci. Azithromycin is indicated for CAP due to S. pneumoniae, H. inßuenzae, C. pneumoniae, or M. pneumoniae in patients for whom oral therapy is appropriate.

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COMMUNITY-ACQUIRED PNEUMONIA

Data summarized from eight comparative studies and two noncomparative studies of patients with pneumonia demonstrate that a standard course of azithromycin delivered overall clinical response rates similar to response rates for comparators in patients of varying ages: 88% for both groups. The comparators used were cefaclor (Lilly’s Ceclor, generics); amoxicillin (GSK’s Amoxil, generics); erythromycin (generics); amoxicillin/clavulanate (generics); and josamycin (Bayer’s Josacine), dosed three times a day or twice a day for seven to ten days as appropriate. The summarized data also demonstrate comparable clinical response rates in the subset of patients aged 16 to 60 years (92% for azithromycin and 95% for comparatives agents) and in the subset of patients older than 60 years of age (82% and 78%, respectively). Bacteriologic response rates (i.e., eradication of the susceptible pathogen present at baseline), compared with the other agents used, were as follows: S. pneumoniae (93% for azithromycin, 95% for comparative agents) and H. inßuenzae (100% for azithromycin, 85% for comparative agents) (Hopkins S, 1995). Azithromycin is rapidly and extensively taken up by leukocytes, which deliver the drug to the site of infection, where the drug maintains high sustained concentrations. In addition, azithromycin has a long half-life (68 hours), thus allowing a convenient Þve-day dosing regimen. It offers excellent tolerability, with a low incidence of adverse events. Azithromycin compares favorably with amoxicillin/clavulanate and other competitive agents from a side-effect and tolerability standpoint. The most common side effects are diarrhea/loose stools (4–5%), nausea (3%), and abdominal pain (2–3%); the overall discontinuation rate is 0.7%. Ketolides Ketolides are a new class of macrolide derivatives designed speciÞcally to combat macrolide-resistant respiratory tract pathogens. The ketolides exhibit good activity against gram-positive and some gram-negative organisms and have excellent activity against drug-resistant S. pneumoniae, including macrolide-resistant strains (Poehlsgaard J, 2002). Spontaneous resistance to the available ketolide, telithromycin, is rare (Davies TA, 2000). Ketolides display excellent pharmacokinetics that allow once-daily dose administration and extensive tissue distribution relative to serum. The ketolide telithromycin is well tolerated; its side effects are primarily GI symptoms (diarrhea, nausea, and headache). Mechanism of Action. Like the macrolides, ketolides act by binding to the 23S ribosomal RNA (rRNA) in the 50S subunit of the ribosome. Binding to the 23S rRNA inhibits the translocation of RNA during protein synthesis and blocks bacterial protein synthesis. Ketolides and macrolides have essentially the same binding sites, located on a portion of the rRNA and on ribosomal proteins, but ketolides bind to ribosomes with higher afÞnity than macrolides. This attribute could contribute to ketolide activity against macrolide-resistant strains (Poehlsgaard J, 2002). Telithromycin. Telithromycin (SanoÞ-Aventis/Fujisawa/Sankyo’s Ketek) (Figure 11), an oral ketolide, is the Þrst member of this new class to be approved

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FIGURE 11. Structure of telithromycin

for clinical use. Ketolides are structural variants of macrolides, and their modiÞcation results in target binding without tripping the inducible resistance to macrolide drugs that many groups of pathogens exhibit (Douthwaithe S, 2001). Telithromycin was Þrst approved in Europe in 2001; Japanese approval was attained in 2003. In the United States, telithromycin’s approval was delayed for more than two years. The FDA issued Aventis an approvable letter for telithromycin in 2001 and requested additional data to support the Þling. Aventis conducted the largest-ever comparative antibiotic trial, enrolling more than 24,000 participants to support U.S. approval, which was granted in April 2004. The product was launched in August 2004. The agent is expected to retain patent protection through 2015 in the United States and Japan and 2016 in Europe. Telithromycin is active against a broad range of pathogens, including S. pneumoniae (including multidrug-resistant strains), H. inßuenzae, M. catarrhalis, C. pneumoniae, S. aureus (methicillin- and erythromycin-susceptible isolates only), and M. pneumoniae. Against isolates of S. pneumoniae, telithromycin has been shown to demonstrate concentration-dependent bactericidal activity in vitro. Telithromycin’s advantage over currently available macrolides lies in its enhanced activity against erythromycin-resistant S. pneumoniae.Antibacterial activity against other respiratory pathogens, such as H. inßuenzae and atypical bacteria, is similar to that of available advanced-generation macrolides—except that telithromycin’s activity is slowly bactericidal rather than bacteriostatic against most respiratory pathogens (Felmingham D, 2001; Felmingham D, 2000; Hammerschlag MR, 2001; Leclercq R, 2001). The agent appears to have a lower propensity to induce certain types of resistance (inducible methylase gene expression) (Buxbaum A, 2003; Clark JP, 2003).

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COMMUNITY-ACQUIRED PNEUMONIA

Telithromycin has demonstrated equivalent activity to clarithromycin, trovaßoxacin, and amoxicillin, with clinical success rates ranging from 88.3% to 94.6%. The agent’s safety and efÞcacy for the treatment of CAP have been shown in four randomized, double-blind, controlled Phase III studies. In the Þrst trial, telithromycin (800 mg once daily) was compared with clarithromycin (500 mg twice daily) for ten days; clinical efÞcacy of telithromycin and clarithromycin was 88.3% vs. 88.5%, respectively. In the second trial, telithromycin was compared with trovaßoxacin (200 mg once daily for seven or ten days); efÞcacy was reported at 90% for telithromycin and 94.2% for trovaßoxacin. This study was stopped prematurely after trovaßoxacin was restricted to use in hospitalized patients with severe infection. The third study compared telithromycin with amoxicillin (1,000 mg three times daily for ten days); efÞcacy was reported at 94.6% for telithromycin and 90.1% for amoxicillin. Finally, a fourth trial compared telithromycin (800 mg once daily for seven days) with clarithromycin (500 mg twice daily for ten days); efÞcacy was reported at 88.8% for telithromycin and 91.8% for clarithromycin. Telithromycin’s side effects include nausea and diarrhea, which occurred in Phase III studies at rates comparable to the rates of comparator antibiotics. The product is also associated with visual disturbances—namely, blurred vision and/or difÞculty focusing—in approximately 1.1% of patients. In a health outcomes analysis of two Phase III studies that compared telithromycin with clarithromycin in CAP patients (Chang JR, 2003), telithromycin was associated with a mean per-patient costs savings of $185 to $419. Fluoroquinolones Overview. Fluoroquinolones are broad-spectrum antibacterials that have experienced an upsurge in use in recent years. Because of their broad-spectrum activity, high efÞcacy, favorable dosing, and availability in oral and IV form, these agents are indicated for a range of bacterial infections, including respiratory, GI, and urinary tract infections. Earlier generations of ßuoroquinolones (e.g., oßoxacin) had limited activity against some respiratory pathogens, such as S. pneumoniae. However, recent ßuoroquinolone agents (so-called third-generation agents, or the “respiratory ßuoroquinolones”) are active against a broad spectrum of grampositive and gram-negative bacteria, including atypical organisms. Therefore, they are highly effective in RTIs. An important factor inßuencing use of recently launched ßuoroquinolones is the withdrawal from the market of several agents in this class due to toxicity—for example, trovaßoxacin (PÞzer’s Trovan) and grepaßoxacin (GSK’s Raxar). Furthermore, ßuoroquinolones as a class are associated with alterations in normal cardiac conduction (i.e., prolongation of the QT interval) that can cause cardiac arrhythmias in vulnerable patients. The degree of QT prolongation varies with each agent; sparßoxacin (Mylan’s Zagam) and grepaßoxacin have been associated with the longest QTc prolongation (Ball P, 2000), and both agents have been removed from the U.S. market. Another important limitation of the ßuoroquinolones is that they are generally contraindicated in children and pregnant women because animal studies have

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shown that they are toxic to growing cartilage (Lipsky BA, 1999; Zhanel GG, 2002). They are used in some severe pediatric infections—for example, in cystic Þbrosis patients—but this is generally the exception. Research in this area is ongoing. Until recently, resistance to the ßuoroquinolones by the respiratory pathogens has been extremely rare (Doern GV, 2004; Hoban DJ, 2001[a]; Rybak MJ, 2004). However, increasing ßuoroquinolone resistance in S. pneumoniae isolates has been reported, and clinical failures with levoßoxacin have been noted in Canada (Odland BA, 1999; Chen DK, 1999). Fluoroquinolone-resistant isolates are more common in people older than 65 years, who have the highest level of ßuoroquinolone use. Prior exposure to ßuoroquinolones is one of the major risk factors for acquisition of levoßoxacin-resistant S. pneumoniae (Ho PL, 2001). Emerging resistance to older ßuoroquinolones, such as ciproßoxacin, has been demonstrated worldwide, ranging from 3% to 12% (Blahova J, 2004; Linares J, 1999; Powis J, 2004). In comparison, penicillin-resistant strains are more common in isolates from young children, who have the highest rate of β-lactam use. Fluoroquinolone resistance often involves alteration of their targets, the topoisomerases. Mechanism of Action. Fluoroquinolones interact with two related yet distinct targets within the bacterial cell: DNA gyrase and topoisomerase IV. DNA gyrase and topoisomerase IV are vital enzymes involved in bacterial DNA replication. These agents are potent inhibitors of nucleic acid synthesis. The exact nature of the interaction of quinolones with their target enzymes is not completely understood; however, this interaction blocks the progression of DNA replication, leading to strand breaks and rapid cell death. The inhibition of DNA gyrase is bactericidal, whereas inhibition of topoisomerase IV is primarily bacteriostatic. Levofloxacin. Levoßoxacin (Johnson & Johnson’s Levaquin, SanoÞ-Aventis’ Tavanic, Daiichi’s Cravit) (Figure 12) is a widely successful ßuoroquinolone approved for the treatment of CAP and several other bacterial infections (e.g., bronchitis, urinary tract infection, skin infections). In the United States, the agent is a leading antibiotic for CAP owing to its high bioavailability, broad-spectrum activity, tolerability, and once-daily dosing. Levoßoxacin was Þrst launched in Japan in 1993 and has been widely available since 1998. Most recently, a new, high-dose formulation (750 mg) has been approved that provides patients an abbreviated Þve-day treatment course for CAP. The agent is expected to lose

FIGURE 12. Structure of levofloxacin, the (S)-enantiomer of ofloxacin

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COMMUNITY-ACQUIRED PNEUMONIA

patent protection in the United States and Japan in 2010 and in Europe in 2011. The drug is available in oral and IV formulations. Levoßoxacin is the levo enantiomer of oßoxacin. The agent’s mechanism of action involves inhibition of bacterial topoisomerase IV and DNA gyrase, which are enzymes required for DNA replication, transcription, repair, and recombination. Levoßoxacin has in vitro activity against a wide range of gram-negative and gram-positive microorganisms. Levoßoxacin’s efÞcacy in adult inpatients and outpatients with CAP was evaluated in two pivotal Phase III clinical studies. In the Þrst study, 590 patients were treated with levoßoxacin (500 mg) once daily orally or intravenously for 7 to 14 days or with the cephalosporin ceftriaxone (1–2 grams), administered intravenously once or twice daily, followed by the cephalosporin cefuroxime axetil (500 mg) administered orally twice daily for a total of 7 to 14 days (OrthoMcNeil, 2004). Patients assigned to treatment with the cephalosporins were allowed to receive erythromycin (or doxycycline if intolerant of erythromycin) if an infection due to atypical pathogens was suspected or proven. Clinical success (deÞned as the percentage of patients cured or improved at 5 to 7 days post-therapy) with levoßoxacin was superior (95%) to success in the control group (83%). In a second, noncomparative study, 264 patients were treated with 500 mg levoßoxacin orally or intravenously once daily for 7 to 14 days; the clinical success rate of evaluable patients in this study was 93%. Moxifloxacin. Moxißoxacin (Bayer Schering-Plough’s Avelox) (Figure 13) is a ßuoroquinolone used for the treatment of RTIs. In December 1999, the FDA approved the oral formulation of moxißoxacin for the treatment of CAP and other respiratory diseases; in December 2001, the agency approved the parenteral formulation of the agent for the same indications. In Germany, oral moxißoxacin received regulatory approval in September 1999 for the treatment of communityacquired infections, including CAP; the parenteral formulation was approved in May 2002. Oral moxißoxacin was recently launched in France, Spain, and Italy. In Japan, moxißoxacin is in Phase III and Phase I trials for the oral and IV formulations, respectively. In September 2003, Schering-Plough and Bayer announced a strategic alliance in which Schering-Plough acquired exclusive rights to market, sell, and distribute moxißoxacin (as well as Bayer’s other antibiotic, ciproßoxacin) in the United States. This agent is expected to retain patent protection through 2014 in the United States and through 2009 in Europe and Japan. O

O

F OH H

N

N

HN

O H

CH3

FIGURE 13. Structure of moxifloxacin

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Moxißoxacin displays excellent activity against a broad spectrum of bacteria, including penicillin- and/or macrolide-resistant S. pneumoniae. It is highly active against the following pathogens: • • • •

Common RTI pathogens such as S. pneumoniae, H. inßuenzae, and M. catarrhalis. RTI pathogens with reduced susceptibility to conventional agents, such as penicillin- and macrolide-resistant strains of S. pneumoniae. Unusual community RTI pathogens such as S. aureus and K. pneumoniae. Atypical microorganisms, such as Mycoplasma, Legionella, and Chlamydia.

Moxißoxacin offers superior activity against S. pneumoniae,compared with current quinolones (e.g., ciproßoxacin, levoßoxacin), and is highly effective in treating other respiratory pathogens. Several multinational clinical trials have compared the efÞcacy of oral moxißoxacin with that of either amoxicillin or clarithromycin in the treatment of CAP (Fogarty C, 1999[a]; Hoeffken G, 2001; Patel T, 2000; Petitprez P, 2001). The studies found that moxißoxacin’s overall clinical success rate (92%) and bacterial eradication rate (94%) were equivalent or superior to clarithromycin and amoxicillin rates (Krasemann C, 2001). Adverse effects associated with moxißoxacin were primarily GI-related (e.g., nausea and diarrhea) and were mild to moderate in severity. Other adverse events reported included rash and dizziness (Patel T, 2000; Hoeffken G, 2001). In postmarketing surveillance of moxißoxacin, less than 5% of patients experienced an adverse event and less than 2% withdrew as a result of a severe adverse event (Landen H, 2001). Additional advantages of moxißoxacin are its high bioavailability and the bioequivalence of the oral and parenteral forms, which require no dose adjustment when stepping down from IV to oral therapy. In a randomized, open-label Phase II study, 628 hospitalized patients with CAP received sequential IV-to-oral therapy with moxißoxacin or amoxicillin/clavulanate, with or without clarithromycin, for 7–14 days. Clinical cure at Þve to seven days post-therapy was 93% for moxißoxacin compared with 85% for the comparator group (Drummond M, 2001). Moxißoxacin’s long half-life allows once-daily dosing (Stass H, 1999), and the drug has limited drug-drug interactions. Moxißoxacin is not associated with phototoxicity, but like other ßuoroquinolones, it has been shown to alter the QT interval. Gatifloxacin. Gatißoxacin (BMS’s Tequin, Kyorin/Dainippon’s Gatißo) (Figure 14) was approved in the United States in December 1999 in both oral and IV formulations. The oral drug was launched in Japan in June 2002, where Kyorin is comarketing the drug with Dainippon Pharmaceutical. Gatißoxacin received regulatory approval in Germany in November 2001 but was reportedly awaiting registration in the European Union, where investigations into the compound’s risk-beneÞt proÞle were being conducted (Reuters Health, press release, April 2002). Gr¨unenthal licensed the product from Kyorin for the European markets. Gatißoxacin is expected to retain patent protection in the United States through 2007 and through 2007–2012 in Japan and Europe.

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COMMUNITY-ACQUIRED PNEUMONIA

O

O

F OH CH3 HN

N

N O CH3

FIGURE 14. Structure of gatifloxacin

Gatißoxacin has a broad spectrum of activity similar to activity observed in other third-generation ßuoroquinolones. The agent is well absorbed following oral administration (almost 100% bioavailability), and its pharmacodynamic and pharmacokinetic properties (e.g., high volume of distribution, long elimination half-life) allow once-daily administration (Nakashima M, 1995). Researchers have conducted several double-blind, randomized, comparative studies of gatißoxacin (400 mg, once daily) in the treatment of CAP. In one Phase III trial, gatißoxacin (oral, 400 mg, once daily) was compared with clarithromycin (oral, 500 mg, twice daily) in 431 patients. Gatißoxacin showed a clinical efÞcacy rate of 95%; clarithromycin’s rate was 93% (Ramirez JA, 1999). In another randomized, open-label, parallel-group, multicenter study, adults with CAP received 7–14 days of treatment with IV gatißoxacin (400 mg, once daily) with the step-down option, or with IV ceftriaxone (1 or 2 g, once daily [with or without erythromycin, 0.5 or 1 g once daily, or clarithromycin, 500 mg, twice daily]) with the step-down option. The study included 170 adults with CAP. Cure rates at 1 to 3 days after treatment were 97.4% in the gatißoxacin group and 90.9% in the ceftriaxone group. In patients whose pathogens were isolated from pretreatment sputum cultures, bacteriologic eradication rates were 100% (gatißoxacin) and 90.9% (ceftriaxone). Treatment-related adverse events occurred in 27.1% of gatißoxacin patients and 21.2% of ceftriaxone patients (Correa JC, 2003). Overall efÞcacy rates in patients in each of the aforementioned trials have demonstrated that gatißoxacin produces cure rates similar to those of the cephalosporin ceftriaxone (with or without erythromycin), the macrolide clarithromycin, and levoßoxacin. In clinical trials, gatißoxacin had a relatively low rate of adverse effects. The most commonly reported adverse effects were nausea (2.8%), headache (2.2%), and dizziness (1.3%) (Casillas JL, 2000). Other side effects, observed at lower frequencies, included gastritis, diarrhea, and rash. Like product labeling for other ßuoroquinolones, gatißoxacin’s labeling warns against giving this agent to patients with long QT syndrome. Postmarketing surveillance has found some reports of serious disturbances of glucose homeostasis in diabetic patients being treated with gatißoxacin. Tetracyclines Overview. Tetracyclines are the prototypical broad-spectrum antibiotics and are occasionally used as Þrst-line agents in some markets (e.g., Germany) for the treatment of mild CAP, particularly when cost, penicillin hypersensitivity, and

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β-lactam resistance are concerns. The widespread use of tetracyclines has resulted in a steady increase in the prevalence of resistance to these agents. Therefore, empiric use of tetracycline is usually restricted to regions where resistance levels remain low or to cases where other appropriate antibiotics are contraindicated. Although some tetracyclines have very low activity against S. pneumoniae, doxycycline maintains good antipneumococcal activity and is the tetracycline most commonly used for treatment of bacterial CAP. Tetracyclines are also sometimes used to treat CAP caused by atypical pathogens. Minocycline (Wyeth’s Minocin, generics) is another widely available tetracycline also used for RTIs, but this agent will not be discussed further owing to its limited use in CAP. All tetracyclines have important adverse reactions with respect to bones and teeth (i.e., bone deposition and permanent teeth staining), and they are contraindicated during pregnancy and for children younger than age eight. Adverse events most commonly reported include stomach upset, diarrhea, nausea, headache, and vomiting. Additionally, phototoxicity, dizziness, and vertigo may also occur. Mechanism of Action. Tetracyclines bind to receptors on the 30S subunit of bacterial ribosomes and prevent the attachment of transfer RNA (tRNA) to the ribosomal complex. Binding to the 30S subunit effectively prevents the addition of new amino acids to the growing polypeptide chain, thus halting the process of protein synthesis. Tetracyclines are usually bacteriostatic, but in some organisms, their effect can be bactericidal at high doses. Doxycycline. Doxycycline (PÞzer’s Vibramycin, generics) (Figure 15) is a broad-spectrum antibiotic derived from oxytetracycline that has been used since the 1960s to treat both gram-negative and gram-positive bacterial infections. It is currently available in oral suspension, capsule, syrup, and IV formulations and has an improved pharmacokinetic proÞle that includes higher oral bioavailability, greater distribution volume, and longer serum half-life, compared with earlier tetracyclines. Doxycycline inhibits bacterial protein synthesis by binding to the 30S subunit of the bacterial ribosome and subsequently preventing the addition of new amino acids to the growing polypeptide chain. In a double-blind, multicenter Phase III study, doxycycline (200 mg once daily) was compared with roxithromycin (150 mg twice daily) in 305 patients with lower respiratory tract infections. The clinical effectiveness of doxycycline was 84%, compared with 83% for roxithromycin (Charpin J, 1988).

FIGURE 15. Structure of doxycycline

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COMMUNITY-ACQUIRED PNEUMONIA

Adverse events most commonly associated with doxycycline include stomach upset, diarrhea, nausea, headache, and vomiting; however, phototoxicity, dizziness, and vertigo can also occur. Doxycyline has been associated with esophagitis and esophageal injury (Ovartlarnporn B, 1991). Aminoglycosides Overview. Aminoglycosides are the preferred agents for treating serious infections caused by aerobic gram-negative bacilli. Aminoglycosides play only a minor role in the treatment of CAP; their use is typically limited to hospitalized patients with severe or complicated CAP in which gram-negative infections (particularly P. aeruginosa) are suspected. In these situations, physicians may prescribe aminoglycosides in combination with a cephalosporin or carbapenem. Gentamicin is a prototypical aminoglycoside. Other commonly used agents in this class include the generics tobramycin and amikacin, but these agents will not be discussed further. All aminoglycosides are potentially nephrotoxic and ototoxic (which manifests as auditory or vestibular dysfunction). These concentration-dependent toxicities are reversible with reductions in dosage. Mechanism of Action. Aminoglycosides impair protein synthesis by binding to receptors on the 30S subunit of the bacterial ribosome. Binding to the 30S subunit interferes with the initiation of polypeptide formation, causes the production of defective proteins (misreading of the messenger RNA template), and disrupts ribosome function. The effect of aminoglycosides is bactericidal (concentrationdependent). However, the precise mechanisms by which aminoglycosides exert their bactericidal effect are not entirely understood. Gentamicin. Gentamicin (Schering-Plough’s Garamycin, generics) (Figure 16) has been available since 1966 to treat infections caused by aerobic gram-negative bacilli. A synergistic bactericidal effect against streptococci occurs when gentamicin or other aminoglycosides are used in combination with a cell-wallsynthesis inhibitor (e.g., cephalosporins, glycopeptides). Gentamicin is produced by Micromonospora purpurea and binds to the bacterial 30S ribosomal subunit, causing translational errors that result in cell death. No quality clinical trials are available that assess gentamicin’s efÞcacy and safety in CAP. Like other aminoglycosides, gentamicin is associated with kidney toxicity that requires serum monitoring if the agent is administered for several days or used in patients with renal dysfunction. Side effects also include vestibular dysfunction and hearing loss. Glycopeptides Overview. Glycopeptides are bactericidal antibiotics used principally for treatment of severe gram-positive infections in patients who cannot receive or who have failed to respond to penicillins and cephalosporins. They may also be used in patients suspected of having pathogens resistant to β-lactams and other anti-infectives. Glycopeptides are believed to be the only antibiotics with universal activity (i.e., no resistance has been reported) against S. pneumoniae, and physicians reserve them for severe CAP episodes in which multidrug-resistant

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organisms are suspected or the patient has life-threatening complications such as bacteremia or meningitis. Glycopeptide therapy is relatively well tolerated. The most severe adverse reactions are ototoxicity (dizziness, vertigo, and/or hearing loss) and nephrotoxicity (decreased renal function), but they occur infrequently. Hypersensitivity, of which skin rash is the most common allergic reaction, is reported in 5–10% of patients. Other possible side effects of glycopeptides include GI disturbances, muscle pain, blood disorders (e.g., eosinophilia), and disturbances in liver enzymes. While glycopeptides demonstrate no activity against gram-negative organisms, no cross-resistance between vancomycin and other antibiotics has been reported in gram-positive microbes. Mechanism of Action. Glycopeptides inhibit bacterial cell-wall biosynthesis by blocking glycopeptide polymerization. This effect produces immediate inhibition of cell-wall synthesis and secondary damage to the cytoplasmic membrane. Glycopeptides also alter the permeability of the cytoplasmic membrane and directly inhibit RNA synthesis; however, the impact these effects have on bacterial viability is largely unknown. These agents have a concentration-independent bactericidal action resulting primarily from inhibition of cell-wall synthesis; they are considered slowly bactericidal against staphylococci. Vancomycin. Vancomycin (Lilly’s Vancocin, generics) (Figure 17) is a narrowspectrum, bactericidal glycopeptide antibiotic. Vancomycin was developed in the 1950s and has been available worldwide since the 1960s; it is available in oral and IV formulations. The oral formulation is not systemically absorbed and is used for the treatment of GI infections such as staphylococcal enterocolitis and antibiotic-associated pseudomembranous colitis caused by Clostridium difÞcile. Use of the injectable glycopeptides is limited in CAP, primarily to treat severe gram-positive infections, including multidrug-resistant staphylococci, S. pneumoniae, S. aureus, and S. pyogenes. Vancomycin has a relatively long duration of action and therefore can be given every 12 hours. R1

NH

R2

O H2N

O

NH2

HO

NH2 O O OH

HO

CH3HN

CH3

FIGURE 16. Structure of gentamicin [gentamicin C1 (R1 = R2 = CH3 ), gentamicin C1a (R1 = R2 = H), gentamicin C2 (R1 = CH3 , R2 = H)].

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COMMUNITY-ACQUIRED PNEUMONIA

H3N

HO

Me

OH

HO Me

O

O

O

O Cl O

O H

HO

OH

Cl H O O H NH −O

HN H

O

O H

H N

Me

N N

H

O H

O

H

N H

H

+

NH2 H

O

2C

H2N

OH HO

OH FIGURE 17. Structure of vancomycin

Vancomycin has been available for a long time, and no high-quality clinical studies pertaining to CAP have been conducted in recent years. Therefore, this section does not cite speciÞc efÞcacy or safety results from clinical studies. Vancomycin is contraindicated in patients with a history of hypersensitivity to this antibiotic or other glycopeptides. Ototoxicity has occurred in patients receiving vancomycin and can be either transient or permanent. Most cases have been reported in patients receiving high IV doses or in patients with a history of hearing complications. Vancomycin is also associated with nephrotoxic effects, and patients should be screened for a history of renal complications. Renal failure has been reported in patients receiving high doses of IV vancomycin. Vancomycin is also associated with infusion-related ßushing, known as “red man’s syndrome.” Resistance to vancomycin can be achieved by a modiÞcation of the components of the bacterial cell wall. Vancomycin binds to a component of the cell wall that when modiÞed or altered results in reduced binding of the drug to the bacteria. Teicoplanin. Teicoplanin (SanoÞ-Aventis’ Targocid) is a narrow-spectrum, bactericidal glycopeptide antibiotic similar in structure to vancomycin. It was Þrst launched in France and Italy in 1988 and by 1997 was available in most of Europe. It was launched in Japan in 1997, where Aventis has partnered with Fujisawa for distribution and copromotion, but has never been available in the United States, where development was suspended in 1999. This agent is expected to lose patent protection in Japan and European markets between 2004 and 2008.

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Teicoplanin is available in parenteral formulations and has a signiÞcantly longer duration of action than vancomycin, thereby allowing once-daily administration. Unlike vancomycin, teicoplanin can be given by IM injection as well as IV injection. Considered a valuable alternative to vancomycin, teicoplanin has a spectrum of activity that includes both aerobic and anaerobic gram-positive bacteria, including methicillin-resistant S. aureus (MRSA). For CAP, use of the agent is generally limited to complicated cases in which Staphylococcus is suspected and to patients who cannot receive or have failed to respond to the penicillins or cephalosporins. A multicenter study compared the efÞcacy of teicoplanin and ciproßoxacin with that of ceftriaxone in the treatment of CAP in elderly patients and in patients with coexisting diseases (such as COPD and diabetes mellitus). Patients received either IV teicoplanin (400 mg/day) plus oral ciproßoxacin (1 g/day) or IV ceftriaxone (2 g/day) in a randomized, single-blind manner. Cure or improvement was observed in 39 (97.5%) of 40 combination therapy recipients and 36 (83.7%) of 43 ceftriaxone recipients. Teicoplanin has less propensity than vancomycin to cause nephrotoxicity, ototoxicity, or infusion-related ßushing (red man’s syndrome). However, it does share some of the same adverse reactions, including ototoxicity and nephrotoxicity. Hypersensitivity reactions should also be a concern of physicians considering teicoplanin therapy. Oxazolidinones Overview. The oxazolidinones—which include only one approved agent, linezolid (PÞzer’s Zyvox)—are an antimicrobial class with a unique mechanism of action. They are active against severe, resistant gram-positive infections, including those caused by methicillin-susceptible and methicillin-resistant S. aureus (MRSA) and penicillin-resistant S. pneumoniae (PRSP). The oxazolidinones are active against anaerobes and display modest activity against H. inßuenzae but are not active against gram-negative enterobacteriaceae. The oxazolidinones are available both orally and parenterally and have a bioavailability of 100%. The role of linezolid in CAP is limited to patients with multidrug-resistant, gram-positive infections (such as PRSP with cross-resistance to other antibiotics). Mechanism of Action. Oxazolidinones bind to a site on the bacterial 23S ribosomal RNA of the 50S subunit, thereby preventing the formation of a functional 70S initiation complex, which is an essential component of the bacterial translation process. No direct action on DNA or RNA synthesis has been observed. Oxazolidinones are bacteriostatic against staphylococci and enterococci and may be slowly bactericidal against other gram-positive bacteria. Linezolid. Linezolid (PÞzer’s Zyvox) (Figure 18) was launched in the United States and the United Kingdom in 2000 and 2001, respectively. In June 2001, linezolid was launched in Japan for the treatment of vancomycin-resistant enterococcal (VRE) infections. Available in injection, tablet, and oral suspension formulations, linezolid is active against infections caused by gram-positive bacteria. Although approved for use in CAP in the United States and Europe, linezolid

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COMMUNITY-ACQUIRED PNEUMONIA

O O

N

N

O

O HN

F

CH3

FIGURE 18. Structure of linezolid

is usually reserved for serious infections caused by multidrug-resistant staphylococci. A randomized, multicenter, open-label Phase III trial of 747 patients with demonstrated or presumptive S. pneumoniae pneumonia requiring hospital admission compared linezolid (600 mg IV, every 12 hours) followed by linezolid (600 mg orally, every 12 hours) with the cephalosporin ceftriaxone (1 g IV, every 12 hours) followed by cefpodoxime (200 mg orally, every 12 hours) (San Pedro GS, 2002). Both test groups were treated for 7 to 14 days. The clinical success rates were 83% for linezolid and 76.4% for ceftriaxone/cefpodoxime. The microbiological eradication rates were 88.7% and 89.9%, respectively. In the subpopulation of patients with bacteremia, linezolid was more effective than ceftriaxone/cefpodoxime: 93.1% versus 68.2%. Linezolid has been associated with hematologic abnormalities (e.g., thrombocytopenia, anemia) that are consistent with mild, reversible, duration-dependent myelosuppression (suppression of bone marrow activity); this effect limits prolonged use. No other signiÞcant side effects for the drug have been reported. EMERGING THERAPIES Many agents are in development for the treatment of community-acquired pneumonia (CAP). The majority of drugs in late-stage development are antibiotics that build on existing drug classes (e.g., ßuoroquinolones). Reßecting the limited number of innovative leads, the high attrition rates, and the high development costs relative to the return, no novel classes have been introduced in the past two decades, with the exception of the oxazolidinone linezolid (PÞzer’s Zyvox), which primarily targets niche antibacterial market segments such as complicated hospital infections. Table 6 summarizes emerging antibacterial agents in late-stage development and their expected sales potential in the CAP market. The antibiotics currently available for treatment of CAP are highly effective and well tolerated. Therefore, the barriers to new agents’ entry into the CAP market are relatively high. However, given the severity of this infection and its relatively high mortality, the CAP market remains relatively receptive to novel agents that can provide a clinical or economic beneÞt. The macrolides and ßuoroquinolones launched over the past decade (e.g., azithromycin, moxißoxacin) have provided advantages such as higher activity against respiratory pathogens, broader spectrums of activity, and more convenient dosing. Emerging agents in these two classes will face intense competition to differentiate themselves in the market. However, one of the drivers of future opportunity within these classes

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TABLE 6. Emerging Therapies in Development for Community-Acquired Pneumonia, 2004 Compound

Development Phase

Marketing Company

Fluoroquinolones Garenoxacin United States Europe Japan

PR III III

Schering-Plough Schering-Plough Toyama

DHFR Inhibitors Iclaprim United States Europe Japan

— IIa —

— Arpida —

a For skin infections.

DHFR = Dihydrofolate reductase; I = Phase I; II = Phase II; III = Phase III; PC = Preclinical; R = Registered; S = Suspended.

will be development of agents capable of overcoming resistant pathogens, particularly as prevalence of resistance to β-lactams and macrolides—and eventually, ßuoroquinolones—continues to grow. Furthermore, opportunity for emerging ßuoroquinolones exists in pediatrics because this class is generally contraindicated in children. The highest unmet needs in CAP are those for hospitalized patients with severe infections, particularly patients admitted to the intensive care unit (ICU). The majority of CAP cases are mild, but mortality in hospitalized patients reaches 5–20%, and mortality in patients admitted to ICUs ranges from 35% to more than 50% (Bochud PY, 2001; Fine MJ, 1993; Fine MJ, 1996; Fine MJ, 1999). The recent upsurge in the development of the streptogramins, oxazolidinones, and other narrow-spectrum agents reßects the demand for new agents that are active against problem pathogens, particularly those that are resistant to current therapies for critically ill patients. In analyzing emerging antibacterial agents, several factors must be considered that will be important for success in the CAP market. First, the spectrum of activity must provide appropriate coverage of key pathogens, particularly Streptococcus pneumoniae and Haemophilus inßuenzae and the atypical pathogens such as Chlamydia pneumoniae, Mycoplasma pneumoniae, and Legionella pneumophila. Second, to be commercially successful, new agents must offer competitive advantages such as activity against resistant pathogens (e.g., drug-resistant S. pneumoniae), a higher barrier to resistance, and/or be extremely well tolerated. Because there are numerous once-daily options already on the market, emerging products must have once-daily dosing. Because the CAP market has a large hospital segment, availability in both oral and intravenous (IV) formulation is important for wide uptake. Finally, a short treatment course is highly desirable because this attribute can improve compliance and reduce overall cost.

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COMMUNITY-ACQUIRED PNEUMONIA

As discussed in the “Overview” section in “Current Therapies,” the measure of in vitro potency of antibiotics is the minimum inhibitory concentration (MIC) of antibiotic required to inhibit growth of a bacteria. In clinical trials, the primary end points used to evaluate antibiotics are clinical response rate or clinical cure rates, which refer to the improvement in subjective symptoms and/or objective measures such as chest X ray. Bacterial, or microbiological, eradication rate refers to the elimination of pathogens in sputum and/or in respiratory tract samples. Fluoroquinolones Overview. The ßuoroquinolones have proved to be one of the most commercially and clinically successful antibacterial classes for the treatment of CAP. Their broad spectrum of efÞcacy (improving with more recent launches against respiratory pathogens), safety, and convenient once-daily dosing have earned them a favorable position in the minds of clinicians. Their main advantages in CAP have been their broad-spectrum activity, particularly their high H. inßuenzae activity, compared with other agents such as the macrolides. Also, their availability in oral and parenteral form has provided an important clinical advantage. The concerns associated with this class of agents include their potential to induce abnormal cardiac conductions (i.e., prolongation of the QT interval on electrocardiograms) and liver toxicity, which was associated with one drug (PÞzer’s trovaßoxacin) that was subsequently withdrawn. Also, ßuoroquinolones currently on the market are contraindicated for use in children owing to cartilage growth concerns. Emerging ßuoroquinolones in development provide potential advances over currently marketed agents, mainly in terms of higher potency against respiratory pathogens, retained activity against resistant pathogens, and a favorable safety proÞle. Mechanism of Action. Fluoroquinolones act by inhibiting bacterial topoisomerase IV and DNA gyrase, which are enzymes required for DNA replication, transcription, repair, and recombination. The exact nature of the interaction of quinolones with their target enzymes is not completely understood. However, this interaction blocks the progression of DNA replication, leading to strand breaks and rapid cell death. These agents are potent inhibitors of nucleic acid synthesis. Garenoxacin. Garenoxacin (Schering-Plough/Toyama’s T-3811, des-F(6)-quinolone garenoxacin) (Figure 19) is a potent, oral, broad-spectrum quinolone that has been submitted to the FDA for approval. Garenoxacin has a broad spectrum of activity, similar to that displayed by the other new ßuoroquinolones (Bassetti M, 2002; Christiansen KJ, 2004; Kirby JT, 2002; Takahata M, 1999; Gordon KA, 2002). Garenoxacin shows the highest antibacterial activity against S. pneumoniae when compared with ciproßoxacin, levoßoxacin, gatißoxacin, and moxißoxacin, and it is more potent than ciproßoxacin, levoßoxacin, and moxißoxacin against anaerobes (Bassetti M, 2002). It is also highly potent against ciproßoxacin-resistant strains of S. pneumoniae and Staphylococcus aureus, and like gatißoxacin, it is associated with a lower risk

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O CO2H

N HN

O CHF2

FIGURE 19. Structure of garenoxacin

of promoting resistance development (Boswell FJ, 2001; Hartman-Neumann S, 2001; Schmitz FJ, 2001). Garenoxacin has favorable pharmacokinetics and is expected to be administered as an oral or injectable once-daily formulation. A 28-day Phase II trial compared the safety and tolerability of garenoxacin, dosed orally at 400 mg once daily, with that of placebo in 45 healthy volunteers; the trial showed the drug to be both safe and well tolerated, with no serious adverse effects (Stewart C, 2001). Similarly, Phase II clinical trial results for the IV formulation show garenoxacin to be safe and well tolerated, with no serious adverse effects (Gajjar D, 2001[a]). No clinically signiÞcant effects on liver function and QT interval have been reported (Gajjar D, 2001[b]). In an open-label, multicenter, noncomparative study involving 208 patients with CAP, researchers observed clinical cure in 161 (91%) of 176 evaluable patients treated with garenoxacin dosed orally at 400 mg once daily for ten days (Arnow P, 2001). In 125 bacteriologic-evaluable patients, 178 (89%) of 199 isolated pathogens were eradicated after treatment with garenoxacin. The same study found that 25% of patients experienced more than one adverse event, including diarrhea (5%), nausea (4%), headache (2%), vaginitis (2%), dizziness (2%), and dry mouth (1%). At present, ßuoroquinolones are not approved for children younger than 18 years because they have been shown to cause arthropathy and cartilage damage in immature animals. (Even so, ciproßoxacin has been used in children for compassionate reasons—for example, life-threatening infections in pediatric patients with cystic Þbrosis.) However, toxicological studies of garenoxacin’s effect on immature and juvenile joint cartilage have shown that this drug does not induce cartilage defects or signiÞcant arthropathy in immature rats or dogs (Kappel EM, 2002; Kastner M, 2004; Nagai A, 2002). If its low chondrotoxic potential proves to be clinically relevant, garenoxacin could be used more widely than other emerging ßuoroquinolones; indeed, it could become the Þrst ßuoroquinolone to be approved for use in all age-groups. Overall, garenoxacin provides a favorable activity and safety proÞle. However, the agent may be the fourth respiratory ßuoroquinolone on the market (following gatißoxacin, moxißoxacin, and gemißoxacin). Schering-Plough has signiÞcant marketing prowess in the respiratory market, which gives it a signiÞcant advantage in promoting the product. However, Schering-Plough recently entered into

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a strategic agreement with Bayer to market Bayer’s moxißoxacin in the United States. It is unclear at this time whether this agreement will affect the development and prioritization of garenoxacin. Overall, however, garenoxacin is likely to face stiff competition in the CAP ßuoroquinolone market. Dihydrofolate Reductase Inhibitors Overview. The dihydrofolate reductase (DHFR) inhibitors target the bacterial DHFR, an enzyme involved in the folic acid pathway, thereby disrupting bacterial replication. DHFR inhibitors target both gram-positive and gram-negative bacteria, and could therefore serve as broad-spectrum antibacterials. The most common folic acid biosynthesis inhibitor, trimethoprim/sulfamethoxazole (Roche’s Bactrim, Monarch’s Septra, generics), was used in the past to treat RTIs such as mild pneumonia and sinusitis. However, this agent’s use has been limited by the emergence of S. pneumoniae that are resistant to trimethoprim/sulfamethoxazole, concerns about sulfa hypersensitivity, and poor activity against atypical RTI organisms. Newer drugs in this class are being developed that overcome current mechanisms of resistance and provide a better spectrum of coverage against RTI pathogens. These agents may become increasingly important over the next 10 to 20 years as resistance to current drug classes builds. Mechanism of Action. The folic acid pathway inhibitors block the action of the bacterial dihydrofolate reductase (DHFR) enzyme, an enzyme essential for the synthesis of thymine nucleotides. Inhibiting the synthesis of thymine nucleotides prevents DNA and RNA synthesis, thereby killing the bacteria. More speciÞcally, in thymine synthesis, folate donates a methyl group to uracil to make thymine; in the process, the folate molecule is oxidized and must be reduced by DHFR to allow another cycle of synthesis. If DHFR is inhibited, the absence of reduced folate causes a precipitous drop in the levels of thymine nucleotides, thereby causing uracil to be erroneously incorporated in place of thymine nucleotides in growing DNA strands. Replication is eventually disrupted by this process and the DNA fragments, killing the bacteria (Quinlivan EP, 2000; Schneider P, 2003). Antibacterials that target the bacterial DHFR protein are generally not toxic to human cells because human and bacterial DHFR are different enough to allow selective targeting of bacterial DHFR. Iclaprim. Arpida licensed all commercialization rights to the DHFR inhibitor iclaprim (AR-100) from Roche. The drug has the potential to be used against serious antibiotic-resistant infections, including RTIs. The agent has completed Phase II trials in Europe in treatment of skin infections. While no information is available regarding its development for CAP, iclaprim’s spectrum of activity makes it a potential emerging candidate for the treatment of CAP. Previous clinical trials have evaluated an IV formulation of iclaprim, but Arpida is reportedly developing an oral formulation as well. Iclaprim targets a range of gram-positive and gram-negative respiratory pathogens, and results from in vitro studies suggest that the potency of iclaprim against C. pneumoniae is favorable and comparable to the potency of leading antibacterials used in RTIs, such as azithromycin and levoßoxacin (Kohlhoff SA, 2004). Arpida reports the agent to have excellent

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activity against S. pneumoniae and H. inßuenzae. Additionally, the agent is active against MRSA and vancomycin-resistant Staphylococcus aureus. Phase II clinical trial results reported in December 2003, showed that the agent is well tolerated and has high clinical and microbiological response rates in skin infections. No data on CAP are available. Although this agent is being developed for skin infections, it demonstrated good activity against gram-positive and gram-negative pathogens involved in RTIs, such as S. pneumoniae and H. inßuenzae, and intracellular pathogens such as Chlamydia. Therefore, it has the potential to be developed for CAP. Availability in both oral and IV forms can provide the agent with an advantage in the hospital setting and in oral step-down therapy. REFERENCES Abbanat DR. In vitro and in vivo antibacterial activities of 15-methyl-2-ßuoro ketolides. Abstracts of the 43rd Interscience Conference on Antimicrobial Agents and Chemotherapy (ICAAC). Chicago, 2003. Abstract 1203. Abramowicz M. The choice of antibacterial drugs. Medical Letter on Drugs and Therapeutics. 1999;41:95–104. Adam D. Global antibiotic resistance in Streptococcus pneumoniae. Journal of Antimicrobial Chemotherapy. 2002;50(suppl):1–5. Adams PF, et al. Current estimates from the National Health Interview Survey, 1996. National Center for Health Statistics. Vital and Health Statistics. 1999;Series 10(200):12. Almirall J, et al. Risk factors for community-acquired pneumonia in adults: a populationbased case-control study. European Respiratory Journal. 1999;13:349–355. Almirall J, et al. Epidemiology of community-acquired pneumonia in adults: a populationbased study. European Respiratory Journal. 2000;15:757–763. American Thoracic Society. Guidelines for the management of adults with communityacquired pneumonia. American Journal of Respiratory and Critical Care Medicine. 2001;163:1730–173;1754. Anderson G, et al. A comparative safety and efÞcacy study of clarithromycin and erythromycin stearate in community-acquired pneumonia. Journal of Antimicrobial Chemotherapy. 1991;27 Suppl A:117–124. Apfel CM, et al. Peptide deformylase as an antibacterial drug target: target validation and resistance development. Antimicrobial Agents and Chemotherapy. 2001;45(4):1058–1064. Appelbaum P, et al. Antipneumococcal activity of NVP-PDF713 compared with 18 other agents. Clinical Microbiology and Infection. 2004;10(suppl 3):239 (plus poster) abstr. P916. [a] Appelbaum P, et al. Antistaphylococcal activity of NVP-PDF713, a new peptide deformylase inhibitor compared with other agents. Clinical Microbiology and Infection. 2004;10 (suppl 3):239 (plus poster) abstr. P915. [b] Arnow P, et al. An open-label, multicenter, noncomparative study of oral BMS-284756 in the treatment of community-acquired pneumonia. Abstracts of the 41st Interscience Conference on Antimicrobial Agents and Chemotherapy (ICAAC), Chicago, December 2001. Abstract 449.

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Complicated Skin and Soft Tissue Infections

ETIOLOGY AND PATHOPHYSIOLOGY Definition Skin and soft-tissue infections (SSTI), including those considered to be “complicated,” are thought of as a summation of many smaller, individual infection diagnoses that have numerous etiologies and infecting pathogens. Impetigo, folliculitis, furuncles, carbuncles, cellulitis, erysipelas, abscesses, necrotizing fasciitis, and gangrene are all considered to be SSTIs (Figure 1). This study deÞnes a complicated skin and soft-tissue infection (cSSTI) as an infection for which the primary diagnosis is an SSTI that requires hospitalization. Many other deÞnitions exist in which an SSTI is considered complicated based on the depth of skin involvement, resistance of the infection to therapy, and the number of pathogens involved. However, for the purposes of this study, a cSSTI is deÞned as any hospitalized SSTI case in which the primary diagnosis is cellulitis, abscess, necrotizing fasciitis, or gangrene (ICD-9 codes 680–686, 707.0, and 785.4); mastitis (ICD-9 code 675); or infected diabetic ulcers (ICD-9 codes 250.7 and 250.8). In addition, impetigo, folliculitis, furuncles, carbuncles (also covered in ICD-9 codes 680–686), and mastitis are included in the cSSTI deÞnition, even though they are rarely “complicated.” Wound, burn, and surgical site infections are not included because they can often extend to other tissue layers. Pathophysiology Impetigo. Impetigo is the most common cutaneous infection in children, with seasonal incidence peaking in August through November. Impetigo is a superÞcial Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

571

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FIGURE 1. Anatomy of skin layers and corresponding infections.

infection of the skin that has two subforms: impetigo contagiosa and bullous impetigo. Impetigo contagiosa is most commonly found on the legs, while bullous impetigo is more often found on the face and trunk. Patients with impetigo generally do not have a fever and generally do not have systemic symptoms or toxicity. Impetigo contagiosa does not have a known etiology, because inoculation of normal skin with group A beta-hemolytic streptococcus or coagulase-positive Staphylococcus aureus (the two commonly isolated pathogens of impetigo) does not actually cause the disease. It is possible that insect bites or infected abrasions may play a role in the infection. Impetigo contagiosa begins as very small papules (solid raised bumps on the skin) that rapidly progress to pustules (papules with an inßamed base that are Þlled with pus). Within 24 hours, these pustules rupture and the contents crust over the infection site. The crust darkens over Þve to six days, and the lesions tend to clear from the center toward the outer edges. Bullous impetigo, commonly seen in newborns, starts as small pustules that rapidly grow to a bulla (a vesicle containing ßuid) that can reach a diameter of three centimeters. When the bulla ruptures, it leaves a red base on the skin. However, bullous impetigo clears rapidly, often faster than impetigo contagiosa (Meislin HW, 2002). Impetigo of either type rarely becomes complicated, and most cases are treated as uncomplicated SSTIs in outpatient settings.

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Folliculitis, Furuncles, and Carbuncles. Folliculitis is a generally mild infection of the hair follicle. It presents clinically as a series of painful red bumps centered on a hair follicle and is found naturally in the hairy parts of the body. It is most common in areas of heavy friction and sweat, like the back of the neck. Furuncles (boils) result from the evolution of folliculitis into a more severe infection, with involvement of deeper layers of the skin and follicle. Furuncles are characterized as hardened painful bumps surrounding the hair follicle, with a diameter of one to two centimeters. As the infection progresses, a furuncle can rupture, effusing pus and necrotic tissue. Following rupture (either spontaneous or surgical), inßammation subsides rapidly, though satellite furuncles often appear at other sites. Neither folliculitis nor furuncles are associated with systemic symptoms, though diffuse furunculosis may be associated with mild fever and malaise. Carbuncles are the most serious extension of folliculitis and are generally localized to the back of the neck. Carbuncles are the deep-seated (spreading into the subcutaneous tissue) infection of multiple hair follicles, with multiple sinus tracts forming within the tissue. Carbuncles generally resolve as a central necrotic crater, which gradually hardens over, leaving an enlarged purple scar. Because it is such a deep infection, carbuncles are more often associated with general symptoms like fever and chills than is folliculitis or furuncles. These systemic symptoms can be indicators of staphylococcal septicemia (Mandell GL, 2000). Most folliculitis, furuncles, and carbuncles are uncomplicated and are treated in outpatient settings; however, in a few instances these infections can progress to complicated abscesses because of resistant bacteria or other underlying conditions. Cellulitis and Erysipelas. Cellulitis is an infection of the superÞcial skin layers. It comes in many forms, often differentiated by the microbes causing the infection, as well as the site of infection. Cellulitis is more common in the lower extremities, with upper extremities second in frequency, followed by the face. The disease is marked by tenderness of the red and swollen infected area and can spread to surrounding skin if untreated. Fever is uncommon in cellulitis, but it can be an indicator of septicemia. Although cellulitis can sometimes be traced to trauma or breaks in the skin, an entry point is not found in a majority of cases. Cellulitis can also spread to skin through the circulatory or lymphatic systems. Staphylococcus aureus and Streptococcus pyogenes are the most commonly isolated pathogens responsible for cellulitis, although the microbial etiology of cellulitis generally reßects the bacteria found at the site of infection. Although there are numerous etiologies and presentations of cellulitis, several subtypes are prominent enough to warrant discussion. Cellulitis caused by Haemophilus inßuenzae occurs primarily in children. This cellulitis is distinguished by a high fever and elevated white blood cell count and is found mostly on the face or extremities. However, since the advent of the H. inßuenzae vaccine, its incidence has decreased dramatically. Orbital cellulitis involves the face, speciÞcally the area surrounding the eyes. This cellulitis

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is commonly caused by streptococci and must be treated aggressively because the veins serving this area drain through communicating vessels into the brain; complications can be extreme. Orbital cellulitis is characterized by swelling of the eyelid, redness of the skin, and warmth at the site of infection. Vision and eye movement are not affected. This type of cellulitis usually occurs in children and is predominantly found in conjunction with sinus infections. Streptococcal cellulitis (also known as ascending cellulitis) progresses quickly from infected extremities, often with noticeable lymphatic streaking and swollen tissue. It is commonly found after surgery or trauma, though neither is necessarily the cause. Without treatment, bacteria replicate quickly and release large amounts of toxin, putting patients at risk for toxic shock syndrome. Staphylococcal cellulitis is a painless infection and is generally less toxic than streptococcal cellulitis. Lesions are more localized and lead more readily to abscesses. Staphylococcal scalded skin syndrome is caused by phage group II, type 71 staphylococci and is found in children aged six months to six years. These staphylococci release an exfoliating toxin that separates the skin at the layer called the zona granulosa, resulting in widespread pain and blistering. The skin can form numerous ßuidÞlled sacs, leading to the sloughing of large sections of skin and resulting in a “scalded” appearance. Erysipelas is a form of cellulitis commonly caused by group A streptococci and is distinct for its involvement of cutaneous lymphatic vessels. This disease occurs primarily among infants and people older than age 30, and it can have varying causes. When it occurs on the face, it is generally preceded by a streptococcal sore throat; when it appears on the trunk or limbs, it can often be traced to a wound or surgical site. However, the mode of spread to the skin is not clear. Erysipelas can be distinguished clinically from cellulitis by its markedly raised or swollen appearance and a clear border between infected and uninfected tissue. The disease often begins as a red, swollen patch and spreads rapidly outward. Infected tissue often has signiÞcant ßuid accumulation. Erysipelas has general symptoms of fever, chills, and systemic toxicity (Meislin HW, 2002). Abscesses. Abscesses are deeper and more localized infections than cellulitis and are commonly complicated. An abscess is a pocket of pus, forming a ßuctuant (with a ßuid-like movement when touched) and a painful soft-tissue mass with a Þrm, red tissue border. The etiology of an abscess depends on its location and is often the result of trauma to the skin that allows inÞltration of microbes colonizing that particular skin surface. Abscesses are not associated with a fever or systemic toxicity, but they generally present with pain and lymphangitis (inßammation of the lymphatic vessels). Abscesses can be uncomplicated and treated in an outpatient setting, but because they can often be deep and require surgical drainage, they contribute signiÞcantly to the overall cSSTI patient population. Abscesses of the head and neck are often the result of blocked apocrine sweat glands and are more prevalent in adults because their sweat glands are more active than those in prepubescent children. Skin abscesses are generally colonized by typical skin ßora, often Staph. aureus. Perirectal and perioral abscesses, in contrast, are usually colonized by anaerobes that dominate the mucous membranes (Meislin

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575

HW, 2002). Intravenous drug users, especially those who inject drugs under the skin when veins are no longer available, get abscesses more often than any other type of skin infection. Infecting microbes in these patients can vary widely because they are often introduced from unclean needles (Ebright JR, 2002). Necrotizing Fasciitis and Gangrene. Necrotizing fasciitis is a disease that has received signiÞcant publicity in recent years as one of the more lethal cSSTIs, and most of this publicity has focused on cases resulting from a group A streptococcus, Streptococcus pyogenes (which the media has termed “ßesh-eating bacteria”). Strep. pyogenes produces a cell-wall protein that separates the connective tissue of the dermis, producing signiÞcant inßammation and necrosis. However, necrotizing fasciitis can also be polymicrobial in nature, including both aerobes and anaerobes. Generally, the deeper the infection, the more normal the skin surface appears. The underlying toxicity is often far greater than is apparent from the appearance of the skin surface. Thus, necrotizing fasciitis can be difÞcult to spot when a patient Þrst presents. Additionally, the lack of pain felt by patients with necrotizing fasciitis can be deceiving because nerve endings die quickly in infected tissue. Therefore, the elimination of pain in patients does not necessarily signal that the infection is subsiding; it can, in fact, mean the opposite. Necrotizing fasciitis is an infection of the underlying fascia of the skin. It can evolve rapidly from even the most minor of skin punctures and may progress from a mild reddened area to pronounced, purple-colored inßammation highlighted with bullae that are Þlled with ßuid within 24–72 hours. As soon as 24–48 hours later, tissue will become necrotic or gangrenous and slough off. Not only will necrotizing fasciitis spread quickly over the skin surface, but infection can also spread into the muscle. In addition to the cutaneous pathogenesis of necrotizing fasciitis, this infection is associated with systemic toxicity, which can lead to fever, elevated heart rate, disorientation, and eventually shock if it is not treated quickly and effectively. Gangrene is a general term for any necrotized tissue that has lost its vascular supply. In infectious disease terms, gangrene is often used to refer to gas gangrene resulting from infection with Clostridium family members, normally C. perfringens. This type of gangrene occurs primarily as a superinfection after trauma, surgery, or necrosis, with at least three days before presentation. This infection spreads through the fascia and does not involve the muscle. (Clostridial myonecrosis, which does involve the muscle, is not included in this study.) Infecting Clostridium bacteria release gas in the intrafascial layers, resulting in a spongy texture when pressed. Patients have systemic symptoms, including fever and elevated heart rate, but the site of infection has minimal pain with mild reddening. The site may have a foul-smelling discharge containing bacteria and neutrophils (Mandell GL, 2000; Meislin HW, 2002). Diabetic Ulcer Infections. Diabetic patients are at increased risk for SSTIs, largely because of diabetic neuropathy. Peripheral neuropathy of the extremities leads to loss of muscle tone and reßexes, which, combined with the loss of

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FIGURE 2. Diagram of mastitis.

feeling, can lead to increased incidences of blistering and trauma of the skin from abrasion. The lower extremities are particularly prone to such a scenario, leading to the common condition of “diabetic foot.” Diabetic patients also suffer from autonomic neuropathy, which can cause peripheral vascular disease and reduced sweating in the foot. These factors combine to create dry and cracked skin. Because of a loss of sensation, the affected areas are subject to increased and initially unnoticed abrasion or trauma. When an infection develops, low tissue perfusion inhibits an appropriate immune response and healing. Thus, many diabetic patients develop complicated infected foot ulcers, often requiring radical debridement or eventual amputation (Larsen PR, 2003; Richardson EG, 2003; Wooten MK, 2001). Table 1 details the levels of severity of diabetic ulcers based on the Wagner scale. SuperÞcial infection of diabetic ulcers is often due to Staph. aureus or group A or B streptococci, while deeper infections are more often caused by a mix of aerobic and anaerobic bacteria. Pseudomonas aeruginosa is more likely to be seen in these infections than in other cSSTIs (File TM, 1995). Mastitis. Mastitis is an infection of the breast that is relatively common in nursing mothers, usually in the Þrst few weeks post partum (Figure 2). Cracked nipples can allow bacteria from the baby’s mouth, the hospital environment, or the mother’s skin to enter the tissue. The vast majority of patients with mastitis do not require hospitalization. In rare cases, mastitis will progress from a painful nodule to a deep abscess and become complicated, in most cases requiring surgical incision and drainage. Staph. aureus is the typical etiological agent, though others are sometimes identiÞed (Mandell GL, 2000). Other rare cases of mastitis can develop from breast surgery, but these are not covered in this study.

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577

TABLE 1. Wagner Diabetic Foot Ulcer Classification System Grade 0 1 2 3 4 5

Description No ulcer, but at high risk (e.g. bony deformities, neuropathy) Superficial ulcer without infection Deep ulcer, extending to tendons or bone Deep ulcer or abscess, extending to and involving bone (i.e., osteomyelitis) Partial gangrene, involving toes or forefoot Gangrene of entire foot

Source: Adapted from Wagner FW. The dysvascular foot: a system of diagnosis and treatment. Foot Ankle. 1981;2:64–122.

Etiology The microbial cause of any one cSSTI usually depends on the bacterial ßora normally present on the skin surrounding the infection site. Additionally, if the infection is acquired in a hospital setting, the etiological cause is much more likely to be methicillin-resistant Staph. aureus (MRSA) than if the infection is acquired in a community setting. Because diagnostic techniques are not very accurate for most cSSTIs, physicians often treat patients empirically, based on microbes known to be prevalent at the infection site. Table 2 segments common bacterial pathogens by Gram’s staining and aerobic/anaerobic growth. The major bacterial pathogens are discussed in the following sections; however, because anaerobes are rarely the causative pathogen in cSSTIs, they are not discussed in this report. Staphylococci. The staphylococci are among the most important SSTI pathogens. Staphylococci are nonmotile, gram-positive bacteria that are usually found on TABLE 2. Segmentation of Common Bacterial Pathogens Aerobe Gram-positive

Staphylococci Staph. aureus, Staph. epidermis

Streptococci Strep. pyogenes Gram-negative

Pseudomonads Pseudomonas aeruginosa Enterobacteriaceae Escherichia coli Shigella Salmonella Klebsiella

Anaerobe Clostridia Clostrium tetani, Clostrium perfringens, Clostrium botulinum, Clostrium difficile Peptostreptococci Peptococci Bacteroides fragilis Veillonella Acidominococcus

Note: Staphylococci, streptococci, pseudomonads, and several enterobacteriaceae are critical pathogens for cSSTIs, though the other types may be found on rare occasions. Some aerobes, especially in the enterobacteriaceae family, are facultative anaerobes, meaning they can live with or without oxygen.

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COMPLICATED SKIN AND SOFT TISSUE INFECTIONS

the skin and mucous membranes of the body (particularly the perianal area, nasal membranes, and hair follicles). Staphylococci are easily distinguished from streptococci using a catalase test; staphylococci are catalase-positive, and streptococci are catalase-negative. A second enzyme, coagulase, is the primary characteristic used to differentiate staphylococcal strains. Staph. aureus is coagulase-positive, and other, “less pathogenic” strains such as Staphylococcus epidermidis are coagulase-negative. Numerous staphylococcal virulence factors have been identiÞed (Table 3), including factors that promote colonization, protect the bacteria from the host defense systems, or facilitate the disease process. These virulence factors are prime targets for future drug development because they are the etiologic agents that are required for infection and include the molecules that can lead to toxic shock. The precise mechanisms and extent to which these virulence factors contribute to the pathogenesis of various staphylococcal diseases are largely unknown. Staphylococci are the major etiologic pathogens for cSSTIs and are isolated in a majority of cases. For most SSTIs, physicians will empirically suspect staphylococci and choose antibiotics accordingly. Staphylococcus aureus. Staph. aureus is a pyogenic (pus-forming) bacterium that has almost always been recognized as an opportunistic pathogen. Although individual strains of Staph. aureus differ in their ability to invade and establish infection, infection essentially relies on the capacity of this bacterium to penetrate the skin or mucous membranes (Day NPJ, 2001; Peacock SJ, 2002). It is able to do so when the skin surface is broken, when the mucosal surface is damaged, or when it is introduced into the host via an external source. Adherence of Staph. aureus to host tissues is a critical step in the colonization and subsequent establishment of infection. Evidence is also emerging that this pathogen can invade and persist within host cells, where it is protected from host defenses and most antimicrobial agents (Jett BD, 2002; Massey RC, 2001). Investigators have found that Þbronectin-binding proteins and other cell-wall adhesins of the MSCRAMM (microbial surface components recognizing adhesive matrix molecule) family enable Staph. aureus to adhere to host tissues and to invade intracellular space (Cucarella C, 2001; Menzies BE, 2002). Also, nearly all clinical isolates of Staph. aureus have a polysaccharide capsule that likely plays an important role in the pathogenesis of Staph. aureus infections by inhibiting phagocytosis. Methicillin-Resistant Staphylococcus aureus. Methicillin-resistant Staph. aureus (MRSA) is among the most feared bacteria associated with any infection. These bacteria are often resistant to most β-lactam antibiotics (such as penicillin), macrolides, tetracyclines, ßuoroquinolones, and aminoglycosides. Several pandemic clones of MRSA have been identiÞed that are associated with the majority of MRSA infections worldwide (Oliveira DC, 2001). It is widely believed that the spread of MRSA is predominantly caused by the clonal expansion of these lineages rather than horizontal transfer of genetic material. These clones are

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TABLE 3. Select Virulence Factors of Staphylococcus aureus and Staphylococcus epidermidis Virulence Factor

Description

Extracellular enzymes and bacterial cell surface proteins Staphylocoagulase A free coagulase that produces true clotting of plasma. Staphylococcal A bound coagulase in the cell wall that induces clumping factor clumping of cells in the presence of fibrinogen. Fibronectin-binding Similar to staphylococcal clumping factor, FnBP is protein (FnBP) involved in adherence to surfaces coated with host-derived proteins. FnBP is also implicated in the adherence and invasion of staphylococci into vascular endothelial cells. Protein A Binds immunoglobulin molecules and is a very effective B-cell mitogen (stimulates B-cell production). Teichoic acids Cells deficient in the ability to D-alanylate teichoic acids have altered surface charge and demonstrate diminished ability to attach to polystyrene surfaces. Lipid S Similar to lipoteichoic acid, possibly involved in biofilm formation. Autolysins AtlE, a major autolysin, is thought to interact with hydrophobic surfaces and allow attachment of the bacteria to plastic materials. AccumulationLarge protein responsible for accumulative growth associated on polymer surfaces. protein PIA and PNSG Extracellular molecule composed of a major (I) and minor (II) polysaccharide; involved in cell-cell adhesion required for biofilm formation; also responsible for hemagglutination activity; PNSG is a polysaccharide similar to PIA. BAP A surface protein involved in biofilm formation. Hyaluronidase An enzyme that degrades hyaluronic acid found in lysosomes. Lipase An enzyme that catalyses the hydrolysis of fats (monoglycerides, diglycerides, and triglycerides) to glycerol and fatty acids; possibly important for skin colonization. Fatty acid modifying Works by esterifying fatty acids to cholesterol, enzyme thereby destroying their bactericidal properties. Staphylokinase Acts as a plasminogen activator (a serine protease that acts on plasminogen to generate plasmin). Has been implicated in invasiveness and is produced by many normal and invasive cells. Regulatory proteinsa Accessory gene A quorum-sensing system involved in regulating regulator (agr) expression of extracellular and surface-attached virulence factors; cell-density dependent. Secondary sigma Involved in regulation of surface-exposed factor (sigB) and sar virulence factors; may influence biofilm locus formation; precise roles are not known.

Organisms S. aureus S. aureus S. aureus, S. epidermidis

S. aureus

S. aureus

S. epidermidis S. epidermidis

S. epidermidis

S. aureus, S. epidermidis

S. aureus S. aureus S. aureus, S. epidermidis

S. epidermidis S. aureus

S. aureus

S. aureus

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COMPLICATED SKIN AND SOFT TISSUE INFECTIONS

TABLE 3. (continued) Virulence Factor Exotoxins Alpha-toxin

Beta-toxin Gamma-toxin

Epidermolytic toxins

Pyogenic toxins Enterotoxins Hemolytic peptide d-toxin

Description A hemolysin associated with skin necroses; can cause the release of cytokines and arachidonic acid derivatives involved in shock. A sphingomyelinase that damages membranes containing sphingomyelin (a lipid). Acts in conjunction with leukocidins to damage membranes of certain cells and may have importance in dermonecrotic lesions. Cause exfoliative disease of the skin (e.g., blisters, impetigo, scalded skin syndrome); are serine proteases. Associated with rash, toxic shock syndrome. Can induce food poisoning when ingested and shock when acquired parenterally. Involved in necrotizing enterocolitis in neonates; causes lysis of erythrocytes; forms part of polypeptide complex involved in modulating cytokines and NF-κB; also plays a role in biofilm formation.

Organisms S. aureus

S. aureus S. aureus

S. aureus

S. aureus S. aureus S. epidermidis

a Homologues of agr, sar, and sigB have been identified in the S. epidermidis genome but whether these loci

perform the same task as in S. aureus is unknown. BAP = Biofilm-associated protein. NF-κ B = Nuclear factor-κ B; a transcription factor that responds to inflammatory signals, activates the expression of inflammatory mediators, and plays a role in the cell proliferation process. PIA = Polysaccharide intercellular adhesion molecule. PNSG = Poly-N-succinyl-glucosamine.

thought to be particularly successful in persisting, disseminating, and causing infection (Day NPJ, 2001; Oliveira DC, 2002). The prevalence of MRSA continues to rise, with the National Nosocomial Infections Surveillance Survey (NNIS) recording resistance rates of 4% in the 1980s, more than 50% by the end of the 1990s, and 57.1% in 2002—a 13% increase over the 1997–2001 period (NNIS, 2003). Indeed, in 2000, researchers observed that the mean rate of MRSA isolates obtained from ICU patients increased by 40%, compared with the average rate of resistance over the previous Þve years. In Europe, 30–50% of Staph. aureus isolates obtained from hospital-based infections are resistant to methicillin. Coagulase-Negative Staphylococci. Coagulase-negative staphylococci (CoNS) are a major part of the normal bacterial ßora of the skin and mucous membranes. CoNS are not usually pyogenic and tend to cause infection only when introduced into the host or when provided with a channel to invade the host. Indeed, CoNS are most often associated with indwelling devices and infection is usually subacute or chronic. Staph. epidermidis is the most frequently isolated CoNS. Historically, this bacterium was regarded as relatively harmless;

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however, Staph. epidermidis is now recognized as a signiÞcant cause of nosocomial infections. Still, it is often difÞcult to establish the signiÞcance of CoNS isolated from a clinical specimen because of their universal colonization of the skin. The pathogenicity of Staph. epidermidis and other CoNS is essentially caused by the production of cell-surface-associated glycoproteins and proteoglycans. This “slime” allows the bacteria to adhere to surfaces and form bioÞlms on indwelling medical devices (Vuong C, 2002). Isolates associated with nosocomial infection appear to produce greater quantities of slime than do CoNS strains from other sources. The production of slime by Staph. epidermidis also reduces the ability of macrophages to phagocytose (engulf) and kill the bacteria (Heinzelmann M, 1997; Johnson GM, 1986; Shiau AL, 1998). The resistance patterns of CoNS are similar to those of Staph. aureus. In fact, CoNS isolates are thought to be the original source for the methicillin-resistance determinant, and researchers believe that resistance occurring in one species can be quickly transferred to the other. More than 80% of CoNS isolates from ICUs are resistant to methicillin. Streptococci. Streptococci are gram-positive, catalase-negative bacteria that include more than 30 different species. These species can be differentiated by a variety of diagnostic tests; the ability to lyse red blood cells separates streptococci into beta-hemolytic and alpha-hemolytic types. Most streptococci that are pathogenic in humans are beta-hemolytic. Streptococci can also be classiÞed by recognizable group antigens, which researchers have used to divide streptococci into groups A through H and K through V. Only group A, B, C, D, and G streptococci are normally found in humans; group A and B streptococci are the primary pathogens in cSSTIs. Group A streptococci are the etiological agents of well-known diseases like strep throat, scarlet fever, and necrotizing fasciitis. Streptococcus pyogenes is the most common streptococcal pathogen in cSSTIs, but other group A streptococci can cause these infections as well. Erysipelas is caused most often by group A streptococci, but group C or G can be the etiological agent in some cases. Although cellulitis can be caused by many types of bacteria, a signiÞcant number are caused by Strep. pyogenes. Streptococcal pathogenesis can be attributed to many factors, but streptococci produce a variety of toxins that are largely responsible. Streptococcal pyrogenic exotoxins types A through C cause the rash in scarlet fever but are also cytotoxic and pyrogenic. Streptolysin S forms pores in the membranes of mammalian cells (Ginsberg I, 1999). Streptolysin S and streptolysin O are both hemolysins and both can also bore holes in the membranes of leukocytes, platelets, and various subcellular organelles. Streptococci also encode several enzymes for cleaving various tissue proteins and substrates, probably to facilitate spread within tissue, including four DNA cleaving enzymes, an enzyme to cleave hyaluronic acid in connective tissue, streptokinase to break up clots, and other proteinases, amylases, and esterases (Mandell GL, 2000).

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In addition to causing pathogenesis at the site of infection, streptococci can cause systemic abnormalities and damage, termed toxic shock syndrome. Streptococcal toxic shock syndrome is much more common in cSSTIs than other streptococcal infections like strep throat. Toxic shock syndrome is thought to be largely the result of the activity of streptococcal pyrogenic exotoxins types A and C, but several other toxins are also implicated. These toxins are superantigens that are capable of activating T cells to release large amounts of cytokines, inducing tumor necrosis factor, which can lead to a cascade of systemic inßammation, coagulation, and sepsis (Baker MD, 2004). Pseudomonads. Pseudomonads are ubiquitous gram-negative bacteria that are found in soil and water supplies. These organisms require simple growth conditions, are capable of multiplying in moist areas (including distilled water), can survive in disinfectant solutions, and are often resistant to antibiotics. These factors contribute to the success of the pseudomonads as opportunistic pathogens, rating among the top pathogens identiÞed in hospital-based infections. In cSSTIs, pseudomonads are suspected primarily in infected diabetic ulcers. Among the pseudomonads, Pseudomonas aeruginosa is the most frequently isolated. Normally a soil bacterium, P. aeruginosa enjoys moist environments, leading to its colonization of the perineum, ear, and axillae. This preference also explains its establishment within the hospital environment in sites such as ice machines, pharmacy preparations, plaster, mouthwash, nebulizers, whirlpools, mattresses, sinks, and potted plants. P. aeruginosa is a relatively uncommon component of skin ßora for healthy individuals, but it has become a leading pathogen among hospitalized patients since the modern era of intensive treatments and broad-spectrum antibiotic use. This opportunistic pathogen is capable of colonizing the skin and mucous membranes of otherwise healthy people, who then can carry this bacterium in the bowel. Antibiotic therapy usually encourages colonization, which often precedes invasive infection. P. aeruginosa characteristically causes pneumonia in ICU patients and neonates and frequently infects abdominal wounds after surgery or trauma. Serious infection typically occurs only in immunocompromised people (e.g., those with burns, cancer, AIDS). The pathogenesis of Pseudomonas spp. is poorly understood. The complex relationship among factors that promote local, invasive, and systemic disease may be divided into three distinct stages: (1) bacterial attachment and colonization, (2) local invasion, and (3) dissemination and systemic disease. Each stage requires the previous stages to occur, but for complex and unknown reasons the disease process may stop at any stage (Lyczak JB, 2000; Pollack M, 1984; Stanislavsky ES, 1997). Table 4 summarizes select virulence factors of P. aeruginosa that may be targets of future drug development or diagnostic markers of infection by this pathogen. Virulence factors involved in bacterial attachment and colonization include the mucoid exopolysaccharide of mucoid Pseudomonas strains and the pili (adhesins) of nonmucoid strains (Alverdy J, 2000; Comolli JC, 1999; Hahn HP, 1997;

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TABLE 4. Select Virulence Factors of Pseudomonas aeruginosa Virulence Factor Surface-associated proteins Mucoid exopolysaccharide

Flagella/pili

Extracellular proteases Elastase and alkaline protease

Hemolysins Phospholipase C and rhamnolipid

Toxins Cytotoxin (formerly leukocidin)

Exoenzyme S and other effector proteins of the type III secretory system Exotoxin A

Description Under certain conditions, P. aeruginosa produces a mucoid substance (alginate) that functions similar to a polysaccharide capsule. This exopolysaccharide forms a matrix around the bacterium (similar to a biofilm), anchoring it to its environment and protecting it from host immune factors such as the mucociliary mechanism of the respiratory tract, phagocytic cells, antibodies, and complement. Flagella are filamentous immunogenic organelles that mediate motility and are associated with increased virulence. Pili are involved in binding the bacteria to mucosal surfaces. The tip of the pilus that binds to cells is termed the adhesin. Cause tissue destruction and cell death in the skin, lungs, and cornea via their proteolytic activity on elastin (important in tissue supportive structures) and fibrin (involved in blood clotting). The breakdown of host tissue is thought to provide the organism with nutrients. Elastase may also cause intra-alveolar hemorrhage and degradation of laminin and elastin in vascular tissue. Capable of hydrolyzing many proteins involved in host defense mechanisms (e.g., IgA, IgG, TNF-α, IFN-γ ), enabling the bacteria to persist at the site of infection. Act synergistically to break down lipids and lecithin (important components of cell membrane structure). Phospholipase C is also responsible for the degradation of an important component of lung surfactant and may contribute to the inflammation associated with P. aeruginosa infection by augmenting arachidonic acid synthesis. Cytotoxic for most eukaryotic cells; is thought to contribute to sepsis-associated lung injury and the development of acute respiratory distress syndrome. Displayed on the bacterial cell surface, these toxins appear to be involved in promoting bacterial adhesion as well as contributing to bacterial dissemination and persistence by inhibiting wound healing. Kills mammalian cells by gaining entry to the cytosol and inhibiting mammalian protein biosynthesis (via covalent linkage with EF-2, an enzyme that catalyzes the elongation step in polypeptide [protein] assembly). Because this extracellular protein mediates tissue necrosis in both local and systemic sites, strains of P. aeruginosa-expressing exotoxin A are more virulent than nontoxigenic strains.

EF-2 = Elongation factor 2; IFN-γ = Interferon gamma; IgA = Immunoglobulin A; IgG = Immunoglobulin G; LPS = Lipopolysaccharide; TNF-α = Tumor necrosis factor-alpha.

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Ichikawa JK, 2000). Mucoid strains of P. aeruginosa are most often isolated from the sputum of patients with cystic Þbrosis and those with injured respiratory epithelia (de Bentzmann S, 1996[a]; de Bentzmann S, 1996[b]). Other virulence factors of P. aeruginosa have cytotoxic effects on host tissues that are involved in the ability of this pathogen to proliferate and invade localized areas (Barbieri JT, 2000; Frank D, 1997; Seeger W, 1987; Suter S, 1994; Vasil ML, 1991). Many pseudomonads contain a permeability barrier that prevents accumulation of antibiotics inside the cell, rendering the bacteria intrinsically resistant to many drug classes. Virtually all P. aeruginosa isolates also contain an inducible, chromosomally mediated β-lactamase enzyme that is capable of inactivating many cephalosporins. In addition, an increasing number of isolates contain a plasmidmediated β-lactamase that confers resistance to ampicillin and other penicillins, notably the extended-spectrum (antipseudomonal) penicillins (Lister PD, 2000). P. aeruginosa isolates that are resistant to imipenem, quinolones, and ceftazidime have been identiÞed with increasing frequency. The rapid rise in isolates resistant to quinolones from 5% in 1989 to more than 15% by 1999 has been impressive in magnitude and is becoming an important clinical issue. Enterobacteriaceae. The enterobacteriaceae are a large family of gramnegative bacteria that inhabit the large intestine of mammals. Most are harmless commensals, but others are associated with opportunistic infections (e.g., Klebsiella, Enterobacter) or intestinal disease (e.g., Salmonella, Shigella). Still others are highly versatile (e.g., Escherichia coli ), with both pathogenic and opportunistic strains associated with various diseases. Most cSSTIs resulting from enterobacteriaceae are perianal abscesses, resulting from anal Þssures. Recurring cSSTIs in this area can indicate inßammatory bowel disease. E. coli is probably the most frequently isolated gram-negative bacteria in nosocomial infections. The diversity of diseases caused by E. coli is due to the acquisition of speciÞc virulence factors that are harbored on transmissible genetic elements (e.g., plasmids) or within distinct DNA segments called pathogenicity islands that are absent from nonpathogenic strains. As a result, many different strains of E. coli exist, and some are more virulent than others. Klebsiella spp., such as K. pneumoniae and K. oxytoca, have come into prominence in recent years because of outbreaks associated with multidrug-resistant strains. These endogenous bacteria are often transmitted via indirect contact with feces. CURRENT THERAPIES Overview Most cases of complicated skin and soft-tissue infection (cSSTI) require empiric antibiotic therapy because numerous bacterial species can cause infections and because diagnosing the speciÞc pathogen takes some time. Practitioners choose empiric therapy based on likely infecting pathogens, local resistance patterns, the location and depth of the infection, and the patient’s history. Table 5 summarizes the leading antibiotic therapies available to treat cSSTIs. Physicians use other

CURRENT THERAPIES

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TABLE 5. Current Therapies Used for Complicated Skin and Soft Tissue Infections, 2004 Agent

Company/Brand

Daily Dose

Penicillins Benzylpenicillin (penicillin G)

Pfizer’s Pfizerpen, Wyeth’s Wycellin, generics

Flucloxacillin

GSK’s Floxapen/Staphlex, generics

Ampicillin/ sulbactam Amoxicillin/ clavulanate

Pfizer’s Unasyn/Unacim/Unacid, generics GSK’s Augmentin/Augmentin XR/Augmentan, generics

6 g (10 MU) daily in divided doses q6h 500 mg qid (PO); 1,000 mg q6h (IV) 3 g q6h

Ticarcillin/ clavulanate Piperacillin/ tazobactam Carbapenems Imipenem/ cilastatin Meropenem

GSK’s Timentin/Claventin Augpenin

1.5–1.75 g daily in daily doses (PO); 1.2 g q8h (IV) 3.1 g q4h

Wyeth/Taisho’s Zosyn/Tazocin

4.5 mg q8h

US, F, G, I, S, UK, J

Merck’s Primaxin/Zienam/Tienam

500 mg q6h

AstraZeneca/Sumitomo’s Merrem IV/Meronem/Meropen Merck’s Invanz

1 g q8h

US, F, G, I, S, UK, J US, F, G, I, S, UK, J US, G, S, UK

GSK’s Ceftin/Zinnat/Oracef, generics BMS’s Cefzil/Procef/Bisoral Pfizer’s Vantin, Sanofi-Aventis’s Orelox, Sankyo’s Banan Abbott/Sanofi-Aventis’ Claforan, generics Roche’s Rocephin/Rocefin/Rocefalin, generics

500 mg qid

Ertapenem Cephalosporins Cefuroxime axetil Cefprozil Cefpodoxime proxetil Cefotaxime Ceftriaxone

Lincosamides Clindamycin

Pfizer’s Cleocin/Dalacin/Sobelin, generics

Fluoroquinolones Ciprofloxacin Bayer’s Cipro/Cipro IV/Cipro XR/Ciflox/Ciprobay/Ciproxin/Baycip/Ciproxan, generics Aminoglycosides Gentamicin Schering-Plough’s Garamycin/Gentacin, Sanofi-Aventis’ Cidomycin, Merck’s Septopal, generics Amikacin Novartis’s Amikin, BMS’ Amiklin/Biklin/Amikin, generics

1 g qd

500 mg bid 400 mg bid 1 g q6h 1 g q12h

Availability US, F, G, I, S, UK, J G, I, UK US, F, G, I, S, J US, F, G, I, S, UK, J US, F, I, UK

US, F, G, I, S, UK, J US, I, S US, F, G, I, S, UK, J US, F, G, I, S, UK, J US, F, G, I, S, UK, J

300 mg tid (PO); 600 mg q8h (IV)

US, F, G, I, S, UK, J

500 mg bid (PO); 400 mg q12h (IV)

US, F, G, I, S, UK, J

3 mg/kg daily in divided doses q8h

US, F, G, I, S, UK, J

15 mg/kg daily in divided doses q12h

US, F, G, I, S, UK, J

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COMPLICATED SKIN AND SOFT TISSUE INFECTIONS

TABLE 5. (continued) Agent

Company/Brand

Glycopeptides and Lipopeptides Vancomycin Lilly’s Vancocin, generics Teicoplanin

Sanofi-Aventis’s Targocid

Daptomycin Cubist’s Cubicin Nitroimidazoles Metronidazole Sanofi-Aventis/SCS Pharmaceuticals/Shionogi’s Flagyl/Flagyl IV/Metro IV, generics Oxazolidinones Linezolid Pfizer’s Zyvox/Zyvoxid Streptogramin combinations Quinupristin/ King Pharmaceuticals/Sanofidalfopristin Aventis/Fujisawa’s Synercid

Daily Dose 1 g q12h 400 mg loading, then 200–400 mg q24h 4 mg/kg q24h

Availability US, F, G, I, S, UK, J F, G, I, S, UK, J US

500 mg qid (PO); 500 mg q6h (IV)

US, F, G, I, S, UK

600 mg bid (PO); 600 mg q12h (IV)

US, F, G, I, S, UK, J

7.5 mg/kg q12h

US, F, G, I, S, UK, J

Oral (PO) dosing: qid = four times daily; tid = three times daily; bid = twice daily; qd = once daily; Intravenous dosing: q4h = every 4 hours; q6h = every 6 hours; q8h = every 8 hours; q12h = every 12 hours; q24d = every 24 hours; MU = million units (mega-units). US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan.

drug classes (e.g., tetracyclines) very commonly in community-based treatment of mild SSTIs (e.g., acne, rosacea, dermatitis), but these infections are beyond the scope of this report because of their limited use in cSSTIs. Although numerous antibiotics are effective in treating cSSTIs, only a few agents have clinical trial data speciÞc to these complicated infections. Physicians frequently employ antibiotics in the treatment of cSSTIs based on results from trials of uncomplicated SSTI or bacteriological data demonstrating efÞcacy against common cSSTI pathogens. Most clinical trials comparing the efÞcacy of antibacterials in SSTIs suggest that the major antibiotic classes and their individual compounds have very similar efÞcacy rates with respect to their ability to elicit a prompt response and to resolve infection. Clinical trials generally measure clinical response as improvement in symptoms (e.g., reduction in fever) and bacteriological end points as a rate of pathogen eradication. Studies report the susceptibility of bacterial strains to antibiotics using the terms sensitive, intermediately sensitive/resistant, or resistant, based on the minimum inhibitory concentration (MIC) required for the antibacterial to exert its effect. Choice of Antibiotic. Physicians consider many factors when they choose an antibiotic to treat cSSTIs: oral bioavailability, tissue penetration, spectrum of activity, and resistance patterns. Oral bioavailability is the amount of drug that is absorbed from the gastrointestinal tract into the bloodstream. For serious infections, physicians need to ensure that patients achieve a high serum concentration of the drug at the site of infection. Antibiotics with poor oral bioavailability do

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not achieve high enough serum concentrations, so physicians must instead use an intravenous (IV) formulation. Antibiotics with high oral bioavailability, however, offer physicians the ability to treat serious infections more conveniently in an outpatient setting. Physicians can also use these oral antibiotics as a step-down agent for patients who initially receive IV antibiotics in the hospital, which results in shorter hospital stays. Because cSSTIs tend to extend deeper into tissue than uncomplicated infections, physicians choose an antibiotic based on its ability to penetrate tissue. Tissue penetration is most important in the management of infected diabetic ulcers, which can extend as deep as the bone. However, any type of cSSTI can become invasive in select cases. Physicians address this problem in different ways; some choose an antibiotic such as clindamycin or ciproßoxacin, agents known for their superior tissue penetration, while others stick with their preferred antibiotic and add a second drug such as rifampicin, which offers penetration to deeper tissues. Physicians’ choice of antibiotic is also inßuenced by the breadth of pathogens suspected at the infection site. Some antibiotics have a narrow spectrum of activity (e.g., vancomycin), while others have a broad spectrum that covers a wide variety of susceptible pathogens (e.g., carbapenems, β-lactam/β-lactamaseinhibitor combinations). Physicians prefer to use a narrow-spectrum antibiotic if possible, but many cSSTI infections are due to multiple, unidentiÞed pathogens, and in these cases physicians must ensure that the chosen antibiotic has broad enough coverage. Physicians also consider an antibiotic’s ease of use in a hospital setting and appropriateness and convenience for patients with comorbidities (e.g., diabetes). Another factor that inßuences a physician’s choice of drug is antibiotic resistance, as discussed in the following section. Antibiotic Resistance. During the past 10–15 years, there has been a steady rise in the prevalence of antibiotic-resistant pathogens. This is especially true in cSSTIs, where the dominant pathogen is Staphylococcus aureus. Many researchers have documented the troubling rise of methicillin-resistant Staph. aureus (MRSA) in the hospital setting. MRSA is also on the rise in the community and is quickly becoming a concern for SSTI cases that do not respond to the initial antibiotic treatment prescribed by general practitioners. This rise in resistance has limited the use of older antibacterials such as penicillin and early-generation cephalosporins and has spurred the use of more powerful agents such as vancomycin and newer therapies such as the ßuoroquinolones. Unfortunately, microbiologists have now isolated strains of vancomycin-resistant bacteria in several patients, demonstrating that bacterial drug resistance is an inherent and continuing challenge in the clinical management of cSSTIs and is likely to continue to drive innovation and the development of novel agents. Pathogen resistance to an antibiotic reduces the efÞcacy of the agent and occurs most often in patients who have recently received antibiotic therapy (as can be the case in hospitalized, immunocompromised, or diabetic patients). From a public health perspective, antibacterial resistance in the community can occur

588

COMPLICATED SKIN AND SOFT TISSUE INFECTIONS

after widespread use (or overuse) of a given antibacterial over a period of time. Therefore, the efÞcacy of a given antibacterial and the susceptibility of pathogens in the community may change over time as resistance patterns change. Furthermore, the prevalence of resistance varies geographically and may correlate with a variety of community factors including local prescription volume. In recent years, antibiotic resistance has become an important factor inßuencing physicians’ choice of agent, even at local levels (i.e., resistant pathogen prevalence varies from hospital to hospital, even within the same metropolitan area). Bacteria have proven remarkably adept at developing resistance to new antimicrobial agents. Resistance to an antibiotic may be an intrinsic property of a bacterial species, or it may be an acquired characteristic of an individual organism. Once bacteria do become resistant, they can (and usually do) pass this ability from one generation to the next. They may also pass on resistance to other strains. Table 6 presents the mechanisms by which bacteria become resistant to the various classes of available antibacterials. Alterations in or mutations of the bacterial chromosomes were the Þrst identiÞed mechanisms of resistance. Although chromosome mutation can be the transformative process, pathogens acquire antibacterial resistance more frequently through transfer of genetic material from one bacterium to another. One of the most worrisome mechanisms of resistance involves the acquired ability of bacteria to exclude antimicrobial agents from the cell. Some gram-positive and gram-negative pathogens have an efßux pump that can remove drugs fast enough to protect the bacteria from the drug and its effects. Resistance to an antibiotic may also result from a change or alteration in the target site at which the drug acts. Some strains of bacteria alter the protein composition of their outer cell membrane to prevent a drug from binding to its target site. Other bacteria resist antibiotics by acquiring mutations that allow them to bypass a metabolic step or pathway that is critical to the drug’s mechanism of action. Other resistant bacteria produce enzymes that cleave or alter the molecular structure of an antibiotic, thereby rendering it ineffective. For example, β-lactamase is an enzyme produced by bacteria that destroys the ring structure of β-lactam antibiotics (e.g., penicillins) responsible for its antimicrobial effect. Penicillins Overview. Penicillins are members of the β-lactam family of antibiotics, which contain a basic β-lactam structure fused to a Þve-member ring. Because of their broad spectrum of activity, safety, and convenient dosing, physicians commonly use penicillins empirically in the treatment of a range of bacterial infections. The penicillins are among the least toxic classes of antibiotics, although an immediate or delayed hypersensitivity reaction is a potentially life-threatening adverse effect. Hypersensitivity reactions can present in a range of severities, from mild skin rash, interstitial nephritis, hematological disturbances (e.g., neutropenia), and vasculitis to anaphylactic shock. The overall incidence of hypersensitivity reactions ranges between 0.7% and 10% of patients treated with penicillins (Weiss

CURRENT THERAPIES

589

TABLE 6. Resistance Mechanisms to Current Therapies Used for Complicated Skin and Soft Tissue Infections Drug Class Penicillins, carbapenems, and cephalosporins

Lincosamides

Fluoroquinolones

Tetracyclines

Aminoglycosides

Glycopeptides and lipopeptides Nitroimidazoles

Oxazolidanones

Streptogramin combinations

Mechanism Expression of β-lactamases, which hydrolyze the β-lactam ring of these antibiotics, has been found in both gram-negative and gram-positive bacteria, with at least 340 β-lactamases described thus far. Sophisticated restructuring of the penicillin-binding proteins (PBPs) is commonly seen in gram-positive cocci. Modifications in outer-membrane permeability are frequently observed in gram-negative species. (Pseudomonas aeruginosa is resistant to many antibacterial drugs because it lacks porins [protein channels] necessary for drug access through the outer membrane.) Alterations in the ribosomal binding site (MLSB phenotype) are the main mechanisms of resistance to the macrolides and lincosamides. Alterations in antibiotic transport (overexpression of efflux pumps) to remove antibiotics from bacterial cell. Production of enzymes that inactivate the antibiotic (e.g., hydrolysis by esterases). Mutations in genes coding for DNA gyrase and/or DNA topoisomerase IV. Overexpression of efflux pumps. Resistance to the tetracyclines can develop rapidly via the selection of isolates that lack permeability to the drug. Changes in the bacterial cell membrane that prevent accumulation of the drug within the cell (efflux). Production of enzymes that protect the ribosome complex from antibiotic binding. Alteration or deletion of antibiotic binding sites on the 30S ribosome subunit. Production of enzymes that inactivate the aminoglycoside. Modification of the D-alanyl-D-alanine components of the cell wall into D-alanyl-D-lactate components inhibits binding of vancomycin. Inactivation of the nitroreductase gene rdxA in Helicobacter pylori Inactivation by products of the nim gene locus in Bacteriodes Possible alterations in flavodoxins or hydrogenase in Clostridia Resistance due to a 23S rRNA mutation can emerge in enterococci during therapy with oxazolidinones and may be associated with clinical failure. Organisms with the MLSB -resistance phenotype are resistant to quinupristin (streptogramin B) but remain susceptible to dalfopristin (streptogramin A).

MLSB = Macrolide-lincosamide-streptogramin B

590

COMPLICATED SKIN AND SOFT TISSUE INFECTIONS

H

COOK

O

CH3

N CH2CONH

S H

CH3

H

FIGURE 3. Structure of benzylpenicillin potassium.

ME, 1988). Other more common—yet minor—side effects caused by penicillins are gastrointestinal (GI) disturbances (e.g., nausea, vomiting, diarrhea). A growing concern over the past two decades has been the development of bacterial resistance to β-lactam antibiotics. β-lactam drug resistance results from altered penicillin-binding proteins (PBPs) in the cell wall of resistant organisms that reduce the afÞnity of the antibacterial agents to the target sites to which penicillins must bind to exert their inhibitory effects. Bacterial resistance to commonly used β-lactam antibiotics has caused physicians to choose newer generations of drugs with better activity against speciÞc resistant strains of pathogens. Mechanism of Action. Penicillins bind to PBPs in bacteria and prevent bacterial cell-wall formation. PBPs are transpeptidase enzymes that cross-link peptidoglycans in the bacterial cell wall and serve to maintain cell-wall homeostasis (e.g., structural integrity, cell shape, cell division, capsule synthesis, phage resistance, regulation of autolysis). The β-lactam ring of penicillins interacts with PBPs and inhibits cell-wall synthesis, killing the cell. Because most bacteria contain between four and eight different PBPs, effective antibiotics need to target several different proteins. Organisms that are metabolically inactive and those lacking bacterial cell walls (e.g., Mycoplasma) are not susceptible to β-lactam antibiotics. Benzylpenicillin (Penicillin G). Benzylpenicillin (PÞzer’s PÞzerpen, Wyeth’s Wycellin, generics) (Figure 3), or penicillin G, is a parenteral antibiotic available in major markets since the 1970s. Benzylpenicillin is a very old product and while it still has sales, there are many generics and the brands are not promoted any more. Like other penicillins, benzylpenicillin exerts a bactericidal effect against susceptible organisms by binding to PBPs, undermining the stability of cell walls and blocking their formation. Used for severe infections when physicians need rapid and high penicillin levels, benzylpenicillin is active against many of the streptococcal, staphylococcal, and atypical pathogens that commonly cause various cSSTIs. Because this agent has been available for almost thirty years, there are no recent, rigorous clinical studies of benzylpenicillin in the treatment of SSTIs. Rather, evidence of its efÞcacy is apparent from its widespread successful clinical use in a variety of bacterial infections including cSSTIs. Like the other penicillins, benzylpenicillin has low toxicity but does have a signiÞcant risk of allergic reaction in patients with hypersensitivity. A hypersensitive reaction to benzylpenicillin may include skin rashes, urticaria, chills, or

CURRENT THERAPIES

H

F

COOH

O

CH3

N CONH N Cl

S H

O

591

CH3

H

CH3

FIGURE 4. Structure of flucloxacillin.

fevers. As with other parenterally administered penicillins, patients treated with benzylpenicillin may have local irritation at the site of administration. Flucloxacillin. Flucloxacillin (GlaxoSmithKline’s Floxapen/Staphlex, generics) (Figure 4) has been marketed since 1970 but is only available in Germany, Italy, United Kingdom, and Japan today. Like benzylpenicillin, ßucloxacillin is a very old product. Although it still has sales, there are many generics, and the brands are no longer promoted. The drug is available in both oral and parenteral formulations. Patients with cellulitis or impetigo often receive oral ßucloxacillin in an outpatient setting. Flucloxacillin is a narrow-spectrum isoxazolyl penicillin that interferes with the ability of bacteria to form cell walls by binding with PBPs. The agent shows some resistance to degradation by β-lactamases; hence, physicians use ßucloxacillin when they identify or suspect a penicillin-resistant infection. Flucloxacillin is active against many community-acquired gram-positive staphylococcal and streptococcal infections (particularly Staphylococcus aureus and Streptococcus pyogenes). An early, double-blind, dose-comparative clinical trial evaluated the efÞcacy of ßucloxacillin in the treatment of staphylococci- and streptococci-induced SSTIs. The study enrolled 223 patients and randomly assigned them to therapy with either 250 mg or 125 mg of ßucloxacillin four times daily for Þve days. There were 75 and 64 patients in each group, respectively, who were clinically evaluable. In the group that received the 250 mg dose, 97.3% of patients experienced clinical cure, as did 92.1% of patients who received the 125 mg dose (Harding JW, 1970). Flucloxacillin is contraindicated in patients who have a history of hypersensitivity to penicillins and in patients with a history of hepatic impairment. Side effects of ßucloxacillin include diarrhea, nausea, vomiting, and jaundice. Although ßucloxacillin is often used in staphylococcal skin infections, it is not active against methicillin-resistant Staph. aureus (MRSA). Ampicillin/Sulbactam. Ampicillin/sulbactam (PÞzer’s Unasyn Unacim/Unacid, generics) is a combination of the semisynthetic aminopenicillin ampicillin (Figure 5) and the β-lactamase inhibitor sulbactam (Figure 6). This combination reached most major markets by 1987. Ampicillin/sulbactam is available for parenteral administration; physicians rarely use the oral formulation of this combination (known as sultamicillin) for cSSTIs.

592

COMPLICATED SKIN AND SOFT TISSUE INFECTIONS

H H C

COOH

O

CH3

N CONH

S H

NH2

CH3

H

FIGURE 5. Structure of ampicillin.

H

COONa

O

CH3

N

H

S O2

CH3

FIGURE 6. Structure of sulbactam sodium.

Ampicillin achieves its bactericidal effect by inhibiting cell-wall formation. As with other β-lactam/β-lactamase-inhibitor combination therapies, the β-lactamase produced by the bacteria irreversibly binds to the β-lactamase inhibitor with greater afÞnity than it binds to the active β-lactam antibiotic. This protects the antibiotic from degradation and effectively extends its spectrum of activity to include several gram-negative organisms in addition to gram-positive staphylococci. According to the package insert for PÞzer’s Unasyn, a controlled clinical trial conducted in pediatric patients demonstrated comparable therapeutic efÞcacy for intravenous (IV) ampicillin/sulbactam and IV cefuroxime in the treatment of SSTIs. Of 99 total patients, 60 received ampicillin/sulbactam therapy and 39 cefuroxime therapy. The rates of clinical success for the ampicillin/sulbactam and cefuroxime groups were 85% and 87%, respectively. The recommended pediatric dosage of ampicillin/sulbactam is 300 mg/kg/day administered every 6 hours. All patients received at least 72 hours of IV therapy. As with other penicillin-related therapies, ampicillin/sulbactam is contraindicated for patients with a history of hypersensitivity to penicillins. As described in the package insert, a high percentage of patients who receive ampicillin develop a hypersensitive skin rash, and some have local adverse reactions such as pain at the intramuscular (IM) injection site (16%), pain at the IV injection site (3%), and thrombophlebitis (3%). As with other penicillins, patients report gastrointestinal (GI) disturbances such as diarrhea (3%), rash (2%), and additional reactions (e.g., itching, nausea, vomiting). Amoxicillin/Clavulanate. Amoxicillin/clavulanate (GlaxoSmithKline’s Augmentin, Augmentin XR, Augmentan, generics) has a broad spectrum of activity against many gram-positive and gram-negative microorganisms. The combination is a favorite among clinicians because of its efÞcacy, safety, and familiarity.

CURRENT THERAPIES

593

H COOH O H HO

C

CH3

N CONH

NH2

CH3

S H

3 H2O

H

FIGURE 7. Structure of amoxicillin.

H S

O

H C

COONa CH3

N CONH

COONa

S H

CH3

H

FIGURE 8. Structure of ticarcillin disodium.

Amoxicillin (Figure 7) acts by disrupting bacterial cell-wall synthesis. The addition of the β-lactamase inhibitor clavulanate protects amoxicillin from degradation by β-lactamase enzymes produced by bacteria; this effectively extends the antibiotic spectrum to include many pathogens otherwise resistant to amoxicillin. Amoxicillin/clavulanate is available in tablets, chewable tablets, extra-strength and extended-release tablets, and oral suspension formulations. An IV form is available in Europe for parenteral administration. Because this agent has been available for many years, there are no recent, rigorous clinical studies of amoxicillin/clavulanate in the treatment of SSTIs. Rather, its widespread successful clinical use in a variety of bacterial infections including cSSTIs is evidence of its efÞcacy. The combination therapy is generally well tolerated, and, according to the Augmentin package insert, the most frequent adverse effects include diarrhea/loose stools (9%), nausea (3%), skin rashes and urticaria (3%), vomiting (1%), and vaginitis (1%). Ticarcillin/Clavulanate. Ticarcillin/clavulanate (GlaxoSmithKline’s Timentin/ Claventin/Augpenin) is an extended-spectrum β-lactam/β-lactamase-inhibitor combination that pairs the semisynthetic carboxypenicillin ticarcillin (Figure 8) with the β-lactamase-inhibitor clavulanate. Available in the United States since 1985, ticarcillin/clavulanate is marketed by GSK in all major markets except Germany and Spain and is used for more severe infections than is the amoxicillin/clavulanate combination. This combination therapy is available only in a parenteral formulation. As with other β-lactam/β-lactamase-inhibitor combinations, the β-lactamase enzyme produced by bacteria irreversibly binds to the β-lactamase-inhibitor (clavulanate) with greater afÞnity than it binds to the active β-lactam-antibiotic (ticarcillin); this protects the antibiotic from degradation and effectively extends its spectrum of activity to those pathogens resistant to ticarcillin alone. Ticarcillin/clavulanate

594

COMPLICATED SKIN AND SOFT TISSUE INFECTIONS

H COONa O H

CH3

N

C

CONH

NHCO

S H

N

O

N

O

CH3

H

C2H5 FIGURE 9. Structure of piperacillin.

has a broad spectrum of activity that covers gram-positive staphylococci, many gram-negative species (including Escherichia coli ), and a variety of anaerobic and atypical organisms. A double-blind, randomized, comparative, multicenter clinical trial of 251 patients evaluated the efÞcacy of ticarcillin/clavulanate and piperacillin/tazobactam in the treatment of patients with cSSTIs requiring hospitalization. In the study, efÞcacy (deÞned as clinical cure or improvement) was comparable for the two treatment regimens. Patients received treatment of either 3.1 g ticarcillin/clavulanate or 3.375 g piperacillin/tazobactam four times daily for at least Þve days. Of the 44 patients who received ticarcillin/clavulanate, 77% achieved clinical cure or improvement, as did 76% of the 67 patients who received piperacillin/tazobactam (Tan JS, 1993). Although it has the low toxicity characteristic of penicillins, ticarcillin/clavulanate is contraindicated for patients with a history of hypersensitivity to penicillins. Ticarcillin/clavulanate shares many of the same occasional side effects of other penicillins, including skin rash, pruritis, GI disturbances, and local reactions. Adverse reactions are similar in adult and pediatric populations. Piperacillin/Tazobactam. Piperacillin/tazobactam (Wyeth/Taisho’s Zosyn Tazocin) combines the extended-spectrum semisynthetic antibiotic piperacillin (Figure 9) with the β-lactamase inhibitor tazobactam (Figure 10). Piperacillin/ tazobactam, marketed since 1993, is available in all major markets in parenteral form only and is used for more severe infections than amoxicillin/clavulanate. Like other β-lactam therapies, piperacillin achieves its bactericidal effect by interacting with PBPs and disrupting cell-wall formation. Because this is a βlactam/β-lactamase-inhibitor combination therapy, the β-lactamase produced by bacteria are bound to the tazobactam more than they are bound to the piperacillin; this allows piperacillin to cover a broader range of organisms, including many gram-negative organisms, such as Pseudomonas aeruginosa (Lister PD, 2000). As noted earlier, a double-blind, randomized clinical trial of 251 patients evaluated the efÞcacy of ticarcillin/clavulanate and piperacillin/tazobactam in the treatment of cSSTI patients requiring hospitalization. Of the 44 patients

CURRENT THERAPIES

595

FIGURE 10. Structure of tazobactam (R = R1 = H).

who received 3.1 g ticarcillin/clavulanate for Þve days, 77% achieved clinical cure or improvement, as did 76% of the 67 patients who received 3.375 g piperacillin/tazobactam (Tan JS, 1993). During clinical trials of piperacillin/tazobactam, most of the adverse reactions were transient and mild to moderate in severity. According to the package insert for Zosyn, the most signiÞcant side effects reported include skin rashes/pruritus, GI complications (including diarrhea, nausea, and vomiting), and allergic reactions (in 1.3%, 0.9%, and 0.5% of the trial population, respectively). Carbapenems Overview. Carbapenems are penicillin derivatives that have good activity against gram-positive and gram-negative aerobic and anaerobic bacteria, and are available in parenteral form only. They include imipenem/cilastatin (Merck’s Primaxin), meropenem (AstraZeneca’s Merrem), and ertapenem (Merck’s Invanz). Resistance is not a major concern for carbapenem use, but physicians use them infrequently for cSSTIs because they are expensive and many consider their spectrum of activity too broad. Researchers have associated the use of carbapenems with outgrowth of fungal infections because the normal bacterial ßora of the skin is destroyed by the antibiotic (similar to other broad-spectrum agents). Mechanism of Action. Carbapenems have broad activity against a wide range of gram-positive and gram-negative aerobic and anaerobic bacteria. They usually exert a bactericidal effect; however, they are bacteriostatic against Enterococcus faecium and some Staph. aureus strains. As with other β-lactams, carbapenems bind to and inactivate the PBPs that cross-link peptidoglycans in the bacterial cell wall. Inhibition of this process destroys the organization and membrane potential of the bacterial cell wall. Imipenem/Cilastatin. Imipenem/cilastatin (Merck’s Primaxin/Zienam Tienam) is a carbapenem/renal dipeptidase inhibitor combination. Merck Þrst introduced this broad-spectrum combination therapy as an IV agent in 1985 and followed with an IM formulation in 1991; the latter is not intended for severe or life-threatening infections. Because imipenem is susceptible to hydrolysis by the renal enzyme dehydropeptidase-1 (DHP-1), it is administered with the DHP-1 inhibitor cilastatin (Figure 11)

596

COMPLICATED SKIN AND SOFT TISSUE INFECTIONS

FIGURE 11. Structure of cilastatin.

to aid pharmacokinetics and improve the urinary recovery of imipenem. Imipenem is stable against the β-lactamases produced by both gram-positive and gramnegative bacteria, and it can inhibit the β-lactamases of gram-negative organisms that are resistant to most penicillins and cephalosporins (such as Pseudomonas aeruginosa). As such, its broad spectrum of activity includes streptococci and staphylococci (especially Strep. pyogenes, Staph. aureus, and Staph. epidermidis), gram-negative microbes (especially E. coli and P. aeruginosa), and anaerobes. Imipenem/cilastatin is indicated for cSSTIs caused by Staph. aureus (penicillinase-producing strains), Strep. pyogenes, E. coli, and P. aeruginosa. In a noncomparative clinical trial, 243 patients with moderately severe to severe softtissue infections received imipenem/cilastatin. Overall, 95% of the 243 patients achieved clinical cure or improvement. The researchers also reported that 98% (498/506) of the various bacterial strains isolated from the patients were eradicated (Marier RL, 1983). Imipenem can trigger hypersensitivity reactions in individuals who are allergic to β-lactams, and physicians must monitor patients closely for symptoms of anaphylaxis. Adverse CNS effects (such as seizures and confusional states) have also occurred during imipenem treatment, suggesting that imipenem is contraindicated for those with a history of convulsive CNS disturbances. According to Primaxin’s package insert, other side effects noted in clinical trials include adverse local reactions (e.g., thrombophlebitis, pain, erythema) and systemic reactions (e.g., nausea, diarrhea, vomiting, rash). Although imipenem/cilastatin has an extensive spectrum of activity, researchers believe that methicillin-resistant organisms and some Pseudomonas species are resistant to the combination. Meropenem. Meropenem (AstraZeneca’s Merrem IV/Meronem/Meropen) (Figure 12) is a synthetic, broad-spectrum, parenteral carbapenem Þrst launched by AstraZeneca in the United States in 1996. AstraZeneca markets meropenem in Europe, while its partner Sumitomo distributes meropenem in Japan under the name Meropen. Meropenem exerts antimicrobial activity by penetrating the cell walls of gram-positive and gram-negative bacteria and binding to PBPs, thereby inhibiting cell-wall synthesis. Its clinically proven spectrum of activity includes some staphylococci, E. coli, and P. aeruginosa as well as several anaerobic organisms. Additionally, researchers have shown that meropenem is active against a wide variety of other bacteria in vitro, including Staph. aureus and Staph. epidermidis.

CURRENT THERAPIES

597

FIGURE 12. Structure of meropenem.

A multicenter, open-label clinical trial compared meropenem with imipenem/ cilastatin for safety and efÞcacy in the treatment of patients hospitalized with SSTIs. One hundred eighty-four patients received either 500 mg of meropenem every eight hours or 500 mg of imipenem/cilastatin every six hours over an average of six to seven days. Among clinically evaluable patients, favorable clinical responses occurred in 97.5% (120 of 123) of meropenem-treated patients and in 95.2% (120 of 126) of imipenem/cilastatin-treated patients (Nichols RL, 1995). Meropenem can trigger hypersensitivity reactions in individuals who are allergic to β-lactams, and physicians must monitor patients closely for symptoms of anaphylaxis. The Merrem IV package insert documents adverse reactions and side effects of meropenem noted during clinical trials that included nearly 3,000 patients. Some patients had local adverse reactions typical of IV antibiotics such as inßammation (2.4%), thrombophlebitis (10.8%), and pain at the injection site (0.4%). Systemic adverse reactions included diarrhea (4.8%), nausea/vomiting (3.6%), headache (2.3%), and rash (1.9%). Ertapenem. Ertapenem (Merck’s Invanz) * is a synthetic, parenteral carbapenem with a broad spectrum of activity. The FDA approved the compound in November 2001 for the treatment of moderate-to-severe infections, including cSSTIs and other complicated bacterial infections. Merck began marketing the agent in the United States in early 2002. Ertapenem is now available in Europe for the treatment of community-acquired pneumonia, intra-abdominal infections, and acute gynecological infections. The Zeneca Group (now AstraZeneca) was the original developer of ertapenem, but it discontinued clinical studies in 1996, when Merck subsequently licensed the agent. Ertapenem shares a comparable spectrum of activity with the other carbapenems against gram-positive and gram-negative bacteria—including key pathogens Staph. aureus and enterococci—but has low activity against Pseudomonas aeruginosa and Acinetobacter species. It is indicated for cSSTIs caused by Staph. aureus (methicillin-susceptible strains only), Strep. pyogenes, Escherichia coli, or Peptostreptococcus species. One clinical study compared the efÞcacy of ertapenem versus that of piperacillin/tazobactam in the treatment of cSSTIs. The study enrolled 540 patients, who received either 1 g of ertapenem once daily or 3.375 g of piperacillin/tazobactam four times daily for 7–14 days. The study

598

COMPLICATED SKIN AND SOFT TISSUE INFECTIONS

evaluated clinical efÞcacy at 10–21 days after completion of therapy. The clinical success rates were 83.9% for the 168 patients receiving ertapenem and 85.7% for the 170 receiving piperacillin/tazobactam (Graham DR, 2002). Carbapenems can provoke penicillin hypersensitivity and are contraindicated in patients with a history of penicillin allergies. Zeneca had discontinued development of ertapenem because of adverse effects in 4.7% of patients enrolled in clinical trials. According to the package insert for Merck’s Invanz, side effects included GI disturbances such as diarrhea and nausea (5.5% and 3.1% of patients, respectively), local effects of drug administration such as infused vein complications and thrombophlebitis (3.7% and 1.3% of patients, respectively), and headache (2.2% of patients). Clinical studies additionally reported seizures in 0.5% of patients receiving ertapenem (compared with 0.3% and 0% of piperacillin/tazobactam-treated and ceftriaxone-treated patients, respectively). Resistance to ertapenem remains low, and cross-resistance between ertapenem and imipenem is uncommon. Cephalosporins Overview. The cephalosporins contain a basic β-lactam structure fused to a six-member ring. Drugs in this class differ widely in their spectrum of activity, susceptibility to β-lactamases produced by bacteria, and serum half-life. Cephalosporins are categorized into four generations, with each newer generation representing an expansion or shift in the spectrum of bacterial coverage. Among the cephalosporins, Þrst-generation agents have the narrowest spectrum of activity. They are most active against staphylococci and streptococci, and most are oral formulations. Second-generation cephalosporins have increased activity to cover more gram-negative bacilli, but they are usually less active than Þrst-generation drugs against gram-positive bacteria. Third-generation cephalosporins are active against gram-negative organisms, but their activity against gram-positive organisms is inferior to that of previous generations. Compared with second-generation cephalosporins, the third-generation agents have greater stability against β-lactamases and have longer serum half-lives. As a result, they have more convenient dosing regimens. Finally, fourth-generation agents have enhanced stability against β-lactamases and provide good coverage against both gram-positive and gram-negative bacteria (particularly Pseudomonas aeruginosa). As a class, the cephalosporins are generally well tolerated (Gustaferro CA, 1991; Okamoto MP, 1994). Common adverse effects are usually minor; GI disturbances and thrombophlebitis are the most prominent with oral and parenteral agents, respectively. Disturbances of the GI tract are reported less often with cephalosporins than with the penicillins. The resistance of a pathogen to cephalosporins, as with other β-lactam antibacterials, is a function of the pathogen’s outer-membrane permeability, the speciÞc cephalosporin’s stability against β-lactamases, and its afÞnity for the PBPs. While bacterial β-lactamase activity frequently limits the efÞcacy of antibacterials such

CURRENT THERAPIES

OCH3

N

H N O

599

O

S N

O

NH2

O CO2R

O

FIGURE 13. Structure of cefuroxime axetil (R = CH(CH3 )OAc).

as penicillin and aminopenicillins, many cephalosporins remain effective against these pathogens. However, the relentless march of emerging resistance has not spared the cephalosporin class. Some strains of gram-negative pathogens in the hospital setting have demonstrated increasing immunity to the third-generation cephalosporins, a formidable problem that has been associated with adverse clinical outcomes and increased hospital costs (Cosgrove SE, 2002). Mechanism of Action. Cephalosporins, like the penicillins, bind to PBPs in bacteria and prevent bacterial cell-wall formation. PBPs are involved in numerous roles within the bacterial cell, including maintenance of structural integrity, determination of cell shape, cell division, induction of capsule synthesis, and regulation of autolysis. Cephalosporins, via their β-lactam structure, inhibit these bacterial processes, leading to bacterial cell death. Cefuroxime Axetil. Cefuroxime axetil (GlaxoSmithKline’s Ceftin/Zinnat/Oracef, generics) (Figure 13) is a semisynthetic second-generation cephalosporin available as a tablet or as an oral suspension. Cefuroxime axetil is an esteriÞed pro-drug of cefuroxime created for oral formulation; the active antibiotic metabolite, cefuroxime, is released after the pro-drug is absorbed from the GI tract. First launched in the United States in 1988, cefuroxime axetil has been available from generic manufacturers in the United States since 2002. The agent is active against a broad spectrum of gram-positive and gram-negative bacteria including Staph. aureus, Strep. pyogenes, and E. coli. Cefuroxime axetil is indicated for uncomplicated SSTIs caused by Staph. aureus (including β-lactamase-producing strains) and Strep. pyogenes. A multicenter comparative clinical study demonstrated cefuroxime axetil’s efÞcacy in SSTIs by comparing it (administered in two different dosing regimens) with cefaclor. Over ten days of therapy, 125 patients received 250 mg of cefuroxime axetil twice daily, 500 mg of cefuroxime axetil twice daily, or 250 mg of cefaclor three times daily. Each group achieved clinically beneÞcial outcomes in 92%, 95%, and 97% of patients, respectively (Parish LC, 1987). Another clinical study compared the efÞcacy of three oral cephalosporins in the treatment of SSTIs. The ten-center study enrolled a total of 330 SSTI patients, who were randomly assigned to receive one of three regimens: 250 mg of cefuroxime axetil (107 patients), 500 mg of cephalexin (111 patients), or 500 mg of cefadroxil (112 patients), each twice daily for ten days. Clinicians

600

COMPLICATED SKIN AND SOFT TISSUE INFECTIONS

documented a favorable clinical outcome of cure or improvement in 97% (89/92), 89% (80/90), and 94% (82/87) of the clinically evaluable patients treated with cefuroxime axetil, cephalexin, or cefadroxil, respectively. Furthermore, patients had favorable bacteriological outcomes in 96% (69/72), 85% (60/71), and 93% (63/68) of bacteriologically evaluable cases, respectively (Gooch WM 3rd, 1991). In clinical trials of cefuroxime axetil, patients who received seven to ten days of therapy had adverse reactions similar in type and frequency to those caused by other antibiotic agents. As reported in the Ceftin package insert, diarrhea (3.7%) and nausea/vomiting (3.0%) were among the most common reactions in trials with adults receiving tablets, while pediatric patients receiving the oral formulation experienced diarrhea (8.6%), dislike of taste (5.0%), diaper rash (3.6%), and nausea/vomiting (2.6%). Cefprozil. Cefprozil (Bristol-Myer Squibb’s Cefzil/Procef/Bisoral) is a semisynthetic, second-generation, oral cephalosporin available in a tablet or liquid oral formulation. Cefprozil has been available in the United States since 1992 and is on the market in Italy and Spain. Cefprozil is active against gram-positive and gram-negative bacteria but not Pseudomonas species. Clinical studies demonstrated that this cephalosporin is active against such common SSTI pathogens as Staph. aureus and Strep. pyogenes. Three separate clinical studies enrolled a total of 882 patients to compare cefprozil with cefaclor in the treatment of SSTIs in adults. Patients in the cefprozil groups received 250 mg twice daily, 500 mg once daily, or 500 mg twice daily, while patients in the cefaclor groups received 375 mg twice daily, 500 mg once daily, or 375 mg twice daily. A review of the results from the three trials showed that 492 of 532 (92%) cefprozil-treated patients and 304 of 350 (86%) cefaclor-treated patients achieved clinical cure or improvement (Nolen TM, 1992). Additional clinical studies showed comparable efÞcacy between cefprozil and cefaclor in pediatric SSTI cases (Faingezicht I, 1992). As described in the Cefzil package insert, patients in cefprozil clinical trials had side effects including nausea (3.5%), diarrhea (2.9%), vomiting (1%), and abdominal pain (1%). Cefpodoxime Proxetil. Cefpodoxime proxetil (PÞzer’s Vantin, SanoÞAventis’s Orelox, Sankyo’s Banan) is an extended-spectrum, third-generation, oral cephalosporin available in a tablet or oral suspension. Sankyo originally developed cefpodoxime proxetil and Þrst launched it in Japan in 1989. Upjohn (later Pharmacia, now PÞzer) began marketing it in the United States in 1992, while Aventis obtained marketing rights to it in Europe. A generic version of cefpodoxime proxetil by Ranbaxy has been approved in the United States. Cefpodoxime proxetil is an esteriÞed pro-drug of cefpodoxime created for oral formulation. Cefpodoxime is stable in the presence of most β-lactamase enzymes, effectively extending its spectrum of activity against several grampositive and gram-negative bacteria commonly resistant to penicillins and other cephalosporins. However, some bacterial strains producing extended-spectrum

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β-lactamase enzymes can inactivate cefpodoxime. Clinical studies have demonstrated that cefpodoxime is active against Staph. aureus, Strep. pyogenes, and E. coli ; it is inactive against enterococci and Pseudomonas species. Cefpodoxime proxetil is indicated for the treatment of uncomplicated SSTIs caused by Staph. aureus and Strep. pyogenes. In clinical trials of treatment for SSTI, effective therapeutic treatment with cefpodoxime proxetil was dependent on the dosing schedule; hence the recommended dosing of cefpodoxime proxetil for SSTIs is greater than that for other indications. The recommended daily dosage of cefpodoxime proxetil for SSTIs is 800 mg, which is greater than the 200–400 mg recommended daily dosages for such bacterial infections as pharyngitis, pneumonia, and bronchitis. A double-blind clinical trial compared the efÞcacy of cefpodoxime proxetil and cefaclor in the treatment of SSTIs. One hundred six patients received 200 mg of cefpodoxime proxetil twice daily while 116 patients received 250 mg of cefaclor three times daily; each group received seven days of treatment. Clinical efÞcacy was 74.5% for the cefpodoxime-treated group and 76.7% for the cefaclor-treated group (Yura J, 1988). In clinical trials, 7.0% of patients had diarrhea; the frequency of this effect was signiÞcantly dosage-dependent. Patients also reported nausea (3.3%), vaginal fungal infections (1.0%), vulvovaginal infections (1.3%), abdominal pain (1.2%), and headaches (1.0%). Pediatric trials involving cefpodoxime as a multiple-dose oral suspension documented patients with diarrhea (6.0% overall, 12.8% of infants and toddlers), diaper rash (2.0%), and vomiting (2.3%), according to the Vantin package insert. Cefotaxime. Cefotaxime (Abbott/SanoÞ-Aventis’s Claforan, generics) (Figure 14) is a semisynthetic, third-generation cephalosporin for parenteral administration. Cefotaxime was Þrst marketed in 1981 in the United States, where generic versions are now available. SanoÞ-Aventis holds marketing rights to branded cefotaxime in the United States and Europe, while Roche markets the antibiotic in Japan. Cefotaxime is stable in the presence of many β-lactamase enzymes, effectively extending its spectrum of activity against several gram-positive and gramnegative bacteria commonly resistant to penicillins and other cephalosporins. Clinical and laboratory studies have demonstrated the activity of cefotaxime against enterococci, Staph. aureus, Staph. epidermidis, and Strep. pyogenes and

COONa OCH3 O N N

C

CH2OCOCH3

N CONH S H

H2N

H

S FIGURE 14. Structure of cefotaxime.

602

COMPLICATED SKIN AND SOFT TISSUE INFECTIONS

other streptococci. The agent is also active against some—but not all—strains of Pseudomonas aeruginosa. Cefotaxime is indicated for the treatment of SSTIs caused by the pathogens noted above, as well as E. coli, Pseudomonas species as noted in preceding paragraph, and anaerobic cocci. A series of noncomparative studies evaluated cefotaxime in the treatment of SSTIs. A total of 360 hospitalized patients received a mean dose of 4 g daily in three or four divided doses administered intramuscularly or intravenously for at least Þve days. Of the 260 clinically evaluable patients, 93.5% displayed favorable clinical outcomes; bacteriological results showed pathogen clearance in 84% of the 225 patients evaluated (McCloskey RV, 1982). Although some strains of enterococci (such as Enterococcusfaecalis) and Pseudomonas species display resistance to cefotaxime in vitro, this cephalosporin has successfully treated patients with infections caused by susceptible strains of these organisms. Cefotaxime is generally well tolerated, but the Claforan package insert documents some common adverse reactions, including local inßammation at the administration site (4.3%) and GI disturbances (1.4%). Systemic adverse reactions to cefotaxime have been rare. Ceftriaxone. Ceftriaxone (Roche’s Rocephin/RoceÞn Rocefalin, generics) (Figure 15) is one of the most widely prescribed antibiotics on the market. The agent is a semisynthetic, third-generation parenteral cephalosporin. Generic ceftriaxone has been available in Europe since 2002. In 2000, Cubist Pharmaceuticals announced that it had acquired the rights to develop an oral ceftriaxone formulation. However, the company discontinued development in 2004 after clinical trials showed highly variable bioavailability of the orally administered formulation. Ceftriaxone has demonstrated a broad spectrum of activity against gram-positive and gram-negative bacteria, including Staph. aureus (but not MRSA), Staph. epidermidis, Strep. pyogenes, and other streptococci, as well as E. coli and many gram-negative bacteria. Ceftriaxone is also active against some strains of Pseudomonas aeruginosa. Ceftriaxone is indicated for SSTIs caused by the pathogens noted above, as well as Bacteroides fragilis and anaerobic cocci. A randomized clinical trial to compare the safety and efÞcacy of ceftriaxone and cefazolin enrolled 84 adults

FIGURE 15. Structure of ceftriaxone.

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hospitalized with SSTIs. One group received 1 g of ceftriaxone daily while the other received 3 to 4 g of cefazolin daily; the clinical success rates were 81% and 77%, respectively. (Bradsher RW, 1984). Local reactions have been reported following parenteral administration of antibiotics including ceftriaxone. The Rocephin package insert notes that patients experienced pain, induration, and tenderness in about 1% of cases during clinical trials. Lincosamides Overview. The lincosamides are a small class of drugs that includes clindamycin, an antibiotic commonly used in the treatment of cSSTIs. Lincosamides are active against gram-positive cocci and also have demonstrated efÞcacy against mycoplasma, toxoplasma, and several anaerobic bacteria. For this reason, physicians treating unidentiÞed cSSTI pathogens often add clindamycin to another antibiotic therapy to broaden coverage. Physicians rarely prescribe clindamycin as a monotherapy for patients without a history of hypersensitivity to penicillins; however, clindamycin is often the treatment of choice for patients with such allergies. Mechanism of Action. Lincosamides act by binding to the 23S ribosomal RNA (rRNA) in the 50S subunit of the ribosome. This binding inhibits the translocation of RNA during protein synthesis and blocks initiation of polypeptide formation. Lincosamides are bacteriostatic but can be bactericidal against highly susceptible bacteria or at high concentrations. They have a long post-antibiotic effect that is sufÞcient to allow therapeutic levels to fall below the mean inhibitory concentration (MIC). Clindamycin’s mechanism of action is a key factor in its use for life-threatening cSSTI cases. In serious infections, bacteria can release large amounts of toxin that can cause toxic shock. Because clindamycin blocks protein production, physicians often add this agent to a therapeutic regimen for patients at risk of toxic shock to curtail the production of bacterial toxin. Clindamycin. Clindamycin HCl (PÞzer’s Cleocin/Dalacin/Sobelin, generics) (Figure 16) has been widely available in all major markets since 1970 and today there are many generic suppliers of the agent. Clindamycin is indicated for serious infections caused by susceptible anaerobic bacteria. It is also used in the treatment of serious infections caused by susceptible strains of streptococci and staphylococci. However, physicians usually reserve clindamycin for use against suspected gram-positive bacteria in patients allergic to penicillins. Clindamycin has good bioavailability (90%) and patients rapidly absorb the agent when it is taken orally. Nonetheless, it is available in both oral and parenteral forms. Clindamycin is widely distributed in body ßuids and tissues; serum levels exceed the MIC for most indicated organisms for at least six hours, and excretion is primarily via the kidneys. An early clinical trial compared clindamycin, erythromycin, and penicillin V in the treatment of SSTIs caused by streptococci (with or without coincident

604

COMPLICATED SKIN AND SOFT TISSUE INFECTIONS

CH3 CH3 N

HCCl CONHCH H HO

CH3CH2CH2

O OH

H

SCH3 OH

FIGURE 16. Structure of clindamycin.

staphylococci) in pediatric patients. Two hundred forty-four children were randomly assigned to therapy with one of the three agents for at least seven days. At 14 days, clinical efÞcacies were 99%, 99%, and 97% for the groups receiving clindamycin, erythromycin, and penicillin V, respectively (Dillon HC, 1975). GI disturbances and generalized hypersensitivity reactions (e.g., maculopapular skin rash) are the most frequently reported adverse reactions to clindamycin. Despite the mild nature of most clindamycin side effects, physicians must monitor patients for symptoms of pseudomembranous colitis, a GI complication associated with many antibiotics that can range in severity from mild to life-threatening. Fluoroquinolones Overview. Fluoroquinolones are broad-spectrum antibacterials that have seen an upsurge in use in recent years. Because of their broad spectrum of activity, high efÞcacy, favorable dosing, and oral and IV availability, these agents have become among the most popular and versatile antibacterial agents available. Furthermore, they are indicated for a wide range of bacterial infections, including skin and soft-tissue, respiratory, GI, and urinary tract infections. General practitioners frequently use them as Þrst-line outpatient therapy for a variety of infections in the United States, but their use in SSTI infections is generally limited to the hospital setting. Fluoroquinolones are available in both oral and parenteral formulations and have favorable pharmacokinetics: they achieve rapid absorption from the GI tract and good penetration into the blood and tissues. Most side effects associated with these drugs are mild, such as minor GI disturbances and CNS effects. However, researchers have associated the ßuoroquinolones—as a class—with alterations in normal cardiac conduction, as manifested by a prolongation of the corrected QT interval (QTc) on an electrocardiogram (ECG), and they can cause cardiac arrhythmias in vulnerable individuals. Another important limitation of the ßuoroquinolones is that they are contraindicated for use in children and pregnant women because animal studies have shown that cartilage erosion and other signs of arthropathy may be a common side effect in the young (Lipsky BA, 1999; Zhanel GG, 2002). Further research in this area is ongoing. Other rare, but serious, adverse events associated with ßuoroquinolone use include leukopenia, anemia, and seizures. Fluoroquinolone-resistant isolates are most common among people

CURRENT THERAPIES

605

FIGURE 17. Structure of ciprofloxacin.

aged 65 or older, who are most likely to have had recent hospitalizations (where ßuoroquinolone use can be high). In comparison, penicillin-resistant strains are more common among isolates from young children, who have the highest rate of β-lactam use. Mechanism of Action. Fluoroquinolones interact with two related yet distinct targets within the bacterial cell: DNA gyrase and topoisomerase IV. DNA gyrase and topoisomerase IV are vital enzymes involved in bacterial DNA replication. Interference with this cellular machinery makes the ßuoroquinolones potent inhibitors of nucleic acid synthesis. The exact nature of the interaction of ßuoroquinolones with their target enzymes is not completely understood; however, this interaction blocks the progression of DNA replication, leading to strand breaks and rapid bacterial cell death. Studies imply that the inhibition of DNA gyrase is bactericidal, whereas inhibition of topoisomerase IV is primarily bacteriostatic. Ciprofloxacin. Ciproßoxacin (Bayer’s Cipro/Cipro IV/Cipro XR/Cißox/Ciprobay/Ciproxan Baycip/Ciproxan, generics) (Figure 17) is a second-generation ßuoroquinolone available in both parenteral and oral formulations. First launched by Bayer in the United States, Germany, and the United Kingdom in 1987, ciproßoxacin was available in all major markets by 1990. Bayer’s ciproßoxacin patent expired in various countries in recent years, and generic competition has been very strong, with multiple entrants offering inexpensive formulations of the product. Ciproßoxacin is the ßuoroquinolone of choice among physicians treating cSSTIs because of its excellent tissue penetration, availability in oral and IV formulations, broad spectrum of activity, and Þrst-rate efÞcacy. The introduction of newer-generation “respiratory” ßuoroquinolones (e.g., levoßoxacin, moxißoxacin) has caused a shift in ciproßoxacin use away from respiratory infections. Ciproßoxacin is indicated for use in a large number of bacterial infections including SSTIs caused by gram-positive bacteria (in particular, Staph. aureus [methicillin-susceptible], Staph. epidermidis, and Strep. pyogenes) and gramnegative SSTI bacteria such as P. aeruginosa and E. coli. One double-blind, multicenter, comparative clinical trial compared the efÞcacy and tolerability of ciproßoxacin and another ßuoroquinolone, sparßoxacin, in the treatment of community-acquired cSSTIs. The study enrolled 603 adult patients

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COMPLICATED SKIN AND SOFT TISSUE INFECTIONS

and randomly assigned them to therapy with either ciproßoxacin (750 mg twice daily) or sparßoxacin (400 mg loading dose followed by 200 mg once daily) for a total of 10 days. The two ßuoroquinolones demonstrated similar clinical success rates among the 475 clinically evaluable patients. The study documented clinical cure or improvement in 87.2% (211/242) of patients receiving ciproßoxacin and 90.1% (210/233) of patients receiving sparßoxacin. Bacteriological success rates were 79.9% and 87.0% for patients treated with ciproßoxacin and sparßoxacin, respectively (Lipsky BA, 1999). Clinical trials that enrolled nearly 3,000 patients documented some side effects associated with ciproßoxacin use. The most frequently reported events were nausea (5.2%), diarrhea (2.3%), and vomiting (2.0%). Ciproßoxacin may cause prolongation of the corrected QT interval and cardiac arrhythmias in vulnerable individuals. Concomitant administration of ciproßoxacin and theophylline (still used in some markets for the treatment of asthma and chronic obstructive pulmonary disease) can lead to elevated levels of theophylline, increasing the risk of seizures and other adverse events associated with theophylline toxicity. Aminoglycosides Overview. Although aminoglycosides are one of the preferred antibiotic classes for treating a range of serious infections caused by gram-negative bacilli and gram-positive cocci, they play only a limited role in the treatment of cSSTIs. Their use is typically limited to those cases in which physicians suspect gramnegative pathogens (particularly Pseudomonas aeruginosa). Anaerobic bacteria are intrinsically resistant to aminoglycosides. All aminoglycosides are potentially nephrotoxic, which manifests as acute renal failure, and ototoxic, which manifests as auditory or vestibular dysfunction. These concentration-dependent toxicities are reversible with dose reductions, requiring practitioners to monitor aminoglycoside concentrations in the bloodstream. Mechanism of Action. Aminoglycosides impair bacterial protein synthesis by binding to receptors on the 30S subunit of the bacterial ribosome. Binding to the 30S subunit interferes with the initiation of polypeptide formation, causes the production of defective proteins (misreading of the messenger RNA template), and disrupts ribosome function. Aminoglycosides exert concentration-dependent bactericidal activity; however, the precise mechanisms of this effect are not entirely understood. Gentamicin. Gentamicin (Schering-Plough’s Garamycin/Gentacin, SanoÞ-Aventis’s Cidomycin, Merck’s Ceptopal, generics) (Figure 18) has been available in parenteral formulation since 1966 to treat infections caused by aerobic gramnegative bacilli and gram-positive cocci. A synergistic bactericidal effect against streptococci and enterococci occurs when gentamicin and other aminoglycosides are used in combination with a cell-wall-synthesis inhibitor (e.g., a cephalosporin, glycopeptide). The bacterium Micromonospora purpurea produces the antibiotic

CURRENT THERAPIES

R1

NH

607

R2

O H2N

O

NH2

HO

NH2 O O OH

HO

CH3HN

CH3

FIGURE 18. Structure of gentamicin [gentamicin C1 (R1 = R2 = CH3 ), gentamicin C1a (R1 = R2 = H), gentamicin C2 (R1 = CH3 , R2 = H)].

gentamicin, which binds to the bacterial 30S ribosomal subunit of susceptible pathogens, causing translational errors that result in bacterial cell death. The pharmacokinetic properties of gentamicin in human subcutaneous adipose tissue allow for sufÞcient concentrations to Þght common bacteria easily obtained after IV administration (Lorentzen H, 1996). Because this agent has been available for almost 40 years, there are no recent, rigorous clinical studies pertaining to gentamicin treatment of SSTIs. A review of trials conducted in the 1970s using gentamicin against various bacterial infections notes that clinical studies enrolled a total of 1,098 patients. Eighty-one percent of 697 patients clinically evaluable in these studies achieved clinical cure, including 70% of the 73 patients treated for SSTIs. The usual dosage in these trials was 7.5 mg/kg IM every 12 hours (Gooding PG, 1976). P. aeruginosa is intrinsically more resistant to gentamicin, but physicians can overcome this resistance by adding β-lactam antibiotics, such as penicillins or cephalosporins, to the treatment regimen. Gentamicin has the potential for nephrotoxic and ototoxic reactions, and physicians need to closely monitor blood concentrations of the drug to ensure that the actual levels do not exceed the intended therapeutic levels. Amikacin. Amikacin (Novartis’s Amikin, Bristol-Myers Squibb’s Amiklin/Biklin Amikin, generics) (Figure 19) has been available in parenteral formulation in the United States since 1975 and is active against aerobic gram-positive and gram-negative bacteria. Its spectrum is somewhat broader than that of other aminoglycosides and includes Serratia and Acinetobacter species as well as certain staphylococci and streptococci. Amikacin irreversibly binds to the 30S subunit of bacterial ribosomes, preventing the 50S ribosomal subunit from attaching to the translation initiation complex and effectively inhibiting bacterial protein synthesis. Because this agent has been available for almost 30 years, there are no recent, rigorous clinical studies of amikacin in the treatment of SSTIs. Rather, evidence of its efÞcacy is apparent from its widespread successful clinical use in a variety

608

COMPLICATED SKIN AND SOFT TISSUE INFECTIONS

CH2NH2

O

NH2

NHC

O OH

O

HO

OH CCH2CH2NH2 H

OH

OH CH2OH O NH2 HO

O OH

FIGURE 19. Structure of amikacin.

of bacterial infections including cSSTIs. Amikacin is less susceptible to enzymatic inactivation than gentamicin or tobramycin, and physicians often prescribe amikacin when they need to manage infections caused by gram-negative bacilli that are resistant to these other aminoglycosides. Glycopeptides and Lipopeptides Overview. Physicians use glycopeptides and the relatively new class of lipopeptides for the treatment of severe or life-threatening infections that are caused by gram-positive organisms such as streptococci and staphylococci. These agents have potent narrow-spectrum activity against MRSA and are generally reserved for patients who have nosocomial infections. Alternatively, physicians may choose these antibiotics as second-line agents in patients who have failed to respond to Þrst-line therapy. Glycopeptide therapy is relatively well tolerated; the most severe adverse reactions involve the inner ear (dizziness, vertigo, and/or hearing loss) and the kidneys, but these occur only infrequently. Studies report hypersensitivity (of which skin rash is the most common allergic reaction) in 5–10% of patients. Other possible side effects of glycopeptides include GI disturbances, muscle pain, blood disorders (e.g., eosinophilia), and disturbances in liver enzymes. While glycopeptides demonstrate no activity against gram-negative organisms, cross-resistance between the glycopeptide vancomycin and other antibacterials appears rare among gram-positive microbes. Glycopeptides have been the safety net for patients infected with MRSA, but in recent years some clinicians have isolated vancomycin-resistant bacteria in patients who do not respond to treatment. Lipopeptides, however, have enhanced efÞcacy against these resistant bacteria. Mechanism of Action. Glycopeptides inhibit biosynthesis of the bacterial cellwall by blocking glycopeptide polymerization. This effect produces immediate inhibition of cell-wall synthesis and secondary damage to the cytoplasmic membrane of the cell wall. Glycopeptides also alter the permeability of this membrane and directly inhibit RNA synthesis. However, the impact that these effects have

CURRENT THERAPIES

H3N

HO

Me

OH

HO Me

O

609

O

O

O Cl O

O H

HO

OH

Cl H O O H NH −O

HN H

O

O H

H N

Me

N N

H

O H

O

H

N H

H

+

NH2 H

O

2C

H2N

OH HO

OH FIGURE 20. Structure of vancomycin.

on bacterial viability is largely unknown. Glycopeptides have a concentrationindependent bactericidal action resulting primarily from inhibition of cell-wall synthesis. They are considered slowly bactericidal against staphylococci. Lipopeptides also disrupt the cell membrane of bacteria. The lipopeptide daptomycin inserts its tail into the cell membrane, which disrupts its integrity enough to quickly destroy the membrane potential. This disruption of membrane potential is dose-dependent and clearly correlates with reduced bacterial viability. Unlike glycopeptides, lipopeptides do not inhibit the synthesis of any cell-wall components (Cubist, press release, 2004). Vancomycin. Vancomycin (Lilly’s Vancocin, generics) (Figure 20) is a narrowspectrum, bactericidal glycopeptide antibiotic. Originally developed in the 1950s, vancomycin has been available worldwide since the 1960s and is available in oral and IV formulations. However, oral vancomycin is poorly absorbed from the GI tract; physicians employ IV formulations of vancomycin for systemic infections. Physicians use parenteral glycopeptides extensively to treat systemic infections because the compounds are active against both aerobic and anaerobic grampositive bacteria, including multidrug-resistant staphylococci, Staph. aureus, and Strep. pyogenes (group A β-hemolytic streptococci). Vancomycin has a relatively long duration of action, requiring administration only every 12 hours. Because this agent has been available for more than 40 years, there are no recent, rigorous clinical studies of vancomycin in the treatment of SSTIs. Rather,

610

COMPLICATED SKIN AND SOFT TISSUE INFECTIONS

evidence of vancomycin’s efÞcacy is apparent from its widespread successful clinical use in a variety of bacterial infections including cSSTIs. Patients receiving vancomycin have had transient and permanent ototoxic reactions (inner ear disturbances), but most of these cases are attributed to excessively high IV doses or to patients with a history of hearing complications. Studies have also associated vancomycin with nephrotoxic (kidney) effects. Some patients treated with high doses of IV vancomycin have had serious renal failure, and physicians should be particularly careful about treating patients with another aminoglycoside concomitantly. Clinical experience shows that vancomycin can also cause anaphylactic reactions in allergic patients, resulting in a condition known as “red man syndrome.” Pathogens achieve vancomycin resistance by modifying their bacterial cell wall. Vancomycin exerts its bactericidal effect by binding to components of the bacterial cell wall, which are modiÞed in resistant bacteria; reducing vancomycin’s binding afÞnity and efÞcacy. Teicoplanin. Teicoplanin (SanoÞ-Aventis’ Targocid) is a narrow-spectrum, bactericidal glycopeptide antibiotic similar in structure to vancomycin. First launched in France and Italy in 1988, teicoplanin was available in the other European markets by 1997. Aventis partnered with Fujisawa for distribution and copromotion to launch teicoplanin in Japan in 1997, but the agent has never been available in the United States, where development was suspended in 1999. Teicoplanin is available in parenteral formulations and has a signiÞcantly longer duration of action than vancomycin, allowing once-daily administration. Unlike vancomycin, teicoplanin is available in both IM and IV formulations. Teicoplanin’s spectrum of activity includes both aerobic and anaerobic grampositive bacteria, MRSA, Staph. aureus, Clostridium difÞcile, Clostridium perfringens,and Corynebacterium species. Teicoplanin also often serves as empiric antibacterial therapy in immunocompromised patients with hematological malignancies. This agent is resistant to the β-lactamases produced by bacteria and is several times more potent an inhibitor of enterococci than vancomycin. Physicians generally use teicoplanin in the treatment of serious staphylococcal infections in patients who have failed to respond to the penicillins and cephalosporins or in patients who have infections with staphylococci that is resistant to other antibiotics. Teicoplanin is indicated for the treatment of several bacterial infections including SSTIs. One randomized, comparative clinical trial evaluated the efÞcacy of teicoplanin and cefazolin against gram-positive cSSTIs. The study enrolled 96 hospitalized adults with moderate to severe infections who were randomly assigned to receive one of three therapeutic regimens. One group received teicoplanin intravenously once daily, a second group received teicoplanin intramuscularly once daily, and a third group received cefazolin intravenously three times daily. Of 76 patients who were clinically evaluable, the study documented clinical cure or improvement in 100% (26/26), 95% (21/22), and 93% (26/28) of the IV teicoplanin, IM teicoplanin, and IV cefazolin groups, respectively (Chirurgi VA, 1994).

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Teicoplanin is less likely to cause nephrotoxicity, ototoxicity, and anaphylactic reactions (“red man syndrome”) than vancomycin. However, there remain risks for these adverse reactions and physicians should carefully weigh the beneÞts of therapy against the potential for complications when considering glycopeptide therapy. Daptomycin. Daptomycin (Cubist’s Cubicin) belongs to the newest class of antibiotics—the lipopeptides. Eli Lilly discovered daptomycin in the early 1980s and subsequently licensed the worldwide rights to Cubist Pharmaceuticals in 1997. The agent reached the U.S. market in 2003 and is available only in a parenteral formulation. Daptomycin remains in development for the European and Japanese markets. Daptomycin has a novel mechanism of action that rapidly kills gram-positive bacteria by compromising multiple portions of bacterial membrane function. Daptomycin has activity against most clinically relevant gram-positive bacteria and, most importantly, has excellent efÞcacy against bacteria resistant to methicillin, vancomycin, and linezolid. Two randomized Phase III trials evaluated daptomycin in 1,092 adult patients with gram-positive cSSTIs. These studies compared daptomycin (4 mg/kg IV once daily) with vancomycin (1 g IV twice daily) or a semisynthetic penicillin (4–12 g IV four times daily). The efÞcacy end point for daptomycin was clinical equivalence to the comparator in each trial. Daptomycin met this primary end point in both studies, with a clinical success rate of 83.4%, compared with 84.2% for the comparators among 902 clinically evaluable subjects (Arbeit RD, 2004). According to the Cubicin package insert, clinical trials of daptomycin showed that some patients had elevations in serum creatine phosphokinase (CPK), which may be associated with skeletal muscle effects; physicians should monitor CPK levels for abnormalities and observe patients for symptoms of myopathy. During clinical trials, patients had other adverse reactions, including constipation (6.2%), nausea (65.8%), local injection site reactions (65.8%), and diarrhea (5.2%). Bacteriologic investigations in clinical trials isolated very few daptomycin-resistant bacteria (2 of >1,000 patients). The mechanism of resistance to daptomycin remains unknown and no study has determined whether resistance is transferable. Nitroimidazoles Overview. Nitroimidazoles are potent, bactericidal, anti-anaerobic, and antiprotozoal agents. The most commonly used agent in this drug class is metronidazole, which has been available worldwide since the late 1970s. Formulations of metronidazole contain an inactive pro-drug that aerobic and anaerobic organisms convert to cytotoxic intermediates. Patients with cSSTIs caused by anaerobic pathogens typically receive nitroimidazoles to treat perianal abscesses, diabetic ulcer infections, and gangrene. Mechanism of Action. Although researchers have yet to document the exact mechanism of action of the nitroimidazole class, it appears that these agents require anaerobic conditions for optimal efÞcacy. The key component of the

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COMPLICATED SKIN AND SOFT TISSUE INFECTIONS

antimicrobial effect of nitroimidazoles may be the formation of toxic intermediates and free radicals, which have subsequent cytotoxic effects on susceptible organisms. Metronidazole. Metronidazole (SanoÞ-Aventis/SCS/Shionogi’s Flagyl/Flagyl IV/Metro IV, generics) is indicated to treat intra-abdominal wound infections and other serious nosocomial infections caused by susceptible anaerobic bacteria. In mixed aerobic and anaerobic infections (e.g., abscesses), physicians often combine metronidazole with another aerobic-spectrum antibiotic to provide a regimen with broad coverage. Because this agent has been available since the late 1970s, there are no recent, rigorous clinical studies of metronidazole pertaining to SSTIs. Rather, evidence of its efÞcacy is apparent from its widespread successful clinical use against anaerobic pathogens. Because metronidazole does not target aerobic bacteria or mammalian cells, the drug has few side effects, except when it interacts with alcohol or other contraindicated drugs. According to the Flagyl package insert, two serious adverse reactions reported in patients treated with metronidazole have been convulsive seizures and peripheral neuropathy (numbness, tingling sensation), but these effects are rare. More common, but mild, adverse reactions include nausea, headache, anorexia, and occasionally vomiting, diarrhea, epigastric distress, abdominal cramping, and constipation. Reports of a sharp, unpleasant metallic taste are not unusual, and overgrowth of Candida in the mouth or vagina may also occur. Oxazolidinones Overview. The oxazolidinones (which include only one approved agent, linezolid [PÞzer’s Zyvox/Zyvoxid]), are an antimicrobial class with a unique mechanism of action. The oxazolidinones are available both orally and parenterally and have a bioavailability of 100%. The development of oxazolidinones helped address the need for antibiotics effective against resistant gram-positive bacteria, speciÞcally MRSA. The oxazolidinones provide yet another option for treating gram-positive hospital infections, as they address concerns about the toxicity and resistance patterns of glycopeptides. Furthermore, by switching patients to the oral formulation of oxazolidinones, physicians are able to discharge patients earlier in the treatment cycle, which helps to reduce health care costs. Oxazolidinones have signiÞcant activity against all gram-positive cocci, including strains resistant to methicillin, vancomycin, macrolides, tetracyclines, aminoglycosides, ßuoroquinolones, and sulfonamides. Of particular importance, they are active against MRSA, methicillin-resistant Staph. epidermidis, Strep. pyogenes, and vancomycin-resistant enterococci (VRE). They are bactericidal in a concentration-dependent manner against streptococci and are bacteriostatic against staphylococci and enterococci. The oxazolidinones are also active against anaerobes and have limited activity against some fastidious gram-negative pathogens, but not gram-negative enterobacteriaceae.

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O O

N

N

O

O HN

F

CH3

FIGURE 21. Structure of linezolid.

Because of potential adverse events, physicians must monitor patients during oxazolidinone treatment (Gerson SL, 2002). In particular, physicians must monitor blood chemistries and platelets in patients who have received more than two weeks of oxazolidinone therapy. In vitro studies with linezolid demonstrate a complete lack of cross-resistance with existing antimicrobial agents. Isolated cases of linezolid resistance among treated patients have appeared in the major markets, but the agent remains the only antibiotic capable of treating multidrugresistant organisms without cross-resistance. Mechanism of Action. Oxazolidinones bind to a site on the bacterial 23S ribosomal RNA of the 50S subunit, preventing the formation of a functional 70S initiation complex, which is an essential component of the bacterial translation process. Existing studies have not demonstrated any direct action on DNA or RNA synthesis. Linezolid. Linezolid (PÞzer’s Zyvox/Zyvoxid) (Figure 21) has been available in the United States and the United Kingdom since 2000 and 2001, respectively, and is indicated for cSSTIs, VRE, and nosocomial infections. In June 2001, PÞzer launched linezolid in Japan for the treatment of VRE infections, and it is currently in Phase III trials there for MRSA infections. Available in intravenous, injection, tablet, and oral suspension formulations, linezolid is active against infections caused by a variety of gram-positive bacteria. One clinical study compared the IV-to-oral treatment regimens of linezolid with IV oxacillin and oral dicloxacillin in 826 patients with suspected grampositive deep soft-tissue infections that may have required surgical intervention. This multicenter, international trial randomly assigned patients to treatment with linezolid 600 mg IV every 12 hours followed by 600 mg orally every 12 hours, or treatment with oxacillin 2 g IV four times daily followed by dicloxacillin 500 mg orally four times daily, for a total of 10 to 21 days. Eligible patients had suspected gram-positive cSSTIs for which microbiologists could conduct Gram’s stains. Among clinically evaluable patients, the clinical cure rate for the linezolid-treated patient population was 88.6%, compared with 85.3% in the oxacillin/dicloxacillin-treated patient population. The microbiological success rate was 88.1% for the linezolid group and 86.1% for the oxacillin/dicloxacillin group. The study demonstrated that linezolid is as effective as an oxacillin/dicloxacillin regimen in eradicating Staph. aureus, Staph. epidermidis, Strep. pyogenes, and Staph. agalactiae (Stevens DL, 2000).

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Another study evaluated the efÞcacy of linezolid and vancomycin in the treatment of MRSA SSTIs. One group of 240 patients received 600 mg of linezolid twice daily (Þrst intravenously, then orally) while another group of 220 patients received 1 g of vancomycin twice daily. Among clinically evaluable patients with proven MRSA, 73.2% of the linezolid-treated group and 73.1% of the vancomycin-treated patients achieved clinical cure (Stevens DL, 2002). Clinical studies have revealed some hematological abnormalities (e.g., thrombocytopenia, anemia) that are consistent with mild, reversible, duration-dependent myelosuppression in patients receiving linezolid. This suppression of bone marrow activity limits prolonged use of linezolid. According to the Zyvox package insert, other adverse reactions to linezolid include diarrhea (8.3%), headache (6.5%), and nausea (6.2%). Physicians must monitor patients who have recurrent nausea or vomiting for lactic acidosis. Other side effects of linezolid are relatively benign and are similar to those reported for other antibiotics.

Streptogramin Combinations Overview. The fungi streptomycetes produces streptogramins, which are classiÞed as streptogramin A (dalfopristin) or streptogramin B (quinupristin). These compounds are bacteriostatic when used separately, but in combination they can act in synergy to become bactericidal, mainly against gram-positive bacteria (Allington DR, 2001). Physicians primarily use this streptogramin combination to treat serious nosocomial infections caused by antibiotic-resistant gram-positive bacteria. As with the oxazolidinones, the development of streptogramin combinations helped Þll the need for antibiotics that are effective against MRSA and other resistant bacteria. Despite a favorable spectrum of activity, the need to administer streptogramins through a central line has limited their use and popularity among physicians. The more recent launch of linezolid with its more convenient formulations has almost entirely replaced the use of streptogramin combinations in the treatment of cSSTIs. Mechanism of Action. Streptogramins enter bacterial cells by diffusion and bind to different sites on the 50S ribosomal subunit to form a stable quinupristinribosome-dalfopristin ternary complex, resulting in an irreversible inhibition of protein synthesis. The two compounds serve different roles: quinupristin (streptogramin B) inhibits peptide chain elongation, while dalfopristin (streptogramin A) blocks the peptidyl transferase reaction. The synergistic bactericidal effect of the combination appears to result from conformational changes brought about in the peptidyl transferase center. Streptogramin combinations are bactericidal (concentration-independent) against streptococci and staphylococci and are bacteriostatic against enterococci, but the presence of the constitutive macrolide-lincosamide-streptogramin B (MLSB ) erm resistance phenotype (often found in Enterococcus faecium and MRSA) can limit their bactericidal effect.

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Quinupristin/Dalfopristin. The streptogramin combination of quinupristin and dalfopristin (Q/D) (King Pharmaceuticals/SanoÞ-Aventis/Fujisawa’s Synercid) is a parenteral antibiotic that was approved for use in the United States and the United Kingdom in 1999. In July 2001, the product also became available in France, Germany, Italy, and Spain. Fujisawa began marketing the agent in Japan in 2003. Because Q/D is intended for use as a second-line therapy for resistant organisms, clinical trials of Q/D enrolled patients in need of second-line antibiotics. One trial evaluated the efÞcacy of Q/D in 396 patients infected with vancomycinresistant Ent. faecium (VREF). Most patients (84%) received 7.5 mg/kg intravenously every 8 hours, while 16% received the same dosage every 12 hours, based on the physician’s assessment at day 2 of therapy. The average duration of therapy was 14.5 +/− 11 days. The study reported clinical success in 74% of patients, and there was bacterial eradication in 71% of patients. Arthralgias and myalgias were the most common systemic adverse events (9.1% and 6.6%, respectively) (Moellering RC, 1999). Two randomized, open-label, controlled clinical trials compared Q/D with cefazolin or oxacillin (however, patients received vancomycin in place of the comparator if the clinician believed that resistant pathogens were responsible for the infection). One study was based in the United States while the other was conducted internationally. The studies enrolled a total of 893 patients; 450 received Q/D and 443 received one of the comparator drugs. While Q/D was equivalent to these comparators in clinical success, it fell short in bacteriological clearance in both trials (66% and 67% clearance for Q/D versus 73% and 78% in the comparator groups, respectively). The studies speculated that this lower rate was the result of a higher degree of polymicrobial infections in the Q/D test group; clinical data showed that bacteriological eradication rates for polymicrobial infections were 63% for Q/D and 83% in the comparator group (Nichols RL, 1999). Q/D produces considerable myalgias and arthralgias and has the potential to interact with other drugs. Patients taking Q/D in clinical trials suffered from a much greater rate of venous adverse events (66%) compared with the comparator group (28%). These adverse reactions were responsible for a signiÞcant percentage of patients who discontinued Q/D in these trials (19.1%), while discontinuation of comparator drugs was primarily due to treatment failure (11.5%) (Nichols RL, 1999). Nonpharmacological Approaches Because of their depth and severity, many cSSTIs require immediate surgical attention as well as antibiotic therapy. In numerous cases, cSSTIs encompass a large amount of necrotic tissue that serves as an entry point for further infection. Therefore, it is standard clinical practice to surgically debride (cut away) dead and infected tissue in coordination with an immediate course of antibiotic therapy in the more severe cSSTI cases. Surgical debridement is especially common in abscesses, in which physicians incise the wound and drain the purulent contents (a

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mixture of bacteria and host immune cells). Almost all cases of necrotizing fasciitis and gangrene require surgical debridement because of their life-threatening nature, but physicians attempt to limit debridement in diabetic ulcers, except in cases where the underlying diabetes has damaged the tissue (i.e., the foot) to the point that it is likely to be a continual source of infection. Most surgeons pack these cSSTI incisions, leaving them open for inspection and irrigation in the days following surgery. EMERGING THERAPIES Complicated skin and soft-tissue infections (cSSTIs) represent an area of relatively high unmet need compared with other bacterial infections. A new agent to treat cSSTIs must have high activity against gram-positive organisms and speciÞc activity against methicillin-resistant Staphylococcus aureus (MRSA) or vancomycin-resistant enterococci (VRE) to be considered for this market. In addition, it must be available in an oral formulation for step-down therapy from intravenous (IV) treatment to allow continuous therapy as patients are discharged from the hospital. Even if emerging therapies achieve some or all of these characteristics, they are unlikely to replace current therapies that are less expensive or more familiar to physicians in treating life-threatening conditions. Most emerging therapies will be held in second- or third-line reserve for patients who fail Þrst-line therapy with already established, less expensive antibiotics. The majority of antibacterial agents in late-stage development for the treatment of cSSTIs simply expand the range of choices already available to physicians in conventional drug classes. These drugs aim to provide more effective treatment against resistant pathogens and have some advantages in formulation or dosing. Some drugs in development are effective against a broad spectrum of gram-positive and gram-negative bacteria and will be effective treatments for empirical therapy. Other, narrower-spectrum drugs are being developed to treat gram-positive infections only and will likely be used to treat infections caused by resistant strains (e.g., MRSA) when current Þrst-line agents have failed or in deÞned at-risk patient populations. Table 7 lists emerging therapies in development for cSSTIs. As already discussed, bacterial resistance is a major issue in this market. Unfortunately, bacteria are able to efÞciently transfer genetic material, so resistance is easily propagated among different organisms. Moreover, because many antibiotic agents act via similar mechanisms of action, resistance against one drug often crosses over to include resistance against the entire drug class. Therefore, signiÞcant effort has gone into developing novel classes of antibiotics that will not be susceptible to current mechanisms of resistance. These agents will likely Þnd an increasing role over the next 10 to 20 years as bacterial resistance to current drug classes builds. MRSA rates are very high in many hospitals, and, even more concerning, MRSA rates are rising dramatically in the community. Vancomycin was previously regarded as the most effective agent against all gram-positive pathogens

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TABLE 7. Emerging Therapies in Development for Complicated Skin and Soft Tissue Infections, 2004 Compound

Development Phase

Marketing Company

Glycopeptides Dalbavancin United States Europe Japan

PR PR —

Pfizer Pfizer —

Oritavancin (LY-333328) United States Europe Japan

III — —

Targanta — —

Telavancin United States Europe Japan

III — —

Theravance — —

Cephalosporins BAL-5788 United States Europe Japan

III III —

Johnson & Johnson/Basilea Pharmaceutica Johnson & Johnson/Basilea Pharmaceutica —

RWJ-442831 United States Europe Japan

I — —

Johnson & Johnson — —

DHFR Inhibitors Iclaprim United States Europe Japan

— II —

— Arpida —

Topoisomerase Inhibitors AVE-6971 United States Europe Japan

— I/II —

— Sanofi-Aventis —

until VRE emerged in 1988, followed by glycopeptide intermediate-resistant Staphylococcus aureus (GISA) in 1999. In response, biopharmaceutical companies have focused their efforts since the mid 1990s on new antibiotics (e.g., streptogramins, novel peptides) to be used primarily for infections most often associated with resistant gram-positive bacteria, for which there are few therapeutic alternatives. Emerging narrow-spectrum agents with potent in vitro activity against resistant gram-positive pathogens include Johnson & Johnson’s RWJ442831 (a cephalosporin) and the peptide antibiotic dalbavancin (PÞzer). Table 8 summarizes both the reported minimum inhibitory concentration required to inhibit 90% of the isolates (MIC90 ) of these emerging agents against key grampositive pathogens and the MIC90 of available agents. Most of the emerging

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COMPLICATED SKIN AND SOFT TISSUE INFECTIONS

TABLE 8. In Vitro Activity of Narrow-Spectrum Antibiotics Against Key Nosocomial Gram-Positive Bacteria Antibiotic Vancomycin Teicoplanin Linezolid Quinupristin/Dalfopristin RWJ-54428 Daptomycin Oritavancin Dalbavancin

Pathogen MIC90 (mg/L) MSSE MRSE PSSP

MSSA

MRSA

0.5–1 0.5–1 2–4 0.25–1 0.5 0.12–0.5 2 0.25

1–2 0.5–4 2–4 0.5–2 2 0.12–1 2–4 0.02–0.5

1–2 2–8 1–4 0.12–0.5 0.5–1 0.5 2–4 N/A

VRE >64 >16 2 16–32 0.125–1 1–4 2 32

VSEF VREF 2–4 >64 0.5–2 >16 1–2 1–2 0.25–8 1–2 8 8 2–4 2–4 0.25–0.5 1–4 0.02–4 32

Antibiotic VSE Vancomycin 2–4 Teicoplanin 0.25–1 Linezolid 2 Quinupristin/Dalfopristin 4–32 RWJ-54428 0.25–1 Daptomycin 1–2 Oritavancin 1–2 Dalbavancin 0.02–4

1–2 2–16 1–4 0.25–2 0.5–4 0.25–0.5 2–8 N/A

0.25–0.5 0.12 1 0.5 0.06–0.03 0.25 0.015 0.02–0.06

PRSP 0.25–0.5 0.12 1 0.5–1.0 0.25–0.5 0.25 0.06 0.02–0.5

Other Streptococci 0.25–5 0.5 1–2 0.25–0.5 0.06–2 0.06–2 0.25–1 0.03

MIC90 = Minimum inhibitory concentration required to inhibit 90% of the isolates; MRSA = Methicillin-resistant Staphylococcus aureus; MRSE = Methicillin-resistant Staphylococcus epidermidis; MSSA = Methicillin-susceptible Staph. aureus; MSSE = Methicillin-susceptible Staph. epidermidis; PSSP = Penicillin-susceptible Streptococcus pneumoniae; PRSP = Penicillin-resistant Strep. pneumoniae; Strep = Streptococci serogroups A, B, C, and G; VRE = Vancomycin-resistant enterococci; VREF = Vancomycin-resistant Enterococcus faecium; VSE = Vancomycin-susceptible enterococci; VSEF = Vancomycin-susceptible Ent. faecium. Note: MIC90 data are compiled from several studies that evaluated the in vitro antibacterial activity of emerging agents, which was determined using broth microdilution procedures according to the recommendations of the National Committee for Clinical Laboratory Standards.

narrow-spectrum therapies are as good as or better than current narrow-spectrum agents against common resistant pathogens. However, several of the new agents are signiÞcantly better than current therapies against VRE. These and other emerging agents are proÞled in the following sections. Glycopeptides Overview. Current glycopeptides, such as vancomycin, are used for the treatment of severe or life-threatening infections that are caused by gram-positive organisms, such as streptococci and staphylococci. These agents have potent narrow-spectrum activity against methicillin-resistant Staph.aureus (MRSA) and are generally reserved for patients who have nosocomial infections, or they may be used as second-line agents in patients who have failed to respond to Þrst-line therapy. MRSA is most commonly associated with cellulitis and abscess, but it can be found in other cSSTIs as well. Because of the rising rates of MRSA in hospital and community settings, glycopeptides have become a very important antibiotic class in cSSTI treatment. New glycopeptides need to demonstrate

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TABLE 9. Comparative Activities of Tigecycline Against Key Nosocomial Pathogens Type of Pathogen Gram-positives

Gram-negatives

Anaerobes/intracellullar pathogens

Tigecycline’s Spectrum of Activity Has potent activity against most gram-positive bacteria, including staphylococci, streptococci, enterococci, and Listeria species. Against gram-positive bacteria, its in vitro activity is comparable to that of daptomycin (slightly better than vancomycin). Is effective against isolates resistant to currently available agents, such as MRSA, GISA, PRSP, and VRE. Has good in vitro activity against a wide variety of gram-negative pathogens, including Haemophilus influenzae, Moraxella catarrhalis, Escherichia coli, Klebsiella pneumonia, Acinetobacter spp., Stenotrophomonas (Pseudomonas) maltophilia, Enterobacter spp., and Citrobacter spp., including ESBL-producing strains. Against most gram-negative bacteria, generally less active than ciprofloxacin, imipenem, and ceftazidime. Like other tetracyclines, tigecycline is also active against a range of intracellular pathogens (e.g., Mycoplasma) and anaerobic bacteria (e.g., Clostridium difficile). Is generally more active than cefepime but less active than imipenem against clinically important anaerobes.

ESBL = Extended-spectrum β -lactamase; GISA = Glycopeptide-intermediate susceptible Staphylococcus aureus; MRSA = Methicillin-resistant Staphylococcus aureus; PRSP = Penicillin-resistant Streptococcus pneumoniae; VRE = Vancomycin-resistant enterococci.

increased activity against MRSA and vancomycin-resistant enterococci (VRE) for uptake in cSSTIs, and they will ideally have better dosing and side-effect proÞles than vancomycin, which is the leading antibiotic against resistant infections. To compete effectively with linezolid, new glycopeptides must be available not just in parenteral formulations, but in oral formulations as well. Mechanism of Action. Glycopeptides inhibit biosynthesis of the bacterial cell wall by blocking glycopeptide polymerization. This effect produces immediate inhibition of cell-wall synthesis and secondary damage to the cytoplasmic membrane of the bacterial cell. Glycopeptides also alter the permeability of the cytoplasmic membrane and directly inhibit RNA synthesis. However, the impact of these effects on bacterial viability is largely unknown. Glycopeptides have a concentration-independent bactericidal action resulting primarily from inhibition of cell-wall synthesis. They are considered slowly bactericidal against staphylococci. Dalbavancin. PÞzer (formerly Vicuron) is developing dalbavancin*, a secondgeneration injectable glycopeptide for clinically important gram-positive infections. Dalbavancin’s mechanism of action is the same as that for the glycopeptide class, inhibiting bacterial cell-wall synthesis. However, dalbavancin is a chemically modiÞed version of a naturally occurring glycopeptide that is speciÞcally

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designed to have better activity than vancomycin and teicoplanin against staphylococci, including MRSA. Dalbavancin’s primary advantage over current antibiotics is that it has a long half-life, allowing it to be given just once per week. In vitro studies have shown that dalbavancin is more potent than vancomycin and teicoplanin against clinically important gram-positive bacteria, including MRSA, methicillin-resistant Staph. epidermidis (MRSE), and penicillin-resistant Strep. pneumoniae (PRSP) (Malabarba A, 1998; Jones RN, 2001). Dalbavancin is also more potent than Q/D against most gram-positive bacteria; it is equivalent to Q/D against methicillin-susceptible Staph. aureus (MSSA). Dalbavancin’s activity against VRE is variable: it is effective against VanB-encoded resistance but not against VanA-encoded resistance. Like other glycopeptides, dalbavancin is not effective against gram-negative bacteria. In animal models of infection, a single dose of dalbavancin is reported to be as effective as six doses of linezolid or four doses of vancomycin for the treatment of PRSP and MRSA, respectively (Candiani GP, 2001; Jabes D, 2001). Also, dalbavancin was more potent than teicoplanin or vancomycin in murine models of staphylococcal, streptococcal, and enterococcal septicemia and staphylococcal endocarditis (Malabarba A, 1998). Vicuron began two randomized, double-blind, Phase III clinical trials in December 2002. Vicuron announced results of these trials in August 2004. In a double-blind trial, 854 cSSTI patients were randomly assigned to receive dalbavancin (1 g IV on day 1 and 500 mg IV on day 8) or standard dosing of linezolid for 14 days; 88.9% of dalbavancin patients showed a clinical response at followup visit, versus 91.2% of patients taking linezolid. Dalbavancin was reportedly well tolerated. The second trial was an open-label study that enrolled 156 patients with SSTIs suspected or conÞrmed to be due to MRSA. Patients receiving dalbavancin had an 89.9% rate of clinical response, compared with 86.7% of patients receiving vancomycin. Dalbavancin fared even better in the intent-to-treat group, with 86% of patients receiving dalbavancin showing a clinical response, compared with only 65.3% of patients receiving vancomycin (Versicor press release, August 12, 2004). In September 2002, Vicuron announced positive Phase II clinical trial results for dalbavancin in the treatment of SSTIs (Versicor, press release, September 5, 2002). The randomized clinical trial enrolled 62 patients with infections that involved deep skin structures or required surgical intervention, such as abscesses, infected ulcers, burns, and cellulitis. Dalbavancin administered once weekly for two weeks had higher clinical and microbiological response rates than other standard care regimens, including vancomycin, given for a mean duration of 15 days (Selzer E, 2003). Table 10 shows clinical trial results for dalbavancin in the treatment of skin and soft-tissue infections. In Phase I clinical trials involving 12 patients, dalbavancin administered as a once-daily, 30-minute IV infusion demonstrated linear kinetics with low interindividual variability and good tolerability (Leighton A, 2001[a]; White RJ, 2000). The drug has a serum half-life of 6.6 to 7.8 days; signiÞcant drug accumulation was observed following multiple doses. Patients who received 360 mg of

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TABLE 10. Clinical Trial Results for Dalbavancin in the Treatment of Skin and Soft-Tissue Infections Success of Therapy Evaluable Populations

Dalbavancina

Dalbavancinb

Standard Carec

Clinically evaluabled Microbiological success

16/17 (94.1%) 8/11 (72.7%)

8/13 (61.5%) 3/11 (27.3%)

16/21 (76.2%) 9/14 (64.3%)

a Patients received two doses of dalbavancin given one week apart (day 1 and day 8). b Patients received one dose of dalbavancin given on day 1. c Standard care was any antibiotic (physician’s choice) given daily for 7–21 days. d Cure or improvement observed at first follow-up. Source: Based on Seltzer E, et al. Once-weekly dalbavancin versus standard-of-care antimicrobial regimens for treatment of skin and soft-tissue infections.Clinical Infectious Diseases. 2003;37(10):1298–1303.

dalbavancin exhibited bactericidal activity even when their serum was diluted 8fold 24 hours after dosing. Indeed, seven days after a single dose, plasma levels remained above the minimum bactericidal concentration required for eradication of 90% of the MRSA and MRSE isolates (MBC90 ). These data support the use of dalbavancin in a wide range of dosing regimens, including once-weekly therapy (Versicor press release, November 28, 2001). In all clinical studies to date, dalbavancin has proved well tolerated, whether dosed daily or weekly. No adverse events were considered to be clinically signiÞcant, although pyrexia and headache were reported most frequently. Notably, no ototoxicity (either auditory or vestibular) has been detected for any dose of dalbavancin (Leighton A, 2001[b]). If it is approved, dalbavancin will be the Þrst once-weekly, injectable antibiotic on the market and could become the standard of care for cSSTIs caused by grampositive bacteria, especially staphylococci and streptococci. This possibility is strengthened by dalbavancin’s superiority in potency and tissue penetration, as well as its longer serum half-life, compared with vancomycin. The availability of a once-weekly antibiotic will not only enable more ßexible and convenient dosing regimens; it will encourage shorter hospital stays and may reduce the incidence of local and systemic infections associated with prolonged use of IV lines. Oritavancin. Targanta Therapeutics is developing oritavancin*, a parenteral, Nsubstituted derivative of a semisynthetic glycopeptide antibiotic, LY-264826, for the treatment of gram-positive bacterial infections, including infections resistant to current antibiotics. Oritavancin’s mechanism of action is the same as that described in the “Mechanism of Action” section for the glycopeptide class. In vitro studies have demonstrated that oritavancin has rapid bactericidal activity against all strains of staphylococci, streptococci, and enterococci, including MRSA, multidrug-resistant Strep. pneumoniae, and VRE (Coyle EA, 2001; Jones RN, 1997; Noviello S, 2001; Zeckel ML, 2000). Its potency against these organisms was comparable to or superior to that of vancomycin, teicoplanin, moxißoxacin, ciproßoxacin, imipenem, linezolid, and Q/D (Fasola E, 1996; Rubio MC, 1999; Schwalbe RS, 1996). The drug is also active against a wide range of anaerobic gram-positive bacteria, including some that are not susceptible

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to vancomycin or teicoplanin (Biavasco F, 1997). Activity against anaerobic gram-positive bacteria is similar to or better than that of clindamycin and cefoxitin (Sillerstr¨om E, 1998). Like other glycopeptides, oritavancin is not effective against gram-negative bacteria. In animal infection models, N-substituted derivatives of LY-264826 were effective against lethal Strep. pneumoniae and Strep. pyogenes infections at doses two to six times lower than effective vancomycin doses. These compounds were effective against lethal Staph. aureus infections at doses 1.5 to 3 times lower than effective vancomycin doses (Nicas TI, 1996). Other animal studies show that oritavancin is effective in the treatment of various Staph. aureus and enterococcal infections, including lethal VRE infection and endocarditis due to MRSA (Boylan CJ, 1998; Kaatz GW, 1998; Saleh-Mghir A, 1997; Schwalbe RS, 1997). Resistant variants have not been observed after prolonged exposure to the drug, but development of moderate resistance in some VREs has occurred in vitro˜(Arthur M, 1999). Table 11 depicts the results of a double-blind, randomized Phase III clinical trial involving 517 patients with cSSTIs, which demonstrate that oritavancin therapy can halve treatment time—from 11.5 days to approximately 5.5 days—compared with the current standard combination of vancomycin plus cephalexin follow-up treatment (Wasilewski MM, 2001[b]). This decrease in treatment time stems from the elimination of the oral cephalexin stepdown therapy. However, because step-down therapy is inexpensive and convenient, the impact of this reduced treatment time could be negligible. In a small Phase II clinical trial involving 27 patients with gram-positive bacteremia, oritavancin demonstrated clinical success. Bacterial eradication was conÞrmed in four of Þve patients with vancomycin-resistant Enterococcus faecium infections, in the three patients infected with vancomycin-sensitive Enterococcus faecalis infections, in the only patient infected with Strep. pneumoniae, TABLE 11. Clinical Trial Results for Oritavancin in the Treatment of Complicated Skin and Soft-Tissue Infections Caused by Gram-Positive Bacteria

Evaluable Populations

1.5 mg/kg Oritavancina

Intent to treat (n = 480) Clinically evaluabled (n = 384) Microbiological success (n = 256) Clinically evaluable with MRSA (n = 33) Relapse rate at late follow-up (day 60) Average length of therapy

63% (n = 163) 76% 74% 74% 10% 5.3

Success of Therapy 3.0 mg/kg 30 mg/kg Oritavancinb Vancomycinc 63% (n = 153) 76% 74% 74% 4% 5.7

65% (n = 164) 80% 76% 80% 5% 11.5

a Administered IV once daily, followed by oral placebo. b Administered IV once daily, followed by oral placebo. c Administered IV in two divided doses daily, followed by oral cephalexin (500–1,000 mg, twice daily). d Cure or improvement observed at first follow-up (day 28).

Source: Based on Wasilewski M, et al. Equivalence of shorter course therapy with oritavancin compared to vancomycin/cephalexin in complicated skin/skin structure infections (CSSI). Clinical Microbiology and Infection. December 2001; Chicago, IL. Abstract UL-18.

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and in the only patient infected with methicillin-resistant Staph. epidermidis (MRSE) (Wasilewski MW, 2001[a]). Phase I clinical trials have demonstrated that oritavancin (once daily IV infusion over 30 minutes) has favorable pharmacokinetics, with a serum half-life of approximately 10.5 days and low inter-individual variability (Chien J, 1998; Thomasson HR, 1999). The bacteria-killing kinetics of oritavancin are significantly affected by inoculum size and the presence of serum, which result in higher MICs, longer killing time, and shorter post-antibiotic effect; this problem is easily solved by increasing the concentrations of administered doses (Mercier RC, 1997; Zhanel GG, 1998). Safety data for oritavancin are limited, but preliminary data from clinical trials suggest that the drug is generally well tolerated. However, additional data relating to phlebitis, edema, anemia, tremor, elevated liver enzyme, and renal function abnormalities are needed to better evaluate the compound’s safety and tolerability. Telavancin. Theravance (formerly Advanced Medicine) is developing its oncedaily injectable glycopeptide telavancin * for gram-positive infections. Telavancin has an extended mechanism of action, inhibiting both peptidoglycan synthesis and phospholipids synthesis in cell-wall production, which Theravance hopes will lower the level of spontaneous resistance against the drug. Telavancin is expected to be effective against MRSA and methicillin-resistant Ent. faecalis. Theravance initiated Phase III trials for telavancin in cSSTIs in late 2004. Researchers reported results of a Phase II study of telavancin in September 2004. A total of 167 patients with suspected or conÞrmed gram-positive cSSTIs were randomized and given either telavancin or standard therapy (a penicillin or vancomycin). For the 102 patients with Staph. aureus, telavancin cured 80% of patients while standard therapy cured 77%. Patients with MRSA (44 total) who received telavancin were cured 82% of the time, compared with only 69% of MRSA patients receiving standard therapy. It is not clear from available data whether standard therapy for these patients was solely vancomycin, or if some patients received only standard penicillin that would normally be ineffective against MRSA. Approximately 5% of patients withdrew from each arm because of adverse events, though fewer serious adverse events were reported for patients taking telavancin than for those on standard therapy, 4 versus 9, respectively (Stryjewski M, 2004). Phase I studies in 119 healthy volunteers who received a 7.5 mg/ kg or 15 mg/kg dose of telavancin as a 60-minute infusion over three days demonstrated linear pharmacokinetics, which were equal in men and women. This study also showed that at the higher dose, 33% of patients had nausea, 21% had headache, 15% had vomiting, dizziness, and rash, and 10% had abdominal pain/cramps. These adverse effects were classiÞed as mild. More detrimental, at the higher dose, 10% of patients discontinued administration because of infusionrelated “red man syndrome,” which is a serious allergic response (Barriere S, 2003). This response is one of the chief complaints about vancomycin, and it will severely limit the uptake of telavancin.

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Cephalosporins Overview. The cephalosporin class is currently very competitive, with many branded and generic drugs available. Drugs in this class are widely used for a variety of infections; thus, doctors are comfortable prescribing new members of the class. However, this high physician acceptance is tempered by formulary restrictions that are placed on new cephalosporins that are not perceived to offer a signiÞcant advantage over less expensive or generic drugs in the same class. Novel cephalosporins with greater potency than those that currently exist, especially against MRSA, may Þnd Þrst-line use in cSSTIs, but they must prove especially effective to compete with the other members of this large class. Mechanism of Action. Cephalosporins, like all β-lactams, bind to PBPs in bacteria and prevent bacterial cell-wall formation. PBPs are involved in numerous roles within the bacterial cell, including maintenance of structural integrity, determination of cell shape, cell division, induction of capsule synthesis, and regulation of autolysis. Cephalosporins inhibit these bacterial processes, leading to bacterial cell death. BAL-5788. Basilea Pharmaceutica is developing a novel cephalosporin, BAL5788 (ceftobiprole, a pro-drug of BAL-9141), that has broad-spectrum activity and additional potency against MRSA and penicillin-resistant Strep. pneumoniae (PRSP). The FDA has granted BAL-5788 two fast-track designations, one in March 2003 for the treatment of cSSTIs due to MRSA and one in June 2004 for hospital-acquired pneumonia due to MRSA. Roche had an option to license BAL-5788 but turned it down in May 2004, leaving Basilea with the global development, marketing, and manufacturing rights. BAL-5788 recently completed additional Phase II clinical trials for cSSTIs, though results are not yet available. Basilea initiated Phase III trials in November 2004. BAL-5788 is an intravenous fourth-generation cephalosporin with coverage against both gram-positive (including Ent. faecalis) and gram-negative pathogens. Like other β-lactams, BAL-5788 binds to bacterial PBPs to inhibit cell-wall production. Unlike many other β-lactams, though, BAL-5788 provides very good potency against MRSA. MRSA is resistant to most β-lactams using two mechanisms. The Þrst is the expression of β-lactamases that cleave antibiotics such as methicillin and other β-lactams. The second resistance mechanism is that MRSA acquires a new PBP from the chromosomal determinant mecA. This protein, dubbed PBP2a, has a much lower binding afÞnity for methicillin and similar compounds, and these drugs are therefore very inefÞcient at blocking the protein’s activity. BAL-5788 is effective against MRSA because it is 10,000 times more resistant to hydrolysis by β-lactamases than penicillin G and it has a more than 100 times greater afÞnity for PBP2a than methicillin. Basilea announced in November 2004 that it was initiating Phase III trials that will compare BAL-5788 with vancomycin in 700 MRSA patients worldwide (company press release, November 10, 2004). In June 2004, Basilea reported positive results from a Phase II trial of BAL-5788 (company press release, March 4, 2004). Forty hospitalized patients with cSSTIs requiring surgical intervention

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were enrolled in the trial. Most had deep infections caused by streptococci or staphylococci, and some patients were infected with MRSA. Of the 35 patients who were clinically evaluable, Basilea reported that all were cured. No major organ toxicity was reported, and the most frequent side effect was mild to moderate nausea. Further details were not disclosed. In a Phase I trial in 40 healthy male volunteers, 30-minute infusions of BAL-5788 equivalent to 150, 250, 500, 750, and 1,000 mg of the active compound BAL-9141 were evaluated (SchmittHoffmann A, 2002). No adverse effects were reported, and analysis of the data suggested that the 750 mg dose infused twice a day would be effective against MRSA. A second Phase I trial was conducted in 5 healthy volunteers and 15 patients with varying degrees of renal impairment (Roos B, 2003). Each patient was given a single 250 mg dose by IV and monitored for Cmax (peak concentration, usually taken at end of infusion) and the steady-state distribution of the drug. All groups were reportedly equivalent in both parameters. RWJ-442831. RWJ-442831 is a pro-drug that is converted metabolically into the active cephalosporin, RWJ-54428. RWJ-54428 entered Phase I clinical trials in the United States in November 1999, but observations of injection-site irritation resulted in the development of RWJ-442831 (Essential Therapeutics, press release, June 10, 2002). In July 2002, Essential Therapeutics announced that RWJ-442831 had entered Phase I clinical trials after preliminary studies in animals had demonstrated reduced venous irritation at the injection site than RWJ-54428. Essential developed this parenteral cephalosporin in collaboration with Johnson & Johnson Pharmaceutical Research & Development (formerly R.W. Johnson) for the treatment of antibiotic-resistant gram-positive infections. However, Essential Þled for bankruptcy in May 2003, and Johnson & Johnson has since taken over development. The pro-drug RWJ-442831 is expected to have in vitro antibacterial potency comparable to that of RWJ-54428. Studies have shown that the activity of RWJ54428 is equivalent to, or better than, the activity of vancomycin, imipenem, linezolid, ceftriaxone, and cefotaxime against most gram-positive bacteria, including VRE, MRSA, MRSE, GISA (glycopeptide intermediate-resistant Staph. aureus), and PRSP (penicillin-resistant Strep. pneumoniae) (Chamberland S, 2001; Johnson AP, 2002; Swenson JM, 2002). Against mecA-positive Staph. aureus and ampicillin-resistant Ent. faecium, RWJ-54428 is less potent than Q/D. Although only marginally effective against most gram-negative bacteria, RWJ-54428 has also demonstrated signiÞcant in vitro activity against the common respiratory pathogens Haemophilus inßuenzae and Moraxella catarrhalis. Other Þndings about RWJ-54428 may also apply to RWJ-442831. For example, in vitro studies indicate that coadministration of RWJ-54428 with antibiotics that are effective against gram-negative bacteria, although not synergistic, is not antagonistic (Foleno B, 2000). In animal models of infection (e.g., septicemia, pyelonephritis, endocarditis, pneumonia), RWJ-54428 also has proved highly effective in clearing infection caused by gram-positive bacteria, including MSSA and MRSA (Frosco MB, 1997; GrifÞth D, 1997; Liu C, 1999). Singledose pharmacokinetic data on RWJ-54428 in animal studies suggest a serum

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half-life of 1 to 2.3 hours for humans, which is similar to that of other marketed cephalosporins (Harford L, 1998). As a result, RWJ-442831 is likely to be administered twice daily. RWJ-442831 is a promising new parenteral cephalosporin for the treatment of infections caused by gram-positive bacteria, particularly those resistant to available drugs. However, the success of this agent, should it proceed in development, will depend on clinical trials demonstrating safety and efÞcacy. Also, because several antibiotics with good activity against resistant gram-positive bacteria have recently been launched (e.g., Q/D, linezolid), the extent to which RWJ-442831 is used for cSSTI treatment will depend on the success of competing antibiotics. Dihydrofolate Reductase Inhibitors Overview. Dihydrofolate reductase (DHFR) inhibitors are a drug class in development that researchers believe will be less susceptible to resistance. Members of this drug class are currently used to treat cancer (e.g., methotrexate) or some bacterial infections. Trimethoprim/sulfamethoxazole is currently used for urinary tract infections and some respiratory tract infections, but it is subject to high levels of resistance. If this resistance does not cross over to new DHFR inhibitors, then these may be effective drugs to treat cSSTIs while limiting resistance because the drug class is relatively novel. Mechanism of Action. Members of this drug class are effective against bacteria by blocking the action of the bacterial DHFR enzyme. DHFR plays an essential role in the synthesis of thymine nucleotides. By inhibiting synthesis of thymine, DHFR inhibitors target DNA and RNA synthesis, thereby causing bacterial cell death. More speciÞcally, folate donates a methyl group to uracil to make thymine. In the process, the folate molecule is oxidized and must be reduced by DHFR to allow another cycle of synthesis. If DHFR is inhibited, the absence of reduced folate causes a precipitous drop in the levels of thymine nucleotides, causing uracil to be erroneously incorporated in its place in growing DNA strands. Replication is eventually disrupted by this process, killing the bacteria (Goodsell 1999). Bacterial DHFR is distinct enough from human DHFR that drugs targeting the bacterial DHFR protein are generally not toxic to human cells. Iclaprim. Arpida has licensed all commercialization rights of the DHFR inhibitor iclaprim (AR-100) from Roche; the drug has completed Phase II trials in Europe for cSSTIs. Previous clinical trials have evaluated an IV formulation of iclaprim, but Arpida is working to develop an oral formulation as well. Iclaprim works by the mechanism of action described previously, and because there is only one other antibiotic in the DHFR inhibitor class, Arpida hopes that existing bacterial pathogens, including MRSA and VRSA, will prove sensitive to it. Arpida also reports that iclaprim has good tissue penetration, a critical asset for treating deep cSSTIs. In January 2004, Arpida announced results from a Phase II trial in which iclaprim was tested for efÞcacy and tolerability in hospitalized cSSTI patients (including those with infected burns, diabetic foot ulcers, cellulitis, and abscesses)

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(Arpida press release, January 9, 2004). Arpida enrolled 92 patients in this double-blind trial and compared 1 or 2 mg/kg of iclaprim to a standard dose of vancomycin. Both drugs were given intravenously twice a day for 10 days. Twenty-six of 28 patients (92.9%) taking 1 mg/kg iclaprim were clinically cured, compared with 29 of 31 patients (93.5%) taking 2 mg/kg iclaprim and 26 of 28 patients (92.9%) taking vancomycin. Microbiological success rates were similar for 1 mg/kg iclaprim and vancomycin (57.1% and 50%, respectively) and were superior for 2 mg/kg iclaprim (76.7%). Iclaprim was well tolerated, and adverse events were similar for all three groups. Topoisomerase IV Inhibitors Overview. The topoisomerase IV inhibitors—which target bacterial replication—are a new class of antibiotics that addresses the problem of drug resistance. This class is related to the ßuoroquinolones in their mechanism of action, though it is too early to determine the extent of similarity between the two classes. Drugs in this class will likely have efÞcacy against both gram-positive and gramnegative bacteria and will compete with other broad-spectrum agents in Þrst-line treatment of cSSTIs. Like the new DHFR inhibitors, these drugs will be aimed primarily at treating resistant bacteria such MRSA and VRSA. Mechanism of Action. Along with DNA gyrase (topoisomerase II), topoisomerase IV is a protein involved in bacterial DNA replication. Both of these proteins are the target of ßuoroquinolones, although individual drugs vary in their afÞnity for either one. It is unclear how individual topoisomerase IV inhibitors will differ from current ßuoroquinolones in their mechanism of action, if at all. Topoisomerase IV is responsible for decatenation and segregation of the bacterial chromosomes after replication. Inhibition of this activity stops the replication process and ultimately kills the bacteria. AVE-6971. SanoÞ-Aventis is developing AVE-6971 * as a broad-spectrum antibiotic with the potential to treat pathogens that are resistant to currently available drugs. AVE-6971 is currently in Phase I/II trials in Europe, but SanoÞ-Aventis has not released any details about the study or the indication. AVE-6971 has the same mechanism of action as previously described for this class, but it is said to be distinct from the mechanisms of ßuoroquinolones and linezolid. In preclinical studies using a mouse thigh abscess model, oral AVE-6971 was found to be ten times more potent than subcutaneous vancomycin; the 50% effective dose (ED50 ) against MRSA was found to be 10 mg/kg for oral AVE-6971 compared with an ED50 of 100 mg/kg for subcutaneous vancomycin (Aventis company presentation, R&D Day 2002, June 18, 2002). REFERENCES Allington DR, Rivey MP. Quinupristin/dalfopristin: a therapeutic review. Clinical Therapeutics. 2001;23(1):24–44.

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Alverdy J. Gut-derived sepsis occurs when the right pathogen with the right virulence genes meets the right host: evidence for in vivo virulence expression in Pseudomonas aeruginosa. Annals of Surgery. 2000;232:480–489. Arakawa S, et al. Clinical studies of S-4661, new parenteral carbapenem antibiotic, in complicated urinary tract infections. Abstracts of the 37th Interscience Conference on Antimicrobial Agents and Chemotherapy (ICAAC). September 1997; Toronto, Canada. Abstract F218. Arbeit RD, et al. The safety and efÞcacy of daptomycin for the treatment of complicated skin and skin-structure infections. Clinical Infectious Diseases. 2004;38(12):1673–1681. Arthur M, et al. Moderate-level resistance to glycopeptide LY333328 mediated by genes of the vanA and vanB clusters in enterococci. Antimicrobial Agents and Chemotherapy. 1999;43:1875–1880. Baker MD, Acharya KR. Superantigens: structure-function relationships. International Journal of Medical Microbiology. 2004;293:529–537. Barbieri JT. Pseudomonas aeruginosa exoenzyme S, a bifunctional type-III secreted cytotoxin. International Journal of Medical Microbiology. 2000;290:381–387. Barriere S, et al. Pharmacokinetics and safety of a new antibacterial, TD-6424, in healthy subjects. 43rd Interscience Conference on Antimicrobial Agents and Chemotherapy (ICAAC). September 2003. Chicago, Illinois. Abstract A-20. Betriu C, et al. [Epidemiologic study of infection by resistant gram-positive bacteria (G + R Study)] Enfermidades Infecciosas Microbiologia Clinica. 2003;21(1):7–11. Biavasco F, et al. In vitro antibacterial activity of LY333328, a new semisynthetic glycopeptide. Antimicrobial Agents and Chemotherapy. 1997;41:2165–2172. Biedenbach DJ, et al. In vitro antimicrobial activity of GAR-936 tested against antibiotic-resistant gram-positive bloodstream infection isolates and strains producing extended-spectrum β-lactamases. Diagnostic Microbiology and Infectious Diseases. 2001;40:179–187. Boucher HW, et al. In vitro activities of the glycylcycline GAR-936 against gram-positive bacteria. Antimicrobial Agents and Chemotherapy. 2000;44:2225–2229. Boylan CJ, et al. Pharmacodynamics of LY333328 in a neutropenic mouse thigh model of Staphylococcus aureus infection. Abstracts of the 38th Interscience Conference on Antimicrobial Agents and Chemotherapy. September 1998; San Diego, CA. Abstract A-41. Bradsher RW Jr, Snow RM. Ceftriaxone treatment of skin and soft tissue infections in a once daily regimen. American Journal of Medicine. 1984;77(4C):63–67. Brook I. Soft tissue and muscular infections (including group A streptococci). Bailliere’s Best Practice and Research Clinical Rheumatology. 1999;13(1):95–114. Calvet HM, Yoshikawa TT. Infections in diabetes. Infectious Disease Clinics of North America. 2001;15(2):407–421. Candiani GP, et al. EfÞcacy of a single dalbavancin (DA) dose compared with multiple linezolid (LN) doses against penicillin-resistant pneumococci (PRSP) in a lobar pneumonia (LP) model in the immunocompetent rat (IR). Abstracts of the 41st Interscience Conference on Antimicrobial Agents and Chemotherapy. December 2001; Chicago, IL. Abstract B989.

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Hepatitis C

ETIOLOGY AND PATHOPHYSIOLOGY Introduction Hepatitis is a disease of the liver that is characterized by hepatic inßammation and hepatocellular necrosis (destruction of liver cells). The disease may be acute or chronic and can be caused by various agents, including alcohol, drugs, and viruses. Acute hepatitis is usually a self-limiting course of liver inßammation. Chronic hepatitis denotes ongoing inßammation of the liver, which leads to progressive hepatic Þbrosis, cirrhosis, end-stage liver disease, and an increased risk of hepatocellular carcinoma (HCC). The Þve major viral hepatitides are hepatitis A through E; diagnosis depends on serologic assays speciÞc to each of the different viruses. Etiology Viral hepatitis was historically separated into two types of infection based on epidemiological characteristics: “infectious” hepatitis (associated with oral-fecal transmission) and “serum” hepatitis (associated with the percutaneous transfer of material containing human serum). With the advent of serological testing, it became evident that most cases of infectious hepatitis were due to hepatitis A virus (HAV) and that many cases of serum hepatitis were associated with hepatitis B virus (HBV) (note that HBV is also associated with sexual transmission). Still, clinicians recognized that not all cases of serum hepatitis were due to HBV, and the term non-A, non-B (NANB) hepatitis was used in these cases (Feinstone SM, 1975). In 1989, scientists conclusively identiÞed hepatitis C virus (HCV) as the organism responsible for most cases of NANB hepatitis, an achievement that Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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resulted from a joint effort involving Chiron and the Centers for Disease Control and Prevention (CDC) (Choo QL, 1989; Kubo Y, 1989). A major obstacle to HCV drug development has been the fact that HCV is exceedingly difÞcult to grow in cell culture; indeed, it was the Þrst infectious agent to be discovered entirely through analysis of cloned nucleic acid sequences. The lack of reliable cell culture systems and animal infection models has hampered studies of the viral life cycle, as well as the development of new antivirals or vaccines. These issues have been somewhat overcome with the development of a viral replicon system (Lohmann V, 1999). The replicon model for HCV replaced genetic sequences encoding HCV structural proteins with drug-resistance genes, enabling selection of cells in culture that maintained infection with the clone. This system allows for viral replication in cell culture and the study of individual viral proteins but does not allow for the study of viral assembly, propagation, and infection in vitro or in vivo. Use of the HCV replicon system as the model for characterization of drug-induced inhibition of HCV-encoded proteins has driven the current expansion of therapeutics against viral targets like the polymerase and protease proteins. More recently, scientists studying a genotype 2a HCV isolate (genotypes are discussed in greater detail later in this chapter) have established a full-length replicating model of HCV genotype 2 virus, which can produce infectious particles in cell culture (Lindenbach BD, 2005; Wakita T, 2005; Zhong J, 2005). This new experimental model system may enable further development in drug discovery because researchers will be able to broaden the focus of their search for potential antiviral targets or targets of host cell proteins involved in viral propagation. However, although experts widely consider this genotype 2a HCV isolate to be a major development in the creation of tools to study this important virus, many thought leaders state that the key will be development of similar replicating HCV genomic models for the more-difÞcult-to-treat HCV genotype 1 virus. Key Viral Proteins and Drug Targets. HCV is a small, enveloped, positivestrand RNA virus of the Flaviviridae family and the only member of the genus Hepaciviridae. It is genetically unrelated to the hepatitis B virus. Despite the lack of culture systems available to efÞciently replicate the virus, the genome has been studied extensively and is now well characterized. The hepatitis C viral genome is approximately 9.4 kb in length and encodes a polyprotein of approximately 3,000 amino acids. This polyprotein is processed by host signal peptidase (localized in the endoplasmic reticulum [ER]) and by two viral proteases to generate three major structural proteins and several nonstructural (NS) proteins necessary for viral replication (Figure 1). In the following sections, HCV genomic sites and their translation products are described. Experts are enthusiastic about the prospects of drugs targeting HCV proteases and polymerases. HCV RNA polymerase and proteases play important roles in the HCV life cycle and thus persistence of infection: protease activity is required to enable activity of HCV-encoded proteins, and the RNA polymerase is essential for viral replication. Drugs targeting these important functions are likely to be launched within the ten-year forecast period. Other long-term targets include entry inhibitors and

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Open reading frame (ORF)

Nonstructural protein precursor

Gene 5'

UTR

Protein function

C

E1

E2

HVR1

RNAHyper5' untrans- binding Envelope variable glycocore region lated region, protein proteins IRES

NS2

NS3

NS4A NS4B NS5A

Serine protease/ helicase

Protein translation and transport inhibitor

Antiapoptotic factor and forms protease NS2-NS3 with NS3

NS3 protease cofactor

NS5B

UTR

Untranslated region 3'

RNA polymerase

Interferon sensitivitydetermining region

Regulation of RNA transcription and stability

FIGURE 1. Hepatitis C viral genome.

therapeutic vaccines. Table 1 lists emerging HCV drug targets and the potential advantages and disadvantages of drugs modulating these targets. NS2 and NS2/3 Protease. Cleavage of the HCV polyprotein between the NS2-NS3 junction is mediated by the NS2/3 protease, an autocatalytic protease encoded within NS2 and the N-terminus of NS3 (Wu Z, 1998). Host proteins located on the endoplasmic reticulum are required to sustain efÞcient cleavage of the NS2-NS3 junction (Santolini E, 1995). The largely hydrophobic HCV NS2/3 protease is essential for the normal infectious cycle of HCV and is a major focus for development of HCV-speciÞc therapies. Once processed by the NS2/3 protease, the NS2 protein is targeted to the endoplasmic reticulum, forming an integral transmembrane protein (Santolini E, 1995; Yamaga AK, 2002). Research has suggested that NS2 may inhibit gene expression and interact with the liver-speciÞc proapoptotic protein CIDE-B by inhibiting its ability to induce cell death of infected hepatocytes (Dumoulin FL, 2003; Erdtmann L, 2003). In this manner, NS2 may be essential for promoting viral persistence and may be a key factor in disease progression to HCC. NS3 Protease. The N-terminal region of the NS3 protein encodes a 180-aminoacid serine protease required for processing Þve of the six HCV nonstructural proteins (Tomei L, 1993). It cleaves four sites within the polyprotein, yielding the NS3, 4A, 4B, 5A, and 5B proteins. NS3 is an induced-Þt protease, requiring both the NS4A cofactor protein and the substrate to fully activate its catalytic machinery (LaPlante SR, 1999; Pizzi E, 1994; Yan Y, 1998). Also, the NS3 protease contains a tetrahedral zinc-binding site, which guides folding of the NS3 protease, maintains protein activity, and provides structural stability (De FR, 1998). The NS3 protease is essential for HCV replication and is a primary target for novel antiviral therapies. Many investigational inhibitors of the NS3 protease have taken advantage of the susceptibility of that protease’s active site to inhibition

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by its cleavage products (corresponding to the NS4A/NS4B, NS4B/NS5A, and NS5A/NS5B N-terminal cleavage sites). NS3 Helicase. The NS3 protein also contains a helicase domain at its Cterminus. Helicase is an ATP-dependent enzyme that catalyzes the unwinding of duplex RNA prior to its replication by RNA-dependent RNA polymerase (RdRp; encoded by NS5B). The strand-separating activity is associated with hydrolysis TABLE 1. Advantages and Disadvantages of Emerging Drug Classes with Specific HCV-Life-Cycle-Regulating Targets Emerging Drug Class Polymerase inhibitors

HCV-Specific Targets NS3 helicase, NS5B (RNA polymerase), NS5A (transcription activator)

Protease inhibitors

NS2, NS2/3, NS3, proteases

Replication cofactors

NS4A

HCV-encoded immunomodulators

NS2, NS4A, NS4B, NS5A

Advantages/Disadvantages of Drugs Modulating Activity of These HCV-Specific Targets + HCV-specific, no immunomodulating properties, potentially reducing side effects.

+ Can potentially block propagation of virus. − High susceptibility for development of antiviral drug resistance, forcing a requirement for a drug with high bioavailability and long-term persistence. − Less likely to be available as a monotherapy as it may require combination therapy to boost clearance of the nonpropagating virus. + HCV-specific, no immunomodulating properties, potentially reducing side effects. + Can potentially block propagation of virus. − High susceptibility for development of antiviral drug resistance, forcing a requirement for a drug with high bioavailability and long-term persistence. − Less likely to be available as a monotherapy as it may require combination therapy to boost clearance of the nonpropagating virus. − Difficult to disable owing to structural chemistry surrounding the active site of the target. + HCV-specific; no immunomodulating properties, potentially reducing side effects. − Possibly not as efficient in therapeutic use, leading to potential as an enhancer to existing or emerging therapies. + HCV-specific. + Potentially increase efficacy of existing combination therapies by reducing or blocking interferon resistance or by blocking antiapoptotic properties of some targets. − Host-pathways affected by these proteins are poorly defined, thus efficacy may be unimpressive.

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of nucleoside triphosphate (NTP), which disrupts the hydrogen bonds that keep the two strands together. The RNA helicase activity of HCV NS3 is enhanced in the presence of NS4A and, unlike the majority of NTPase/helicases, the HCV NS3 helicase is capable of unwinding DNA/DNA, RNA/RNA, and DNA/RNA duplexes. In addition, the NS3 helicase is thought to confer the speciÞcity of NS5B for HCV viral RNA (Piccininni S, 2002). Because the HCV NS3 helicase structure has been well characterized, it is regarded as a promising target for antiviral therapy, though drugs in development against this target are still early in preclinical development. NS4A. NS4A is an essential cofactor for the NS3-dependent processing of the NS3-NS4A, NS4A-4B, and NS4B-NS5A junctions. In addition, although its presence is not required, NS4A increases the efÞciency of cleavage at the NS5A-5B site. This membrane-associated protein is believed to enhance the stability of the NS3 N-terminal structure and may have several molecular functions: • • • • •

Modulating NS3 serine protease activity through binding and cleavage of NS3 (Yang SH, 2000). Increasing NS3 stability by targeting it to the membrane of the ER. Enhancing RNA synthesis by tethering the replication complex to ERassociated NS3. Promoting phosphorylation of NS5A. Mediating inhibition of protein synthesis, an important process for regulating viral replication.

NS4B. NS4B is a hydrophobic, transmembrane protein with possible roles in HCV replication and inhibition of protein translation and transport. Using electron microscopy, researchers found that NS4B induces morphological changes in the ER (Gretton SN, 2005), which may play a role in HCV replication. Researchers have also observed NS4B-mediated inhibition of protein translation (Florese RH, 2002; Piccininni S, 2002) and have demonstrated inhibition of ER-to-Golgi protein transport in the presence of unprocessed NS4A/B (Konan KV, 2003). The authors of these studies suggest that inhibition of protein translation and transport may downregulate a cell’s ability to secrete cytokines and express MHC molecules, allowing HCV to circumvent the host immune response. Furthermore, NS4A and NS4B have been shown to induce expression of interleukin-8 (IL-8) (Kadoya H, 2005), which has previously been shown to play an important role in angiogenesis, tumor growth, and metastasis. NS4B also plays a critical role in HCV RNA replication through binding and hydrolysis of GTP (Einav S, 2004; Elazar M, 2004). In other ßaviviruses, such as dengue, West Nile, and yellow fever virus, NS4B has been shown to block activation of interferon- (IFN-) alpha and beta responsive transcription factors such as STAT1, and IFN-stimulated response elements (Munoz-Jordan JL, 2005). Although similar activity has not yet been described for the HCV NS4B, if the HCV NS4B functions similarly to its counterpart in related ßaviviruses, it is possible that NS4B plays a role in IFN resistance.

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NS5A. Although associated with other HCV proteins as part of the viral replication complex, NS5A’s role in viral replication has not yet been deÞned. It is thought that differential phosphorylation of NS5A may affect its role in RNA replication (Appel N, 2005). Proper phosphorylation of NS5A requires the expression of the protein in its polyprotein context (Koch JO, 1999). NS5A is known to be associated with membranes and to act as a transcriptional activator. It displays a multitude of activities that are believed to trigger crucial steps in the pathophysiology of HCV infection. For example, NS5A has been shown to interact with growth factor receptor-binding protein 2, p53, p21, interleukin (IL)-8, NF-κB, and the signal-transduction pathway for tumor necrosis factor-alpha (TNF-α) (Qadri I, 2002; Reyes GR, 2002; Waris G, 2002). Many of these interactions block the cellular response to persistent HCV infection by inhibiting apoptotic processes. NS5A has distinct homology to the antiapoptotic protein Bcl-2, and NS5A was able to interact with the proapoptotic protein Bax to inhibit apoptosis in p53negative HCC cells (Chung YL, 2003). Clearly, the activity of NS5A and NS2 in preventing apoptosis may explain the chronic persistence of HCV in the liver and, with the activity of NS4A and NS4B, its progression to HCC. Drug therapies that are unable to kill infected cells (i.e., antibodies that only clear extracellular virus) may be ineffective at preventing HCV disease progression. NS5A has generated considerable interest because it has been implicated in causing HCV tolerance to IFN treatment (He Y, 2002). Although the idea is controversial, the molecular mechanism of this IFN tolerance is thought to involve inhibition of the IFN-induced, double-stranded, RNA-activated protein kinase (PKR) by NS5A. PKR is a major mediator of the IFN-induced biological response. NS5B. NS5B encodes the RNA-dependent RNA polymerase (RdRp) activity of HCV and is the central catalytic enzyme responsible for HCV replication. As such, this protein represents a promising and popular target for antiviral drug discovery (Leveque VJ, 2002). Alone, NS5B lacks speciÞcity for HCV RNA; it requires additional viral factors, such as the NS3 helicase and NS4A, to recognize and initiate viral replication. Indeed, the NS3 helicase is believed to dramatically modulate template recognition by NS5B (Piccininni S, 2002). Advances in understanding the structure-function of NS5B, such as the identiÞcation of features important for binding the RNA template, have provided a framework for structure-based drug design. Nucleoside and non-nucleoside inhibitors of NS5B have been identiÞed, and both are the subject of ongoing research efforts. Untranslated Regions. Within the 5
and 3
ends of the HCV genome, two untranslated regions (UTRs) are critical for viral transcription and translation. The 5
UTR contains an internal ribosomal entry site (IRES) that is involved in the initiation of translation. This highly conserved region of the HCV genome is the subject of much research because IRES-mediated translation occurs via a unique prokaryotic-like mechanism that makes it an attractive antiviral target (Gallego J, 2002).

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HEPATITIS C

The 3
UTR consists of a poorly conserved short segment (approximately 28–42 bases), followed by a polyU(C) region of variable length and a highly conserved 98-base sequence that is essential for viral replication (Blight KJ, 1997). This highly structured region of the HCV genome is thought to promote HCV RNA genome stability and may serve as a replicase recognition site during the initiation of minus-strand RNA synthesis. Core Protein. The core protein (or nucleocapsid protein) is immunogenic, and antibodies directed against the core protein are typically present in infected persons. A number of biological activities have been associated with the core protein; these biological activities include its role in alterations in cell-cycle regulation, transcription of cellular proto-oncogenes, induction or suppression of apoptosis, and suppression of the host immune response (Ray RB, 2001; Soo HM, 2002). The signiÞcance of these biological activities in the pathophysiology and persistence of HCV infection is not clear. However, research indicates that the core protein’s most important role is encapsidation of the viral RNA prior to viral budding. Aspartic acid 111 has been mapped as a critical residue for virus assembly and budding, and may represent a novel site for drug development (Blanchard E, 2003). E1 and E2. E1 and E2 are envelope proteins that play pivotal roles in viral assembly, budding, and HCV entry of host cells; for these reasons, they are important targets in HCV vaccine development. The mechanism by which HCV penetrates hepatocytes remains loosely deÞned. Strong evidence indicates that the human CD81 receptor, which is expressed on various cell types, is involved in the attachment of HCV via the hypervariable regions (HVR1) of envelope protein E2 (Drummer HE, 2004; Drummer HE, 2005; Hughes MG, Jr., 2005; McKeating JA, 2004; Meyer K, 2004). Additional host-cell surface components have been implicated to play a role in cellular tropism of HCV entry, including scavenger receptors and lectins (Bartosch B, 2003; Cormier EG, 2004b; Cormier EG, 2004a). E2-speciÞc antisera can block binding and entry of HCV to host cells, conÞrming the important role of E2 in HCV host cell invasion (Keck ZY, 2004). In addition to facilitating entry, these proteins may have other roles. E1 and E2 form a stable heterodimeric complex that anchors viral particles to the ER during replication of the virus (Op De BA, 2001). Furthermore, E2 has been implicated as a potential inhibitor of protein kinase R (PKR), which may contribute to the resistance of HCV to IFN (Pavio N, 2002). Hypervariable Regions. Analysis of the HCV genome has revealed the presence of three hypervariable regions (HVRs) located within the E1, E2, and NS5A regions. Hypervariable region-1 (HVR-1), near the amino terminus of E2, is approximately 30 amino acid residues in length and is thought to exist as a polypeptide loop on the surface of the virion. Functional analysis of E2 suggests that HVR-1 is involved in modulating CD81 receptor binding, perhaps through SR-B1 binding (Bartosch B, 2003; Roccasecca R, 2003).

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Infected people frequently possess neutralizing antibodies (antibodies that block viral replication) to the HVR-1 sequences of the virus with which they are infected. Researchers believe that the appearance of such antibodies results in the selection of variant viruses with HVR-1 sequences that are less reactive; this important pathogenic mechanism leads to evasion of the host immune response and persistent infection (Mondelli MU, 2001). Genetic Variability. Genetic variability is a key feature of HCV genomes and one that they share with many other RNA viruses, such as HIV and dengue virus. Viral heterogeneity appears to result primarily from the high transcription error rate associated with the RdRp, as well as from the absence of any proofreading mechanisms. The resulting genetic hypervariability poses a challenge to drug and vaccine development. New drugs targeting HCV proteins (polymerases and proteases) may be susceptible to drug resistance, and vaccines designed to target a speciÞc viral epitope may be rendered ineffective with shifts in the peptide sequence of the antigen. HCV exhibits genetic heterogeneity on three levels: genotypes, subtypes, and quasi species. Within each subclassiÞcation, the percentage nucleotide difference in the RNA sequences declines: 31–35% of viral RNA differs between genotypes; 20–23% of RNA differs between subtypes; and in quasi species, only 1–9% of the nucleotides differ (Bukh J, 1995). Genotypes. Genotypes represent the accumulation of mutations that have occurred over the long-term evolution of the virus. Phylogenetic analyses of the NS5 and E1 sequences from HCV samples taken from around the world have identiÞed six major genotypes. These genotypes have been further divided into closely related subtypes. The nomenclature designates the genotype with an Arabic numeral; the subtype is denoted by a lowercase letter. Table 2 illustrates the distribution of HCV genotypes in the major pharmaceutical markets. Genotypes 1, 2, and 3a are broadly distributed in patients around the world, whereas the other types are more geographically restricted. Most people are infected by a single genotype. Blood transfusion is responsible for the worldwide spread of genotype 1b; in contrast, needle sharing among drug users is associated with the transmission of genotypes 1a and 3a in most industrialized countries. Genotype 3b is found mainly in the Indian subcontinent; genotype 4 is found mainly in the Middle East and Africa; genotype 5 is found mainly in South Africa; and genotype 6 is found mainly in Hong Kong. The clinical consequences of infection by the various genotypes remain unclear. Although investigations have shown that liver steatosis is particularly frequent in patients with HCV genotype 3 infection (Rubbia-Brandt L, 2004), an association between severity of disease (over a natural course of infection) and HCV genotype has not been found (Freeman AJ, 2003; Reid AE, 1999). The inßuence of genotype on the expected response to therapy is somewhat clearer. Indeed, HCV genotype has emerged as an important factor in predicting a sustained response to antiviral therapy and determining the duration of such treatment. Genotypes 2, 3, and 5 respond better to pegylated interferon alpha (pegIFNα)/ribavirin combination therapy (the current gold-standard therapy) than do

646

HEPATITIS C

TABLE 2. Geographic Distribution of HCV Genotypes in the Major Pharmaceutical Markets, 2004 Country

1a

1b

2a

2b

United States France Germany Italy Spain United Kingdom Japan

x x x x x x x

x x x x x x x

x x

x

2c

2 nsp

x x x x

x x x

x x

3a x x x x x x x

3b

4a

4 nsp

5

x x x

6 x

x x

x x

x x x

nsp = Not specified. Note: The distribution of HCV genotypes is constantly changing, and an accurate assessment of the actual distribution of prevalence of HCV genotypes has not recently been determined. Data in this table reflect data collected from published reports or cases describing HCV genotype prior to 2005.

genotypes 1 and 4. Patients with genotypes 1, 4, 5, and 6 respond better to 48 weeks of combination therapy than to 24 weeks, whereas responses in patients with genotypes 2 or 3 infection are similar after 24 and 48 weeks of therapy. Furthermore, patients infected with HCV genotype 1 require 1,000–1,200 mg of ribavirin daily to achieve optimal response rates, whereas only 800 mg daily is needed for patients infected with genotype 2 or 3. The molecular etiology behind the differential treatment outcomes for genotype 1 versus genotype 2- or 3-infected patients remains unclear. Quasi Species. The term quasi species describes the heterogeneous mixture of genetically related but sequence-distinct variants that circulates within each HCV-infected person’s bloodstream. A quasi species is generated simply by random mutations that occur as virus is replicated in a patient’s cells. This random mutation generates a mixed pool of heterogeneous viral variants, with certain genetic variants constituting the bulk of virions. The genetic variant that is in the majority is termed the quasi species. At any time, one quasi species predominates within an infected host, although this dominant form may change or be replaced over time. Changes in the dominant quasi species, through selection and expansion of less common genetic variants, can result from spontaneous changes in the host environment (e.g., immune responses) or be triggered by factors such as concurrent infections or drug intake. The ability of HCV to survive for decades in the host is a direct result of its genomic plasticity and the evolution of quasi species within an infected person. This evolution enables the virus to continually evade the host immune response. HCV’s ability to avoid detection until it has become well entrenched permits the development of persistent HCV infection in 60–85% of all people who have contracted the virus. The genetic ßuidity that is so characteristic of HCV also presents a signiÞcant barrier to the development of vaccines against this infection. HCV Transmission. Before the identiÞcation of HCV, the majority of non-A, non-B hepatitis cases were transmitted and acquired through blood transfusions.

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Since the isolation of the virus and the institution of antibody screening measures and nucleic-acid testing to exclude blood from infectious donors, the risk of transfusion-related HCV infection has been virtually eliminated. Indeed, most newly acquired cases of hepatitis C in the major markets are related to intravenous drug use. Other risk factors include occupational exposure to contaminated blood (primarily via needle sticks), hemodialysis, sexual transmission, vertical transmission (transmission from mother to child), and nonintravenous drug use (e.g., intranasal cocaine). In addition, percutaneous exposure through body piercing and tattooing are possible (but unconÞrmed) means of transmission. Accumulated evidence indicates that sexual transmission of HCV does occur, but the efÞciency of transmission is much lower than that of other sexually transmitted viruses, such as HBV and HIV. The transmission rate from mother to infant is also low—4% to 7%—and is correlated with high levels of maternal viremia (i.e., high viral load in the maternal blood) and HIV coinfection. Indeed, coinfection with HIV increases the rate of vertical transmission four- to Þvefold (Roberts EA, 2002). Despite the plethora of risk factors, most studies have been unable to identify a speciÞc risk factor in approximately 10% of all patients examined (Alter MJ, 1998).

Pathophysiology HCV Disease Progression. The natural course of HCV infection and disease varies widely. Although HCV-related liver disease is a leading cause of mortality in adults and is the primary reason for liver transplantation in the major markets (Kim AI, 2005; Kim WR, 2001; World Health Organization Initiative for Vaccine Research, 2005), experts tell us that the vast majority of carriers die with rather than from this infection. Disease manifestations are absent or minimal in the majority of cases. HCV RNA can be detected in the blood within one to three weeks after initial exposure; however, a signiÞcant proportion of infected individuals will experience few or no symptoms of infection and therefore will remain unaware of their disease. Chronic HCV infection is diagnosed by the repeated detection of virus in the blood over a period of at least six months. Patients with persistent viremia are at risk of Þbrosis and cirrhosis (extensive Þbrosis that distorts the structure and degrades the function of the liver), but the extent of liver damage and the time course of disease progression vary among individuals. Because acute infection is often clinically silent, it has proved difÞcult to establish the true incidence and time course of progression to disease. Retrospective studies have found that approximately 20% of all infected people will eventually develop cirrhosis, typically after a “silent period” of 20–40 years. Nevertheless, a meta-analysis of community-based cohorts whose populations most closely represent the current HCV-infected populations in the major markets indicates that the risk of progression to cirrhosis within 20 years of infection is less than 10% (Freeman AJ, 2001). Few natural history studies have gone

648

HEPATITIS C

Infection (often asymptomatic)

Typical time to progression

Weeks or months

Natural resolution of infection 5-25%

Progression to chronic infection 75-95%

10 years Infection persists but disease does not develop, or progresses only to mild or moderate disease 80% (usually asymptomatic; serum ALT concentrations may be normal or persistently or intermittently elevated)

20 years

Progression to cirrhosis 20%

30 years

Cirrhosis progresses slowly and patient dies from other causes 50-70%

Progression to decompensated cirrhosis and liver failure 20-30%

Progression to hepatocellular carcinoma 10-25% of cirrhotic patients

ALT = Alanine aminotransferase.

FIGURE 2. The natural history fo hepatitis C infection.

beyond the Þrst 20 years of infection, so the true outcome of HCV infection beyond this time period is not known. Cirrhosis can remain asymptomatic for several years as healthy tissue compensates for diseased tissue. However, once it is established, complications such as jaundice, ascites, variceal hemorrhage, and encephalopathy may ensue. The development of these complications deÞnes decompensated cirrhosis, or end-stage liver disease. In patients with decompensated cirrhosis, the Þve-year survival rate declines to 50%. In addition, approximately 10–25% of patients who develop cirrhosis go on to develop HCC. The course of infection and disease varies in individual patients on three levels: at the level of the viremia, at the level of histology, and at the level of symptoms. In the following sections, the course of events that may ensue from HCV infection are discussed (Figure 2). Spontaneous Resolution of HCV Infection. With regard to viremia, infection may follow two main courses:

ETIOLOGY AND PATHOPHYSIOLOGY

•

•

649

Approximately 5–25% of people who contract HCV recover spontaneously within 2–12 weeks (although antibodies persist, the virus has been eradicated from the blood). The remaining 60–85% of infected people fail to eradicate the virus and develop a chronic infection.

Numerous studies have attempted to identify factors associated with the spontaneous resolution of infection. Accumulated evidence indicates that incubation period, degree of elevation of liver enzymes, and viral load during the acute phase are not predictive of whether the infection will resolve or become chronic. It has been noted, however, that children and young women are more likely to undergo a spontaneous resolution of infection than are adults, whereas AfricanAmerican men are among those with the highest risk of HCC (Fleckenstein J, 2004). Patients who experience symptomatic hepatitis (e.g., jaundice) during the acute phase of infection are also less likely to develop chronic hepatitis C than those who are asymptomatic in the acute phase. Most patients who spontaneously clear the virus after acute hepatitis C infection demonstrate a strong TH 1 immune response to infection (secreting type-1 cytokines, such as IL-2, IFNγ , and TNF-α), with little or no TH 2 response (secreting type-2 cytokines, such as IL-4, IL-5, and IL-10), whereas those who develop chronic hepatitis C show the reverse. Other host-dependent genetic factors—speciÞcally human leukocyte antigen (HLA) class—have been shown to be related to viral clearance or disease progression (Jinushi M, 2003; Mangia A, 1999; Wang Y, 2003). Studies in the chimpanzee model of HCV infection have revealed that a strong intrahepatic T-cell response (rather than a peripheral T-cell response) to HCV correlates with viral clearance (Su AI, 2002; Thimme R, 2002). Moreover, it is suggested that some mechanism other than the destruction of infected hepatocytes is responsible for the virus’s clearance. Chimpanzees that transiently and permanently cleared the infection were observed to initiate a vigorous, multispeciÞc, IFNγ -producing, intrahepatic T-cell response. These chimpanzees were also observed to upregulate genes involved in antigen presentation and the adaptive immune response. In contrast, a strong type 1 IFN (IFNα/IFNβ) response was observed in all chimpanzees, regardless of the outcome of infection, suggesting that HCV is relatively resistant to the antiviral effects of type 1 IFNs. Histology of HCV Infection. At the histological level, patients with chronic HCV have historically been classiÞed as having chronic active or chronic persistent hepatitis; that is, more or less progressive disease as evaluated by the extent and nature of liver injury. This nomenclature captures an important feature of HCV; the disease progresses more quickly in some patients than in others. One patient who has been infected for 30 years may have mild disease, whereas another who has been infected for only 10 years may have severe disease. Under this system, the future rate at which the disease will progress is indicated by how fast it progressed between the date of infection and the date of diagnosis.

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TABLE 3. Histological Activity Index (Knodell Score) for Assessment of the Extent of Liver Disease Histological Feature Periportala necrosis, including piecemeal necrosisb (PN) and/or bridging necrosisc (BN)

Severity None

Mild PN Moderate PN Marked PN Moderate PN + BN Marked PN + BN Multilobular necrosis Intralobular degeneration and focal necrosis

Score 0

1 3 4 5 6 10

None

0

Mild Moderate Marked

1 3 4

Portal inflammation

None Mild Moderate Marked

0 1 3 4

Fibrosis

None Fibrous portal expansion Bridging fibrosis Cirrhosis

0 1 3 4

Maximum score

22

a Surrounding the portal vein. b Disruption of the periportal hepatocytes by inflammatory cells.

In reality, people infected with HCV may Þnd themselves anywhere on a continuum of severity of liver disease. For this reason, most specialists have rejected the terms chronic active and chronic persistent in favor of newer grading systems that describe the virus’s histological activity in the liver (grade) and degree of progression (stage). Grade is based on the degree of necrosis (death of an area of tissue), inßammation, and Þbrosis (formation of Þbrous tissue). The most popular grading system used in clinical trials is the Histologic Activity Index, or Knodell score, shown in Table 3 (Knodell RG, 1981). This score is formulated by scoring based on evaluation of four parameters with regard to function/dysfunction of the liver, with a lower score indicating fewer histological anomalies: periportal and/or bridging necrosis (scored 0 through 10), intralobular degeneration and focal necrosis (scored 0 through 4), portal inßammation (scored 0 through 4), and Þbrosis (scored 0 through 4). The scores from the Þrst three categories are then summed to provide a quantitative value for the severity of inßammation: • •

0 = No inßammation. 1–4 = Minimal inßammation.

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• • •

651

5–8 = Mild inßammation. 9–12 = Moderate inßammation. 13–18 = Marked inßammation.

The Þbrosis portion of the Knodell score comprises the fourth category and is scored from 0 to 4, although no value is set for a score of 2 in this scale: • • • •

0 = No Þbrosis. 1 = Fibrous portal expansion. 3 = Bridging Þbrosis (portal-portal or portal-central linkage). 4 = Cirrhosis.

This scaling is similar to the Metavir scoring system, which aims to evaluate liver function and histology on two scales: grade and stage. In the Metavir system, grade denotes the activity or amount of inßammation and is scored 0 through 4 with 0 being no activity and 3 or 4 indicating severe activity. The stage scoring indicates the amount of Þbrosis and is also scored 0 through 4: • • • • •

0 = No scarring. 1 = Minimal scarring. 2 = Scarring present and extends outside of areas of the liver that are peripheral to blood vessels. 3 = Bridging Þbrosis, spreading and connecting areas of Þbrotic tissue. 4 = Cirrhosis or advanced scarring of the liver.

Because of the similarities between the Þbrosis portion of the Knodell score and the Metavir scoring system, clinical trials will often report the effect of a drug on changes in Þbrosis status as quantiÞed using the Metavir scale or by using only the Þbrosis portion of the Knodell scoring. Outside the context of clinical trials, hepatitis may be graded simply as mild, moderate, or severe. The Child-Pugh score is a grading/evaluation system for cirrhosis, based on analysis of various liver function tests (Table 4). It involves summation of score values assigned to results from Þve different liver function/status tests, and the summation of these scores is divided into three classes (A, B, and C, with A being more mild cirrhosis and C indicating end-stage liver disease). Although this scoring system is not typically used in describing the effects of a drug in clinical trials, it is often referred to concerning medical practice and treatment options in cirrhotic patients. Symptoms of HCV Infection. Although the majority of patients do not develop symptoms during the acute phase of infection, 20–30% of them experience malaise, fatigue, weakness, anorexia, or right upper quadrant pain, followed by the appearance of jaundice. Patients who go on to develop chronic infection generally have few symptoms during the Þrst 20–40 years after infection. A subset of chronic HCV patients (approximately 20%) will develop nonspeciÞc symptoms, including mild fatigue and malaise, nausea, and right upper quadrant

652

HEPATITIS C

TABLE 4. Child-Pugh Grading System for Evaluation of Cirrhosis Score

Albumin (g/dL)

Bilirubin (mg/dL)

PT (sec. prolonged)

Hepatic Encephalopathies

Ascites (grade)

1 2 3

>3.5 2.8–3.5 <2.8

<2 2–3 >3

1–4 4–6 >6

None 1–2 3–4

None Mild Severe

After summation of scores for each category, the Child-Pugh class is defined in the following manner: A = 5–6. B = 7–9. C = 10+ (end-stage liver disease). PT = Prothrombin time.

pain. Many patients with chronic hepatitis C Þrst exhibit symptoms when cirrhosis develops, although patients can continue with cirrhosis without symptoms for several years. Risk Factors for Progression. In 1999, L. Pagliaro and colleagues published a meta-analysis of variables predicting the probability of progression to cirrhosis. Their study showed that the stage of disease at diagnosis was indeed a key indicator of the risk of progression, particularly if the patient had been infected for at least ten years. Each of nine longitudinal studies that Pagliaro assessed found that higher necroinßammatory grading or staging of Þbrosis at diagnosis was associated with more frequent and accelerated progression to cirrhosis (Pagliaro L, 1999). Other factors consistently associated with a higher risk of progression to severe liver disease include excessive alcohol consumption, smoking, advanced age at the time of infection, gender, the presence of competing etiologies for liver disease (e.g., superinfection with HAV or HBV), and coinfection with HIV (Hezode C, 2003; Poynard T, 1997; Seeff LB, 1999). Disease progression is not associated with the level of viremia; however, disagreement about the role of HCV genotype in disease progression is evident in the literature. Persistent elevation of serum aminotransferase (ALT) levels (10–20 times above normal) is often indicative of progression to chronic hepatitis, but the magnitude and frequency of elevation of serum ALT levels correspond poorly with the extent of Þbrosis (Shiffman ML, 2000). Furthermore, whereas serum HCV RNA levels remain fairly constant once persistent viremia is established, serum ALT levels may ßuctuate independent of symptoms or disease progression. Approximately 30% of patients with chronic hepatitis C have serum ALT levels that are normal, and another 40% have serum ALT levels that are less than two times the upper normal limit (Bacon BR, 2002). Of patients with normal/nearnormal ALT levels, up to 35% may show signiÞcant inßammation and Þbrosis upon liver biopsy. Disease Mechanisms. The Host Immune Response. HCV is thought to principally exert its pathological effects indirectly, via the host’s immune response, rather than through its

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653

own cytopathic properties. In most infected patients, a wide array of antibodies are generated to structural and nonstructural regions of the virus, but the clinical signiÞcance of this humoral (B-cell-mediated) response with respect to controlling infection and disease remains poorly understood. Unfortunately, infection persists in most patients despite the development of HCV-directed antibodies. Furthermore, in studies of chimpanzees, animals that were previously infected could later be reinfected, despite the development of a humoral response (Farci P, 1992). Specialists believe that the neutralizing immune responses are strainspeciÞc and that the quasi-species nature of the virus allows other strains to emerge when the dominant strain comes under immune pressure. The cellular arm of the immune system appears to play the major role in both controlling infection and damaging the HCV-infected liver. The mechanisms by which CD4+ helper T cells (TH ) and CD8+ cytotoxic T cells (TC ) eradicate infection in some patients and contribute to liver damage in others are unclear, but evidence suggests that an imbalance between the TH 1 and TH 2 response results in the inability to clear infection and the progression of liver disease (Sobue S, 2001). An early TH 1 response indirectly promotes TC activity and establishes the antiviral cellular responses. Conversely, researchers hypothesize that in chronic hepatitis C, the T-cell response is polarized early to a TH 2 response, which allows viral persistence rather than viral clearance (Jacobson Brown PM, 2001). The persistence of HCV in infected hepatocytes causes continuous hepatic inßammation and injury. Cytokine-Induced Hepatocellular Damage and Fibrosis. After years of HCV infection, Þbrosis may develop. Fibrosis is characterized by the accumulation of collagen and other insoluble extracellular matrix proteins that ultimately destroy the liver architecture. During this process, proinßammatory cytokines (e.g., TNFα, IL-1, IL-6) are released by Kupffer cells in the liver or by inÞltrating neutrophils and macrophages. These cytokines stimulate normally quiescent stellate cells, which grow adjacent to hepatocytes, to secrete collagen and other proteins that impair the ability of liver cells to process materials. If Þbrosis progresses, cirrhosis can result; cirrhosis is characterized by bands of Þbrosis that enclose nodules of regenerating hepatocytes. Researchers are investigating ways to disrupt the process by which stellate cells become stimulated as a means of preventing disease. One such target is transforming growth factor-beta (TGF-β), a potent Þbrogenic cytokine produced by Kupffer and stellate cells. Note, however, that the liver can continue to function for many years despite the presence of cirrhosis. HCV Cytopathic Effects. Some evidence also suggests that HCV can, like other members of the Flaviviridae family, induce tissue damage directly. Histopathologists sometimes Þnd damaged hepatocytes in the absence of adjacent inßammation. Several liver transplant centers have also reported that immunocompromised transplant recipients with high levels of hepatic RNA expression develop rapidly progressing graft dysfunction; their biopsies reveal only mild inßammation (e.g., presence of lymphocytes). The relative scarcity of intrahepatic lymphocytes in

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HEPATITIS C

these immunosuppressed people and the increased expression of HCV suggest that HCV itself causes liver damage. Nevertheless, biopsy studies have failed to correlate hepatic viral load with biochemical or histological markers of disease activity—possibly because virus-mediated damage occurs only beyond a certain level of intracellular antigen accumulation, which most infected cells may not have reached. CURRENT THERAPIES Current therapies for chronic hepatitis C virus (HCV) aim to reduce or halt progression of liver Þbrosis. Most experts agree that the best method for achieving this goal is through eradication of HCV. Thus, the primary end point for all current therapies for HCV is achieving a sustained virological response (SVR), deÞned as the absence of detectable HCV RNA in polymerase chain reaction (PCR) screens of serum 24 weeks after the conclusion of antiviral therapy. The secondary end points for current HCV therapies are the normalization of serum alanine aminotransferase (ALT) levels and improvements in liver histology, both of which can occur in the absence of an SVR. Studies of the long-term efÞcacy of HCV therapy conÞrm that an SVR is associated with the normalization of serum ALT levels, resolution of liver damage (regression or slowing of Þbrosis), and a low likelihood of relapse of HCV infection (Coverdale SA, 2004). Recent studies suggest that the achievement of SVR signiÞcantly increases survival among chronic HCV-infected individuals (Brok J, 2005; Kasahara A, 2004). Another recent study has shown that the eradication of HCV virus (with peg-IFN-α and ribavirin therapy) reduces the incidence of hepatocellular carcinoma (HCC) in patients who have HCV-related cirrhosis, corroborating the value of SVR as a surrogate end point for efÞcacy and clinical outcome (Hung CH, 2004). The interferon (IFN) family of glycoproteins includes four naturally occurring cytokines that exhibit antiviral, antiproliferative, and immunomodulatory activities: IFN-alpha (α), IFN-beta (β), IFN-gamma (γ ), and IFN-omega (ω). Interferons are produced predominantly by leukocytes in response to viral infection and other biological stimulants. Recombinant or natural IFN-α is the mainstay of HCV treatment in the seven major pharmaceutical markets under study (United States, France, Germany, Italy, Spain, United Kingdom, and Japan), but it has many shortcomings, including limited efÞcacy, signiÞcant side effects, and the need for an extended course of therapy. ModiÞcations to recombinant IFN-α, through covalent attachment of a polyethylene glycol (peg) molecule, substantially changes the drug’s pharmacokinetics, reducing its rate of degradation in the bloodstream and allowing for less frequent administration. Ribavirin is a nucleoside analogue that was approved for hepatitis C in the United States in 1998, in Europe in 1999, and in Japan in 2001. Acting by mechanisms that are not completely understood, the combination of ribavirin and peg-IFN-α elicits an SVR in more than 50% of treated patients (Hadziyannis SJ, 2004). This therapeutic advantage has made once-weekly peg-IFN-α in combination with daily ribavirin the current gold standard for the treatment of HCV.

CURRENT THERAPIES

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Table 5 lists the regimens used to treat chronic hepatitis C; Table 6 describes the mechanism of action for each molecule used to treat HCV; and Table 7 lists current HCV therapies’ key patent expiries. Pegylated-Interferon-Alpha/Ribavirin Regimen Overview. The gold standard for treatment of chronic HCV infection is a combination therapy comprising peg-IFN-α and ribavirin. This regimen has produced the highest efÞcacy of all current therapies, eradicating the virus in nearly 80% of HCV genotype 2- or 3-infected patients and in 40% of patients infected with genotype 1 and 4. Because genotype 1 accounts for nearly 70% of all HCV infections in the United States and Europe, the average efÞcacy of the peg-IFN/ribavirin regimen is estimated at 50–55% (Hadziyannis SJ, 2004). Two forms of peg-IFN-α are available in all seven of the major pharmaceutical markets covered here: • •

Peg-IFN-α-2b (Schering-Plough/Enzon’s PEG-Intron). Peg-IFN-α-2a (Roche/Chugai’s Pegasys).

In mid 2000, Schering-Plough launched peg-IFN-α-2b for the treatment of HCV in the European Union, with the U.S. launch coming shortly afterward in early 2001. Roche launched peg-IFN-α-2a for the treatment of HCV in the European Union in mid 2002 and in the United States in early 2003. Chugai Pharmaceuticals (a member of the Roche group) received approval for peg-IFN-α-2a in Japan in mid 2003, and the molecule was launched soon after. Schering-Plough received approval for peg-IFN-α-2b in Japan in late 2004 and the molecule was launched before the end of that year. Roche and Schering-Plough have been battling for market share, and each company has made signiÞcant attempts to market its peg-IFN-α product as superior among different patient populations and treatment groups. Table 6 describes the mechanism of action of the peg-IFN-α agents. In an effort to boost therapeutic efÞcacy, researchers explored combination therapy of peg-IFN-α with other antiviral or immunomodulatory compounds, and they found success with the addition of ribavirin (Schering-Plough’s Rebetol, Roche’s Copegus). Ribavirin (Figure 3) is a synthetic guanosine nucleoside analogue with a broad spectrum of activity against several RNA and DNA viruses. ICN Pharmaceuticals (now Valeant Pharmaceuticals) developed ribavirin in the early 1970s for aerosol administration in children with respiratory syncytial virus. The company also developed an oral form of ribavirin as an HCV therapy, and the product was initially licensed to Schering-Plough in 1998. Since 2002, Roche has marketed a ribavirin product (Copegus) under license from Valeant, and in 2004 generic ribavirin products entered the market in the U.S. and some European markets. The combination of peg-IFN-α with ribavirin for the treatment of chronic hepatitis C is associated with signiÞcant adverse effects. Peg-IFN-α’s side effects include fatigue, inßuenza-like symptoms, hematologic abnormalities, and neuropsychiatric symptoms (Table 8). Indeed, premature withdrawal from therapy

656 TABLE 5. Current Regimens/Classes Used for Chronic Hepatitis C, 2005 Regimen Components Regimen or

Class

Interferon alphas

Pegylated interferon alphas

Agent

Availability

Dose

Common Toxicities

Interferon-alpha-2b (Schering-Plough’s Intron A)

US, F, G, I, S, UK, J

3 MU tiw for 24–48 weeks

Flulike symptoms Fatigue Anorexia Alopecia Neutropenia Thrombocytopenia Apathy Cognitive changes Irritability Depression

Interferon-alpha-2a (Roche’s Roferon A) Interferon-alpha-con1 (Yamanouchi/Amgen’s Advaferon; Valeant’s Infergen) Pegylated interferonalpha-2b (ScheringPlough’s PEG-Intron)

US, F, G, I, S, UK, J

3 MU tiw for 24–48 weeks 9 µg tiw for 24–48 weeks

US, F, G, I, J

US, F, G, I, S, UK, J

1.5 µg/kg/qw for 24–48 weeks

Flulike symptoms Fatigue Anorexia Alopecia Neutropenia Thrombocytopenia Apathy Cognitive changes Irritability Depression

TABLE 5. (continued) Regimen Components Regimen or

Class

Ribavirin

Agent

Availability

Dose

Pegylated interferonalpha-2a (Roche’s Pegasys) Ribavirin (Schering- Plough’s Rebetol, Roche’s Copegus, generics)

US, F, G, I, S, UK, J

180 µg qw for 24–48 weeks 400–600 mg bid for 24–48 weeks

US, F, G, I, S, UK, J

Interferon- beta

Interferon-beta (Mochida’s IFN-beta; Toray-Daiichi’s Feron)

Ja

3–6 MU qd for 6–12 weeks, then tiw for 6–18 weeks

Ursodeoxy- cholic acid

Ursodeoxycholic acid (Mitsubishi Pharma’s Urso/Ursosan) Glycyrrhizic acid (Minophagen Pharmaceutical’s Stronger Neo-Minophagen C [SNMC])

Jb

600 mg qd for 24 weeks

J

5–20 mL qd for 24 weeks

Glycyrrhizic acid

Common Toxicities

Anemia Anorexia Dyspnea Insomnia Nausea Pharyngitis Pruritis Rash Flulike symptoms Fatigue Anorexia Alopecia Neutropenia Thrombocytopenia Apathy Cognitive changes Irritability Depression Unknown

Unknown

657

a Recombinant IFN-beta is marketed in the United States and Europe for multiple sclerosis. It is not approved for the treatment of hepatitis C. b Ursodeoxycholic acid is marketed in the United States and Europe for cholestatic liver disease and for cholesterol gallstone dissolution. It is not approved for the treatment

of hepatitis C. bid = Twice daily; IFN = Interferon; kg = Kilogram; mcg = Microgram; mg = Milligram; MU = Million units; qd = Once daily; qw = Once weekly; tiw = Three times weekly. US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan.

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HEPATITIS C

FIGURE 3. Structure of ribavirin.

owing to adverse events has been reported in 7–12% of participants in randomized controlled trials, and dose reductions of 50% were required in up to 42% of participants because of adverse events (Table 9) (Fried MW, 2002; Hadziyannis SJ, 2004). The most common reasons for dose modiÞcations are peg-IFN-induced neutropenia and ribavirin-induced anemia. Patients taking peg-IFN-α and ribavirin sometimes require administration of growth factors to counter these adverse effects. Approximately 50% of patients who discontinue therapy do so because of severe depression or other adverse psychiatric events. Dose-reduction and discontinuation-of-therapy rates are often higher with combination therapy than with peg-IFN-α alone. Ribavirin’s side effects include anemia, anorexia, dyspnea, insomnia, nausea, pharyngitis, pruritis, and rash (Khakoo S, 1998). SigniÞcantly, ribavirin causes an acute dose-dependent drop in hemoglobin levels in approximately one-third of patients; this drop in hemoglobin typically occurs within two weeks of initiation of therapy. In approximately 10% of patients, ribavirin causes hemoglobin levels to fall to below 10 g/dL; for this reason, hemoglobin levels of all drug recipients should be monitored closely. Because of ribavirin-induced anemia, patients with cardiovascular disease warrant cautious management and may be contraindicated for combination therapy. Patients with hemoglobinopathies (e.g., thalassemia, sickle-cell anemia) are similarly contraindicated for ribavirin. Ribavirin-induced anemia often mandates dose reductions, which experts believe reduces the efÞcacy of therapy. Animal studies have revealed that ribavirin causes signiÞcant teratogenic and embryocidal effects (Fried MW, 2002); therefore, this agent is contraindicated in pregnant women and in men whose partners are pregnant. For patients and partners of patients taking ribavirin, extreme care must be taken to avoid pregnancy for the duration of therapy and for six months beyond the end of treatment; ribavirin’s product label includes a recommendation that couples employ two forms of contraception. Formulation. The IFN-α products are self-administered subcutaneously, and ribavirin has an oral formulation. A major drawback of natural or recombinant IFN-α is its very short half-life; its serum half-life is only a few hours. As a result,

CURRENT THERAPIES

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TABLE 6. Mechanism of Action by Molecule for Chronic Hepatitis C Virus Molecule Interferon-α

Ribavirin

Ursodeoxycholic acid (UDCA)

Mechanism of Action Experts attribute the antiviral effects of the IFNs to the induction of intracellular enzymes, such as 2
5
oligoadenylate synthetase and protein kinase R, which ultimately cause degradation of viral RNA and inhibit protein synthesis. IFNs also enhance the expression of human leukocyte antigens on the surface of infected cells, resulting in increased recognition of the infected cells by cytotoxic T cells and natural killer cells. IFNα, IFNβ, and IFNω have predominantly antiviral and antiproliferative effects, whereas IFNγ acts as an immunomodulatory mediator. Broad-spectrum antivirals are drugs that work to inhibit the replication of a virus, but do not act directly on the virus itself. Ribavirin is one such drug, and although it was developed more than 30 years ago, the drug’s precise mechanism of action remains unclear and is somewhat controversial. Furthermore, it is not clear which of ribavirin’s reported effects is most important to its efficacy in treating HCV infection. Researchers have proposed that ribavirin’s antiviral activity is exerted by inducing mutations in viral genomes forcing RNA viruses into ‘‘error catastrophe.’’ Other proposed antiviral effects of ribavirin include the following: • Inhibition of host inosine monophosphate dehydrogenase (IMPDH) that leads to depletion of the intracellular supply of guanosine triphosphate (GTP) and ultimately results in the suppression of viral nucleic acid synthesis. • Direct inhibition of the virus-encoded RNA-dependent RNA polymerases, leading to inefficient translation of viral messenger RNA. Scientists also believe that ribavirin can improve the host’s ability to clear viral infection by modulating the CD4+ T-helper-cell response. Research suggests that ribavirin enhances the expression of TH1 cytokines (e.g., IL-2, TNF-α, IFNγ ) and suppresses the expression of TH2 cytokines (e.g., IL-10, IL-4) in cultured T cells (Tam RC, 1999). These immunomodulatory activities may, in part, account for ribavirin’s ability to enhance the antiviral and antifibrotic activities of IFNα. Researchers have shown that the greater efficacy of IFNα/ribavirin combination therapy is associated with ribavirin’s repression of IL-10 production (Cramp ME, 2000). UDCA is a naturally occurring hydrophilic bile acid that is present in small amounts in human bile. It is also an immunomodulator that suppresses the production of IL-1, IL-6, and TNF-α, all of which are potent mediators of inflammation.

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TABLE 6. (continued) Molecule

Mechanism of Action

Glycyrrhizic acid

Glycyrrhizic acid is an aqueous extract from licorice with anti-inflammatory and antiallergic properties. It has been shown to induce interferon to indirectly inhibit virus production and to directly inhibit several viruses in cell culture, including vaccinia, herpes simplex virus, Newcastle disease, and vesicular stomatitis viruses (Abe N, 1982; Pompei R, 1980). Glycyrrhizic acid also inhibits the immunosuppressive effects of cortisone. The necessary activity of glycyrrhizic acid to inhibit HCV-induced liver damage has not yet been elucidated.

TABLE 7. Key Patent and Exclusivity Expiries of Drugs Used to Treat HCV Drug Interferon alpha-2a Interferon alpha-2b Interferon con-1 Interferon alpha-n1 Pegylated interferon alpha-2a Pegylated interferon alpha-2b Interferon beta

United States 21 Dec 2013 21 Dec 2013 13 Dec 2011 21 Dec 2013 Post2014 Post2014 Post2014

France Germany 20 Dec 2013 20 Dec 2013 14 Apr 2013 20 Dec 2013 Post2014 Post2014 28 Jan 2014

20 Dec 2013 20 Dec 2013 14 Apr 2013 20 Dec 2013 Post2014 Post2014 28 Jan 2014

Italy

Spain

20 Dec 2013 20 Dec 2013 14 Apr 2013 20 Dec 2013 Post2014 Post2014 28 Jan 2014

20 Dec 2013 20 Dec 2013 14 Apr 2013 20 Dec 2013 Post2014 Post2014 28 Jan 2014

United Kingdom 20 Dec 2013 20 Dec 2013 14 Apr 2013 20 Dec 2013 Post2014 Post2014 28 Jan 2014

Japan 20 Dec 2013 Post2014 14 Apr 2013 20 Dec 2013 Post2014 Post2014 30 Jul 2012

Note: Our patent and exclusivity expiration dates are based on the latest information from the following sources: FDA Orange Book, IMS Patent Focus, and a review of ongoing news and litigation found in a variety of proprietary sources.

even with daily dosing, troughs occur in the IFN-α serum concentration, making it difÞcult to effectively eradicate HCV infection. Pegylation of IFN-α reduces IFN-α’s accessibility and susceptibility to degradative proteolytic enzymes, which allows for a more sustained concentration of peg-IFN-α in the blood (Bailon P, 2001). As a result, the duration of the drug’s clinical efÞcacy is increased as it provides constant viral suppression, and less frequent administration of the drug is needed. Many factors inßuence the serum half-life of the pegylated protein, but the size of the attached peg molecule plays some role. Peg-IFN-α-2b (Schering-Plough’s PEG-Intron) consists of a 12-kDa linear peg molecule attached to IFN-α-2b. Peg-IFN-α-2b has a large volume of distribution, and dosing is therefore based

CURRENT THERAPIES

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TABLE 8. Summary and Frequency of Treatment-Related Adverse Events Reported in Clinical Trials for Chronic Hepatitis C Trial 1 (Manns MP, 2001)

Trial 2 (Fried MW, 2002)

PEG-Intron plus Ribavirin (%)a

Intron A plus Ribavirin (%)b

Pegasys plus Ribavirin (%)c

Blood disorders Thrombocytopenia Neutropenia Leukopenia Anemia

5 26 6 12

2 14 5 17

5 27 14 11

<1 8 12 11

Flulike symptoms Fever Myalgia (muscle pain) Arthralgia (joint pain) Fatigue/asthenia Rigors

46 56 34 66 48

33 50 28 63 41

41 40 22 65 25

55 49 23 68 37

Gastrointestinal system disorders Diarrhea Weight loss Anorexia Loss of appetite Nausea Abdominal pain Vomiting

22 29 32 32 43 13 14

17 20 27 27 33 13 12

11 10 24 NR 25 8 NR

10 10 26 NR 29 9

Respiratory system disorders Dyspnea (difficulty breathing) Cough Pharyngitis

26 23 12

24 16 13

13 10 NR

14 7 NR

Skin disorders Alopecia Pruritis Rash Dry skin Dermatitis

36 29 24 24 NR

32 28 23 23 NR

28 19 6 10 16

33 18 5 13 13

Central and peripheral nervous system disorders Headache 62 Dizziness 21

58 17

43 14

49 14

Psychiatric disorders Depression Irritability/anxiety/nervousness Insomnia Impaired concentration

31 47 40 17

34 47 41 21

20 33 30 10

28 38 37 13

Resistance mechanism disorders Overall

18

13

12

10

Adverse Event

Intron A plus Ribavirin (%)b

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TABLE 8. (continued) Trial 1 (Manns MP, 2001)

Adverse Event

Trial 2 (Fried MW, 2002)

PEG-Intron plus Ribavirin (%)a

Intron A plus Ribavirin (%)b

Pegasys plus Ribavirin (%)c

75 NR

49 NR

NR 23

Application site disorders Any reaction Injection site reaction

Intron A plus Ribavirin (%)b NR 16

a. PEG-Intron (1.5 mcg/kg once weekly) plus ribavirin (400 mg twice daily) for 48 weeks. b. Intron A (3 MU thrice weekly) plus ribavirin (500–600 mg twice daily) for 48 weeks. c. Pegasys (180 mcg once weekly) plus ribavirin (500–600 mg twice daily) for 48 weeks.

MU = Million units; NR = Not reported. Notes: Adverse events are events reported in more than 10% of patients. Patients may have reported more than one adverse event within a body system category. Full source citations appear in ‘‘References.’’

TABLE 9. Frequency of Discontinuations and Dose Modifications Reported in Clinical Trials for Chronic Hepatitis C Trial 1 (Manns MP, 2001)

Trial 2 (Fried MW, 2002)

Intron A PEG-Intron plus plus Ribavirinb Ribavirina (% patients) (% patients) Discontinuation (total) Adverse event Laboratory abnormality

Intron A plus Ribavirin (% patients)

14

13

10

11

12 2

12 1

7 3

10 1

Pegasys Dose modifications (total) Adverse event Anemia Neutropenia Thrombocytopenia

Pegasys plus Ribavirinc (% patients)

69 42 9 18 NR

55 34 13 8 NR

36 11 1 20 4

Ribavirin

Intron A

46 22 22 1 <1

20 11 3 5 <1

Ribavirin 42 21 19 <1 <1

a PEG-Intron (1.5 mcg/kg once weekly) plus ribavirin (400 mg twice daily) for 48 weeks. b Intron A (3 MU thrice weekly) plus ribavirin (500–600 mg twice daily) for 48 weeks. c Pegasys (180 mcg once weekly) plus ribavirin (500–600 mg twice daily) for 48 weeks.

MU = Million units; NR = Not reported. Full source citations appear in ‘‘References.’’

upon body weight; the drug is typically administered once weekly at 1.0 or 1.5 mcg per kg. Peg-IFN-α-2b has a mean serum half-life of approximately 40 hours (range 22–60 hours). Peg-IFN-α-2b is metabolized predominantly via the liver (approximately 70%); the remaining 30% is eliminated via the kidneys (ScheringPlough Pharmaceuticals, 2003).

CURRENT THERAPIES

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Peg-IFN-α-2a (Roche’s Pegasys) consists of a 40-kDa branched peg molecule attached to IFN-α-2a. Because of its larger size, peg-IFN-α-2a has a small volume of distribution and does not diffuse from the bloodstream as readily as peg-IFNα-2b. Peg-IFN-α-2a is therefore administered at a Þxed dose of 180 µg per week. It has a mean serum half-life of 80 hours (range 50–140 hours), so with weekly administrations, peg-IFN-α-2a levels build up over time. Almost 100% of the drug is metabolized by the liver (Roche Pharmaceuticals, 2005ScheringPlough attempted to overcome patient noncompliance because of the weightbased formulation of peg-IFN-α-2b by offering it in a preÞlled syringe system, called the Redipen. Roche has matched this move by offering preÞlled syringes of peg-IFN-α-2a. The interim results of a comparison trial conducted in Germany were recently reported at the 56th American Association for the Study of Liver Disease (AASLD) annual meeting in November 2005. This study followed 95 patients using both Roche’s preÞlled syringes and Schering-Plough’s Redipen and evaluated the amount of time that physicians spent explaining each system and the problems patients had with self-administration of medication using each system. The study found that patients had problems or difÞculty with 16.0% of drug administrations using Schering-Plough’s Redipen system compared with 2.8% of drug administrations using Roche’s preÞlled syringes (Janisch HD, 2005). Clinical Performance. The regimen of peg-IFN-α in combination with ribavirin is the most effective treatment option for hepatitis C currently available. Table 10 shows clinical trial results for key therapies in a range of patient populations. Schering-Plough conducted a randomized, open-label Phase III clinical trial enrolling 1,530 previously untreated adults with chronic hepatitis C. The results demonstrated the efÞcacy of the peg-IFN-α-2b/ribavirin regimen, in comparison with IFN-α-2b/ribavirin therapy. In this trial, patients were randomly distributed among three treatment groups: • •

•

Peg-IFN-α-2b (1.5 µg/kg once weekly) plus ribavirin (800 mg daily) for 48 weeks. Peg-IFN-α-2b (1.5 µg/kg once weekly) plus ribavirin (1,000 or 1,200 mg daily depending on weight) for four weeks, followed by peg-IFN-α-2b (0.5 µg/kg once weekly) plus ribavirin (1,000 or 1,200 mg daily depending on weight) for 44 weeks. IFN-α-2b (3 MU thrice weekly) plus ribavirin (1,000 or 1,200 mg daily depending on weight) for 48 weeks.

The percentage of patients in each group that had achieved an SVR at the end of the treatment period was 54%, 47%, and 47%, respectively. In patients with genotype 1, the corresponding SVR percentages were 42%, 34%, and 33% in the respective treatment groups; almost 80% of patients with genotypes 2 or 3 had an SVR, regardless of treatment group. Histological improvement was observed in 68% of patients administered peg-IFN-α-2b and 69% of patients receiving IFN-α2b. Secondary analyses identiÞed body weight as an important predictor of SVR in all therapeutic regimens, a Þnding that prompted an additional comparison of

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TABLE 10. Comparison of Key Current Therapies Regimen/ Clinical Trial

Patient Population

Pegylated interferon-α-2a (Pegasys) with ribavirin Fried MP, 2002 1,121 treatment-naive patients Hadziyannis SJ, 2004 PEAK trial

1,311 treatment-naive patients 344 treatment-naive patients

Conjeevaram HS, 196 African-American 2005 treatment-naive, genotype 1 patients

Trial Outcome 56% SVR overall (46% SVR among HCV genotype 1 infected patients) 63% SVR overall (52% SVR among HCV genotype 1 infected patients) Pegasys: week 8 HCV RNA 3.22 log10 reduction from baseline PEG-Intron: week 8 HCV RNA 2.65 log10 reduction from baseline African-American patients: 28% SVR overall Caucasian patients: 50% SVR overall

205 Caucasian treatment-naVve, genotype 1 patients Zeuzem S, 2004 491 treatment-naive adults with persistently normal ALT levels APRICOT 860 HCV/HIV coinfected patients, naive to interferon treatment HALT-C, lead-in 604 patients with advanced phase fibrosis or cirrhosis, previously nonresponsive to conventional interferon-based therapy Sherman M, 2004 120 relapsers to previous interferon-based therapy 241 nonresponders to previous interferon-based therapy

52% SVR overall (40% SVR among HCV genotype 1 infected patients) 40% SVR overall (29% SVR among HCV genotype 1 infected patients) 18% SVR overall

Relapsers: 40% SVR overall Nonresponders: 23% SVR overall

Pegylated interferon-α-2b (PEG-Intron) with ribavirin Manns MP, 2001 1,530 treatment-naive patients 54% SVR overall (42% SVR among HCV genotype 1 infected patients) WIN-R trial 4,913 treatment-naive patients Weight-based ribavirin dosing: 44% SVR overall Flat ribavirin dosing: 41% SVR overall African-American patients: 19% SVR Muir AJ, 2004 100 treatment-naive African-American patients overall (40% EVR at week 12) 100 treatment-naive Caucasian Caucasian patients: 52% SVR overall patients (69% EVR at week 12) Carrat F, 2004 416 HCV/HIV coinfected patients, 27% SVR overall (17% SVR among naVve to interferon treatment HCV genotype 1 infected patients) Taliani G, 2004 141 nonresponders to previous 20% SVR overall interferon-based therapy Interferon-α-consensus (Infergen) with ribavirin Fattovich G, 2003 193 treatment-naive patients

Sjogren MH, 2005 128 treatment-naive patients Chen K, 2005

76 nonresponders to previous interferon-based therapy

HCV genotype 1 infected patients: 38% SVR HCV genotype 2 or 3 infected patients: 67% SVR 57% SVR overall (46% SVR among HCV genotype 1 infected patients) 50% SVR overall

CURRENT THERAPIES

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TABLE 10. (continued) Regimen/ Clinical Trial

Patient Population

Kaiser S, 2004

182 nonresponders to previous interferon-based therapy

Leevy II C, 2004

137 patients who had not experienced an EVR with peg-IFN-α-2b/ribavirin

Leevy II C, 2005

Trial Outcome Conventional interferon nonresponders: 45% SVR overall Pegylated interferon nonresponders: 31% SVR overall African-American patients: 36% SVR overall

Caucasian patients: 49% SVR overall 61 HCV/HIV coinfected patients 30% SVR overall who had not experienced an EVR with peg-IFN-α-2b/ribavirin

Full source citations appear in ‘‘References.’’

the IFN-alpha regimens. When analyzed on the basis of dose per body weight (greater than 10.6 mg/kg of ribavirin daily), the SVR percentages were 61% for all genotypes, 48% for genotype 1, and 88% for genotypes 2 and 3 (Manns MP, 2001). Roche conducted a large Phase III multinational clinical trial to evaluate the efÞcacy of peg-IFN-α-2a/ribavirin combination therapy with peg-IFN-α-2a monotherapy and IFN-α-2b/ribavirin combination therapy. A total of 1,121 previously untreated adults with chronic hepatitis C were randomly assigned to three treatment groups: • • •

Peg-IFN-α-2a (180 µg once weekly) plus ribavirin (500–600 mg twice daily) for 48 weeks. Peg-IFN-α-2a (180 µg once weekly) plus placebo for 48 weeks. IFN-α-2b (3 MU three times weekly) plus ribavirin (500–600 mg daily) for 48 weeks.

Researchers observed an SVR in 56% of patients who received peg-IFN-α2a/ribavirin, compared with only 44% who received IFN-α-2b/ribavirin; only 29% of patients receiving peg-IFN-α-2a monotherapy had an SVR. Of the patients infected with genotype 1, 46% of those receiving peg-IFN-α-2a/ribavirin had an SVR, compared with 36% of those receiving IFN-α-2b/ribavirin and 21% of those receiving peg-IFN-α-2a monotherapy. The corresponding SVR percentages in patients infected with genotype 2 or 3 were 77%, 61%, and 45%, respectively (Fried MW, 2002). This study also established the predictive value of an early virologic response to therapy. Patients with no detectable HCV RNA at 12 weeks were more likely to have an SVR (75%) than patients who had a 2-log10 decrease in HCV RNA at 12 weeks (33%). In contrast, only 3% of patients who had detectable HCV RNA at week 12 had an SVR. Factors consistently identiÞed as adversely affecting the likelihood of achieving an SVR were infection with HCV genotype 1, age 40

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or older, and body weight greater than 75 kg. Peg-IFN-α-2a’s product labeling differentiates usage for different genotypes; genotype 1 patients are expected to receive 48 weeks of therapy and genotype 2 and 3 patients require only 24 weeks of therapy (Fried MW, 2002). In March 2004, investigators published the results of an additional 1,311person clinical trial. The study evaluated the efÞcacy of peg-IFN-α-2a by randomly assigning trial participants into two treatment groups: one arm received peg-IFN-α-2a and the other received nonpegylated interferon-alpha. Both groups received a standard dose of ribavirin. The overall SVR for the peg-IFN-α-2a treatment group was 63%. Interestingly, among HCV-genotype 1 patients given peg-IFN-α-2a plus standard doses of ribavirin for 48 weeks, the SVR was 52%, which is higher than that of any previous study (Hadziyannis SJ, 2004). Roche is conducting an ongoing long-term follow-up study of peg-IFN-α2a (alone or in combination with ribavirin). A total of 896 patients have been enrolled in the study. Eligibility was limited to patients from the registration trials of peg-IFN-α-2a who were HCV-RNA-negative at the conclusion of their original respective trial. Patients are monitored yearly for HCV RNA, for Þve years; 120 patients have completed the Þve years. Interim results were reported at the 56th AASLD annual meeting in 2005. All patients have been followed for at least one year. At this point, only 7 patients have tested detectable levels of HCV RNA (greater than 50 IU/mL serum HCV RNA) after completing treatment. This study demonstrates a greater than 99% durability of SVR in a broad spectrum of patients treated with peg-IFN-α-2a (Swain MG, 2005). The Þnal results of Schering-Plough’s Weight-Based Dosing of PEG-Intron and Rebetol (WIN-R) trial were reported at the 56th AASLD annual meeting. The WIN-R trial was designed to test the efÞcacy of weight-based ribavirin dosing compared with the administration of ribavirin as a ßat dose. Two hundred and twenty-Þve sites recruited a total of 4,913 patients. The trial randomized patients to two separate arms to receive ribavirin as a ßat dose (800 mg/day) or receive ribavirin dosed based on body weight (800, 1,000, 1,200, 1,400 mg/day); all trial participants received peg-IFN-α-2b (1.5 µg/kg/week). HCV-genotype 2 and 3 patients were also randomized to treatment durations of 24 weeks or 48 weeks. All HCV genotype 1 patients received treatment for 48 weeks. At the conclusion of the trial, 44% of patients on weight-based ribavirin dosing achieved an SVR compared with 41% of patients receiving a ßat dose. For HCV genotype 1 patients, 34% achieved an SVR from the weight-based dosing arm, compared with 29% of patients receiving ßat dosing. Extending treatment duration to 48 weeks in genotype 2 and 3 patients did not improve SVR. Relapse rates were also lower with weight-based dosing (15%), compared with ßat dosing (19%) (Jacobson I, 2005). Early results of a head-to-head comparison study between Roche’s peg-IFNα-2a and Schering-Plough’s peg-IFN-α-2b were reported at the 55th AASLD annual meeting in 2004. The Pegasys Early Virologic Response and Kinetics (PEAK) trial enrolled a total of 344 treatment-naive genotype 1 hepatitis C patients randomized into two treatment groups receiving standard doses of either peg-IFN-α-2a or peg-IFN-α-2b in combination with equal doses of ribavirin.

CURRENT THERAPIES

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The study aims to evaluate the early viral load reductions at week 12 of therapy, which has been shown to be highly predictive of attaining an SVR. At week 8 of therapy, the mean HCV-RNA decrease for the peg-IFN-α-2a group was 3.22 log10 compared with 2.65 log10 for the peg-IFN-α-2b group (Di Bisceglie AM, 2004). Final results are expected by the end of 2006. Schering-Plough is currently enrolling patients in another head-to-head clinical trial with rival Roche’s pegylated IFN-α-2a product. The Individualized Dosing EfÞcacy vs. Flat Dosing to Assess Optimal Pegylated Interferon Therapy (IDEAL) trial compares standard peg-IFN-α-2b dosing (1.5 µg/kg) and weightbased ribavirin dosing with (1) the peg-IFN-α-2b dose ((1.0 µg/kg) plus weightbased ribavirin dosing and (2) the approved Þxed doses of Roche’s ribavirin and peg-IFN-α-2a. Schering-Plough added the third arm with Roche’s products to its prior Phase IV commitment to study the Þrst two combinations. The IDEAL trial aims to enroll 2,880 treatment-naive, genotype 1 patients and will consist of 48 weeks of treatment with 24 weeks of follow-up. Results from this trial are expected in early 2007. African-American hepatitis C patients typically experience lower treatment success rates than HCV patients from other ethnic groups. A recently published study compares the efÞcacy of chronic HCV treatment between AfricanAmericans and non-Hispanic Caucasians. One hundred African-American and 100 non-Hispanic Caucasian treatment-naive patients were given peg-IFN-α-2b (1.5 µg/kg for 48 weeks) and ribavirin (1,000 mg/day for 12 weeks followed by 800 mg/day for 36 weeks). To control for genotype, researchers ensured that 98% of all patients in the trial had genotype 1. At 12 weeks of therapy, only 40% of African-American patients had an early virological response (EVR), deÞned as a 2 log10 IU/mL reduction in viral HCV RNA compared with 69% in the non-Hispanic white group. An EVR had been shown to be predictive of an SVR, and in this study 48% of the African-American patients with an EVR had an SVR compared with 75% of the non-Hispanic white patients. At the conclusion of the trial, 19% of African-American and 52% of Caucasian patients had achieved an SVR. Thus, it seems that factors other than viral genotype contribute to the poor response rates to current HCV therapies observed among African-Americans (Muir AJ, 2004). The results of a study examining peg-IFN-α-2a/ribavirin treatment in AfricanAmericans were recently reported at the 56th AASLD annual meeting in 2005. The study enrolled 196 African-Americans and 205 Caucasian Americans; all patients were infected with HCV-genotype 1 and were treatment-naive. Patients all received peg-IFN-α-2a (180 µg/week) and ribavirin (1,000 or 1,200 mg/day, based on body weight) for 24 weeks. Patients who tested HCV-RNA-negative at 24 weeks continued on therapy for 48 weeks, while patients who were HCVRNA-positive at 24 weeks were considered nonresponders to therapy. At 72 weeks, the SVR was 28% for African-Americans and 50% for Caucasian Americans (Conjeevaram HS, 2005). Roche is conducting additional clinical trials to broaden its indication labeling and use of peg-IFN-α-2a. One recently published trial involved 491 patients

668

HEPATITIS C

with chronic hepatitis C and persistently normal ALT levels. The trial found that although the enrolled patients had normal ALT levels, one-fourth to one-third of them had some degree of liver Þbrosis. Patients in the two treated groups received the standard doses of peg-IFN-α-2a (180 µg/week) and ribavirin (800 mg/daily) for either 24 or 48 weeks. The control group received placebo. Results from this trial showed that 52% of patients treated for 48 weeks achieved an SVR, while no patients in the placebo group achieved an SVR. Seventy-two percent of treated genotype 2 and 3 patients achieved an SVR after 24 weeks of therapy, compared with 40% of treated genotype 1 patients after 48 weeks of therapy (Zeuzem S, 2004). These data suggest the potential advantages of treatment in patients with normal ALT levels. An additional expansion of peg-IFN-α-2a’s label was achieved in February 2005. Roche announced that the FDA had approved peg-IFN-α-2a/ribavirin combination therapy for the treatment of HCV/HIV-coinfected patients. The randomized, single-blind AIDS Pegasys Ribavirin International Coinfection Trial (APRICOT) demonstrated the efÞcacy of peg-IFN-α-2a/ribavirin treatment in patients coinfected with HCV and HIV. The study enrolled 860 patients (85% of whom received antiretroviral therapy) who were divided into three groups and given standard doses of IFN-α-2a/ribavirin, peg-IFN-α-2a alone, or peg-IFN-α2a/ribavirin for 48 weeks. Forty percent of the patients treated with peg-IFN-α2a/ribavirin achieved an SVR, and their ability to take antiretroviral therapy for their HIV infection was not compromised. In contrast, only 20% of the patients taking peg-IFN-α-2a alone and 12% of the IFN-α-2a/ribavirin patients achieved an SVR. For patients infected with genotype 1, the SVR rates were 29% for the peg-IFN-α-2a/ribavirin treatment group, 14% for the peg-IFN-α2a monotherapy group, and 7% for the IFN-α-2a/ribavirin group; for genotype 2 and 3 patients, the respective rates were 62%, 36%, and 20% (Torriani FJ, 2004). Another recently published clinical trial investigated the use of ScheringPlough’s peg-IFN-α-2b/ribavirin in patients coinfected with HCV and HIV. This randomized, open-label trial divided 416 patients into two treatment groups. The Þrst group received standard doses of peg-IFN-α-2b while the second group received standard doses of nonpegylated IFN-α-2b. Both groups also received 800 mg of ribavirin daily. An SVR was obtained in 27% of the peg-IFN-α2b/ribavirin-treated patients compared with 20% of the nonpegylated IFN-α2b/ribavirin-treated patients. For patients infected with genotype 1, the SVR was 17% for the peg-IFN-α-2b/ribavirin group and 6% for the nonpegylated IFN-α-2b/ribavirin group; for genotype 2 and 3 patients, the SVRs were not signiÞcantly different (44% and 43%, respectively). For patients Þnishing at least 80% of treatment, the SVRs were 40% for the peg-IFN-α-2b/ribavirin-treated patients and 29% for the standard IFN/ribavirin-treated patients (Carrat F, 2004). Investigators are studying the efÞcacy of peg-IFN-α-2a/ribavirin treatment in nonresponder patients (those who have failed Þrst-line therapy) in the leadin phase of the Hepatitis C Antiviral Long-term Treatment against Cirrhosis (HALT-C) trial. The HALT-C trial consists of two phases; during the Þrst phase

PEGYLATED INTERFERON-ALPHA, SINGLE AGENT

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(or “lead-in” phase) patients received peg-IFN-α-2a (180 µg once weekly) and ribavirin (1,000 mg/day or 1,200 mg/day for those weighing over 75 kg) for 24 weeks. If patients responded to therapy at week 20, they continued treatment for a total of 48 weeks. Patients who did not respond are entered into the second phase of the HALT-C trial, which is designed to evaluate the efÞcacy of continuing lowdose peg-IFN-α-2a therapy in patients who fail to respond to initial therapy to suppress HCV replication and provide liver protective effects (Halt-C trial Web site, 2005). In 2004, M.L. Shiffman reported preliminary results for 604 patients who had advanced Þbrosis or cirrhosis and were previously nonresponsive to conventional interferon-based therapy (either IFN-α monotherapy or IFN-α in combination with ribavirin). In the lead-in phase of the HALT-C trial, patients received pegIFN-α-2a (180 µg once weekly) and ribavirin (1,000 mg/day or 1,200 mg/day for those weighing over 75 kg) for 24 weeks. Thirty-Þve percent of patients had a response to therapy (HCV-RNA-negative) at 20 weeks (35%) and continued therapy for a total of 48 weeks. At 72 weeks, the SVR for this group was 18% (Shiffman ML, 2004). This result demonstrates the efÞcacy and feasibility of peg-IFN-α-2a/ribavirin treatment in patients who previously failed conventional interferon-based therapy. The results of another study evaluating the efÞcacy of peg-IFN-α-2a/ribavirin therapy in nonresponders and relapsers were presented at the 55th AASLD annual meeting in 2004. One hundred and twenty relapsers and 241 nonresponders to previous interferon-based therapy (either monotherapy or combination therapy with ribavirin) were enrolled. Patients were given either a 24- or 48-week course of peg-IFN-α-2a/ribavirin combination therapy and were monitored for 24 weeks after treatment. The SVR was 40% for the relapser patient group and 23% for nonresponders, demonstrating a beneÞcial effect of peg-IFN-α-2a/ribavirin in previous nonresponders and relapsers to therapy (Sherman M, 2004). Schering-Plough reported the results of a similar study to evaluate the efÞcacy of peg-IFN-α-2b/ribavirin in nonresponder patients at the 55th AASLD annual meeting in 2004. One hundred and forty-one patients who had failed interferon/ribavirin combination therapy were enrolled in this trial and were treated with standard doses of peg-IFN-α-2b and ribavirin for 48 weeks. The SVR for these nonresponders was 20%, which is comparable to similar studies examining the efÞcacy of peg-IFN-α-2a and ribavirin in the same population (Taliani G, 2004). PEGYLATED INTERFERON-ALPHA, SINGLE AGENT Overview Pegylated-interferon-alpha (peg-IFN-α) treatment has long been the standard of care for HCV. Although the current standard of care involves pegylatedinterferon-alpha combination therapy with ribavirin, many patients are contraindicated for ribavirin treatment and will resort to pegylated-interferon-alpha monotherapy. Two forms of pegylated IFN-α are available:

670 • •

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Peg-IFN-α-2b (Schering-Plough’s PEG-Intron) *. Peg-IFN-α-2a (Roche’s Pegasys) *.

The launch histories of the peg-IFN products are described in the previous section, “Pegylated-Interferon-Alpha/Ribavirin Regimen,” and the mechanism of action of peg-IFN-α is shown in Table 6. Formulation. The formulation of the peg-IFN products is discussed in the previous section, “Pegylated-Interferon-Alpha/Ribavirin Regimen.” Clinical Performance. In August 2001, Schering-Plough reported results from a multinational Phase III trial comparing the safety and efÞcacy of peg-IFN-α-2b with IFN-α-2b as monotherapy for the treatment of HCV infection. The trial randomly assigned 1,219 treatment-naive HCV patients with elevated ALT and liver histopathology consistent with chronic HCV to one of four treatment groups. All patients were treated for 48 weeks. Three groups were treated with peg-IFNα-2b (0.5, 1.0, or 1.5 µg/kg) once weekly, and one group was assigned standard IFN-α-2b (3 MU, thrice weekly). A sustained virological response occurred in 18%, 25%, 23%, and 12% of the patients. All three peg-IFN-α-2b doses improved liver histology to a greater extent than did IFN-α-2b, particularly in subjects who achieved an SVR (Lindsay KL, 2001). Researchers have reported results from two Phase III clinical trials evaluating the clinical efÞcacy of peg-IFN-α-2a monotherapy in treatment-naive patients. In one study with 630 HCV patients (20% with cirrhosis or bridging Þbrosis), an SVR was reported in 38% of patients receiving 180 µg of peg-IFN-α-2a once weekly for 48 weeks, compared with 19% of patients receiving IFN-α-2a (6 MU three times a week for 12 weeks, followed by 3 MU three times a week for 36 weeks) (Zeuzem S, 2000). A second study, involving 269 patients with cirrhosis or bridging Þbrosis, found that treatment with 180 µg of peg-IFN-α-2a once weekly for 48 weeks was effective in 30% of patients, compared with an 8% rate of SVR in patients receiving standard IFN-α-2a (Heathcote EJ, 2000). Schering-Plough is currently enrolling patients in a randomized, open-label Phase III trial of long-term low-dose peg-IFN-α-2b in comparison to long-term therapy with colchicine (Clincaltrials.gov Web site, 2005). Colchicine has not been clinically tested in HCV but has been shown to improve liver function in cirrhotic patients (Kaplan MM, 1999). Long-term maintenance therapy is based on the observation that up to 40% of nonresponders to IFN-α therapy have a histological response during treatment; researchers hope that continuing treatment with IFN-α will prevent Þbrosis and progression to liver complications even if the virus is not cleared. The objective of the multinational Colchicine versus PEG-Intron Long-Term (COPILOT) study is to evaluate the safety and efÞcacy of peg-IFN-α-2b for the prevention of disease progression in adults with compensated cirrhosis secondary to chronic hepatitis C who have failed to respond to previous combination therapy. Researchers reported interim results at the 55th AASLD annual meeting in 2004. The study enrolled 534 chronic HCV patients with advanced cirrhosis who had failed previous IFN-α therapy.

PEGYLATED INTERFERON-ALPHA, SINGLE AGENT

671

Two hundred and seventy patients were randomly assigned to receive peg-IFNα-2b (0.5 µg/kg once weekly) while 264 patients received colchicine (0.6 mg twice daily). Low-dose maintenance peg-IFN-α-2b treatment demonstrated superior protective efÞcacy to colchicine with almost twice as many patients in the colchicine arm having reached a clinical end point (15% versus 7.5%), signifying a complication of cirrhosis (Afdahl N, 2004). In June 2000, the National Institutes of Health (NIH) began enrolling patients in the HALT-C trial to examine the safety and efÞcacy of long-term therapy with peg-IFN-α-2a. The study was designed to determine whether continuing low-dose peg-IFN-α-2a therapy in patients who fail to respond to combination therapy is sufÞcient to suppress HCV replication, avert progression to cirrhosis, prevent liver cancer, and reduce the need for liver transplantation. The lead-in phase of the HALT-C trial consists of treating nonresponder patients with combination therapy. Patients who are HCV-RNA-positive 20 weeks into this second round of treatment are enrolled in the HALT-C trial. Enrollment and randomization of more than 1,000 patients in the HALT-C trial were completed in August 2004. Patients were randomized to receive either peg-IFN-α-2a or no treatment for 3.5 years. Final results are not expected until 2008 (Halt-C trial web site, 2005). The results from a smaller randomized, controlled trial examining the beneÞt of peg-IFN-α-2a monotherapy in chronic HCV patients with advanced Þbrosis or bridging cirrhosis were presented at the 55th AASLD annual meeting in 2004. Data from 183 patients treated in three separate study arms were evaluated. Patients received either 180 µg of peg-IFN-α-2a weekly, 90 µg of peg-IFN-α-2a weekly, or 3 MU of IFN-α-2a three times weekly. End points of the trial were deÞned as improvement in Þbrosis (greater than or equal to one stage on the Metavir scale for evaluation of liver Þbrosis) or improvement in inßammatory grade (greater than or equal to one Metavir stage) from baseline. An SVR was obtained in 37%, 16%, and 9% of participants in the high-dose peg-IFN, lowdose peg-IFN, and IFN groups, respectively. Fibrosis improvements were seen in 35%, 25%, and 27% of patients in the high-dose peg-IFN, low-dose peg-IFN, and IFN groups, respectively (Everson G, 2004). Adverse events are more common with higher doses and prolonged durations of IFN-α treatment. Adverse events observed with peg-IFN-α were similar to those reported during trials and treatment with unmodiÞed versions of IFN-α. However, a higher proportion of patients taking peg-IFN-α did experience an adverse event compared with those taking unmodiÞed IFN-α (Fried MW, 2002; Hadziyannis SJ, 2004; Manns MP, 2001). SigniÞcantly, the peg-IFN-α induced a dose-dependent reduction in thrombocytes and neutrophils more often than standard IFN-α. Clinical trial researchers note that depression and ßulike symptoms occur less frequently in patients taking peg-IFN-α-2a than in patients taking peg-IFN-α-2b. Interferon-Alpha-Consensus/Ribavirin Regimen Overview. Interferon alpha-con-1 (IFN-α-con-1) (Valeant’s Infergen) is a modiÞed recombinant interferon that is generated using a consensus nucleic acid

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HEPATITIS C

sequence. IFN-α-con-1 is marketed in the United States, France, Germany, Italy, and Japan. Ribavirin is a synthetic guanosine nucleoside analogue with a broad spectrum of activity against several RNA and DNA viruses. It has been shown to augment the therapeutic efÞcacy of interferon-alpha treatment of HCV. Its mechanism of action is described in Table 6. Clinical Performance. The efÞcacy of IFN-α-con-1 in combination with ribavirin as an initial treatment for chronic hepatitis C was examined in an openlabel, multicenter, randomized clinical trial enrolling 193 treatment-naive patients. Patients were randomized into two groups and received either 9 mcg or 18 mcg of subcutaneously administered IFN-α-con-1 three times weekly in combination with either 1,000 or 1,200 mg of ribavirin (based on body weight) taken orally. Genotype 1-infected patients were treated for a total of 48 weeks while genotype 2- or 3-infected patients were treated for 24 weeks. At the 24-week post-treatment follow-up, the percentages of patients achieving an SVR were 67% for the genotype 2 and 3 patients and 38% for genotype 1 patients (Fattovich G, 2003). The study concluded that a higher dose of IFN-α-con-1 does not increase SVR rates. Another open-label, multicenter, randomized trial enrolled 128 patients who were randomized into two treatment groups. The Þrst group received 15 mcg of IFN-α-con-1 three times weekly while the comparator group received 3 MU of IFN-α-2b three times weekly. Both groups received 1,000 mg of ribavirin daily, and treatment lasted 48 weeks. At 24 weeks post-treatment, 57% of patients treated with IFN-α-con-1 had achieved an SVR compared with 40% in the IFNα-2b group. In a subset of patients with genotype 1 and a high initial viral load, those patients taking IFN-α-con-1/ribavirin fared much better (46% achieved an SVR) than the IFN-α-2b/ribavirin group (14% achieved an SVR). Among all patients with an initial high viral load, 57% of IFN-α-con-1/ribavirin-treated and 31% of IFN-α-2b/ribavirin-treated patients achieved an SVR (Sjogren MH, 2005). Adverse events were similar in both treatment groups. Another study reported at the 56th AASLD annual meeting evaluated IFNα-con-1 to treat previous nonresponders to both peg-IFN-α/ribavirin and IFNα/ribavirin combination therapies. The study randomized 76 patients to receive IFN-α-con-1 (15 mcg/day) in combination with ribavirin (800–1,200 mg/day, based on patient body weight). Patients were treated for 48 weeks, 80% of patients were HCV genotype 1 and 47% of patients had Metavir Þbrosis scores of F3/F4 (see “Etiology and Pathophysiology”). At 72 weeks, 50% of the recruited patients achieved SVR (Chen K, 2005). This trial demonstrates the utility of IFN-αcon-1 as a potential alternative for nonresponders to peg-IFN-α/ribavirin and IFN-α/ribavirin combination therapies. Results of another study evaluating the use of IFN-α-con-1 to treat previous nonresponders to both peg-IFN-α/ribavirin and IFN-α/ribavirin combination therapies were reported in 2004 at the 55th AASLD annual meeting. The study enrolled 182 patients (92% genotype 1) who had previously failed combination therapy. Patients were initially treated with 27, 18, or 9 µg of IFN-α-con-1 daily for 2 weeks, followed by 9 µg IFN-α-con-1 daily for 4 weeks, after which treatment was continued at 9 µg of IFN-α-con-1 daily with 10–15 mg/kg ribavirin

PEGYLATED INTERFERON-ALPHA, SINGLE AGENT

673

daily for 36 weeks. At 24 weeks post-treatment, 38–45% of IFN-α-con-1-treated conventional IFN-α/ribavirin nonresponders had an SVR while 27–31% of IFNα-con-1-treated peg-IFN-α/ribavirin nonresponders had an SVR. Tolerability of this regimen was comparable to peg-IFN-α/ribavirin therapy with the exception of the 27 µg treatment group during the two-week induction period, which was reported as less tolerable (Kaiser S, 2004). Results of a separate study reported at the 55th AASLD annual meeting in 2004 showed promise to the African-American nonresponder population. One hundred and thirty-seven patients being treated with peg-IFN/ribavirin combination therapy who had not experienced an EVR after 12 weeks of therapy were immediately transferred onto a regimen of 15 mcg of IFN-α-con-1 daily with 1,000–1,200 mg ribavirin for 12 weeks. The SVR for African-Americans in this study was 36% at 36 weeks post-therapy compared with 49% of non-African-American participants (Leevy II C, 2004). This response is a substantial improvement over other currently available therapies for this typically low-responding segment of the HCV patient population. A study to determine the efÞcacy of IFN-α-con-1 and ribavirin in HIV/HCVcoinfected patients recruited treatment-naive HIV/HCV-coinfected patients who were initially treated with peg-IFN-α-2a and ribavirin for 12 weeks. Patients who did not achieve an EVR after 12 weeks of treatment were immediately transferred to an IFN-α-con-1 treatment group totaling 61 patients, 92% of whom were HCVgenotype 1. Patients received IFN-α-con-1 (15 mcg/day) and ribavirin (weightbased daily dosing) for 72 weeks. At the end of treatment, 31% of patients were HCV-RNA-negative, and at 96 weeks, 30% of coinfected patients had achieved an SVR (Leevy II C, 2005). These results demonstrate the potential utility of IFN-α-con-1 as a second-line therapy for HCV patients coinfected with HIV. Interferon-Beta, Single Agent Overview. Japanese physicians have used a wider variety of therapies than their Western counterparts, with a greater focus on controlling liver damage and preventing progression to more serious liver complications in chronic HCV patients. In Japan, physicians use natural IFN-β for the treatment of chronic hepatitis C, often as induction therapy, before prescribing IFN-α-based therapy. Clinical Performance. Physicians typically use IFN-β (Mochida’s IFN-β, Toray/Daiichi-Sankyo’s Feron) as part of a high-dose induction therapy consisting of 6 MU of IFN-β daily for 6 weeks, followed by 3 MU IFN-α three times weekly for a total of 24 weeks. In comparing the efÞcacy of IFN-β induction therapy (n = 50) with IFN-α monotherapy (n = 49), researchers report that although IFN-β induction therapy achieved more rapid viral clearance than IFNα monotherapy, no statistically signiÞcant difference in the SVR rates between these two groups (18% and 6%, respectively) was observed (Watanabe H, 2002). The results of a trial to compare the efÞcacy of recombinant IFN-β-1a (Biogen’s Avonex, Serono’s Rebif) alone or in combination with ribavirin in chronic hepatitis C patients were recently published. This randomized trial recruited 102

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HEPATITIS C

treatment-naive patients in multiple centers in Italy. Patients were split into two treatment groups and received 6 MU of recombinant IFN-β-1a daily alone or with 1,000–1,200 mg of ribavirin for 24 weeks. The study did not show a statistically signiÞcant difference between the percentages of patients in the monotherapy and combination treatment groups achieving an SVR (21.6% and 27.4%, respectively) (Pellicano R, 2005). In the largest trial reported to date for natural IFN-β, researchers investigated the efÞcacy of 6 MU of natural IFN-β administered daily for 12 weeks, followed by thrice-weekly IFN-β for a subsequent 14 weeks in patients with high viral load (greater than 6 million copies/mL). An SVR was achieved in 88% (22 of 25) of patients with a low viral load and in 22% (19 of 87) of patients with a high viral load, corresponding to an overall SVR proportion of 37%. Patients infected with HCV genotype 2a (a subset of HCV genotype 2) were more likely to respond to therapy than were patients infected with genotype 1; an SVR was observed in 67% and 21% of patients, respectively (Shiratori Y, 2000). Ursodeoxycholic Acid, Single Agent Overview. Japanese physicians report widespread use of ursodeoxycholic acid (UDCA) to lower ALT levels and prevent progression of liver damage. Japanese physicians commonly prescribe UDCA (Mitsubishi Pharma’s Urso) alone as second- or third-line therapy and, less frequently, in combination with IFN-alpha as Þrst- or second-line therapy. Clinical Performance. Japanese physicians give UDCA at a dose of 600 mg daily for at least six months and sometimes for as long as several years. Although this drug is also available in the United States and Europe, physicians in these markets do not report prescribing UDCA as a treatment for hepatitis C; instead, it is used for cholestatic liver disease and cholesterol gallstone dissolution. UDCA predates IFN-based HCV treatment in Japan by several decades. Several trials have shown that when prescribed as monotherapy, UDCA reduces serum ALT levels in patients with chronic hepatitis C but has no effect on HCV RNA levels. Clinical trials evaluating the efÞcacy of combination therapy of UDCA plus IFN-α from the mid to late 1990s have been similarly inconclusive; in some instances, the addition of UDCA to treatment regimens has generally not signiÞcantly affected likelihood of SVR, but it has delayed relapse and, in some cases, produced histological beneÞts (Angelico M, 1995; Boucher E, 1995; Kiso S, 1997; Mazur W, 2001; Senturk H, 1997). A recent study has shown that combination therapy comprising IFN-α, ribavirin, and UDCA reduces serum ALT and results in transient lowering of HCV viral load. However, six months after cessation of therapy, both ALT and viral load levels had elevated, providing evidence of a viral relapse (Ljubuncic P, 2005). When prescribed as second- or third-line therapy following nonresponse to or relapse after IFN-α therapy, UDCA is used to slow the progression of disease. Although clinical trials evaluating the efÞcacy of UDCA in patients previously treated with IFN-α have shown some improvements in serum ALT levels, the

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long-term beneÞts of this regimen have not been established (Fabbri C, 2000; Lirussi F, 1999; Porcaro G, 2000; Poupon RE, 2000). Glycyrrhizic Acid, Single Agent Overview. Japanese physicians report the use of glycyrrhizic acid to lower ALT levels and prevent progression of liver damage. Physicians in Japan have used glycyrrhizic acid to lower ALT levels in chronic liver diseases since 1975, when it was Þrst demonstrated to reduce serum ALT concentrations. Like UDCA, glycyrrhizic acid does not reduce viral load and must be prescribed long term because ALT levels will rise when treatment is discontinued, indicating that glycyrrhizic acid does not actually address the root causes of elevated ALT. Clinical Performance. Glycyrrhizic acid (Minophagen Pharmaceutical’s Stronger Neo-Minophagen C [SNMC]) is administered by injection at doses ranging from 5 to 100 mL/day. Like UDCA, it is prescribed as second- or third-line therapy to reduce ALT levels. Researchers report that long-term therapy with glycyrrhizic acid reduces the risk of cirrhosis and HCC in patients with chronic hepatitis. In 1997, a retrospective study on the long-term effects of glycyrrhizic acid found that HCV-positive patients treated with glycyrrhizic acid were 2.5 times less likely to develop HCC than HCV-positive patients who did not receive glycyrrhizic acid (Arase Y, 1997). Of the 453 patients diagnosed with chronic hepatitis C between January 1979 and April 1984, 84 had been treated with glycyrrhizic acid, administered at 100 mL daily for 8 weeks, then two to seven times a week for 2–16 years (median, 10.1 years). After 15 years, only 12% had developed HCC. In comparison, of the 109 patients who could not be treated with glycyrrhizic acid for a long period of time (median, 9.2 years), 25% developed HCC by year 15. Further analysis revealed that of 178 patients on long-term glycyrrhizic acid therapy, only 50 (28%) developed cirrhosis after 13 years. In comparison, 40% of the 100 control patients developed cirrhosis after 13 years (Kumada H, 2002).

EMERGING THERAPIES Overview The combination of pegylated interferon-alpha (peg-IFN-α) plus ribavirin achieves a sustained virological response (SVR) in 40–50% of patients infected with the hepatitis C virus (HCV) genotype 1 and is effective in approximately 80% of patients infected with HCV genotype 2 or 3. Nevertheless, because genotype 1 accounts for a large proportion of HCV infections, overall treatment response for HCV is suboptimal. Additionally, a signiÞcant number of patients cannot be treated or cannot tolerate IFN-α or ribavirin for various reasons (e.g., toxicity proÞle, contraindications). Several second-generation nucleoside analogues and inhibitors of inosine-5’-monophosphate dehydrogenase (IMPDH)

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are in development and are intended to have a more favorable side-effect proÞle than that of ribavirin. Researchers are also testing numerous interferons, immunomodulators, and protease and polymerase inhibitors in an effort to identify novel mechanisms of action that may increase the efÞcacy of HCV therapy or at least provide second-line alternatives to slow disease progression in patients who fail initial therapy. Most of the limitations associated with nucleotide analogues (i.e., ribavirin) and immunomodulators (e.g., IFN-α) stem from their nonspeciÞc mechanism of action. For example, ribavirin targets host-cell pathways that facilitate viral replication, while IFN-α activates a systemic antiviral immune response. Neither mechanism of action is particularly speciÞc for HCV, and both drugs have been used to combat other important diseases (e.g., inßuenza, hepatitis B, multiple sclerosis). Drug developers hope to improve safety and efÞcacy by developing HCV-speciÞc inhibitors. However, the investigational techniques that facilitate discovery of HCV-associated drug targets remain in early stages of development. Although the pipeline for HCV drug development is more heavily skewed toward preclinical development, several highly anticipated HCV-speciÞc antiviral candidates are in advanced stages of clinical trials, and thought leaders believe that some of the novel HCV-speciÞc antivirals (e.g., polymerase inhibitors and protease inhibitors) have the potential to revolutionize HCV therapy. HCV replication may be inhibited by targeting any one of the several nonstructural HCV proteins involved in viral replication, including the HCV NS3 serine protease, the NS3 helicase, and the NS5B RNA-dependent RNA polymerase (RdRp). Although signiÞcant effort has been put into identifying speciÞc inhibitors of the HCV NS3 helicase, a prominent target owing to its critical role in viral replication, few drug candidates have emerged to date. Eradication of chronic HCV infection may also be achieved by preventing the translation of viral proteins (translation initiation inhibitors) or by inhibiting entry of the virus into the host cell (antibody therapies). These approaches may effectively suppress viremia and the ultimate progression to chronic liver disease, cirrhosis, or hepatocellular carcinoma (HCC). Researchers are also developing therapeutic and prophylactic vaccines that may be effective in treating and preventing HCV infection, respectively. Table 11 lists emerging therapies in development, and Table 12 lists companies with emerging HCV therapies by class and phase. Interferons Overview. Current treatment consists of drugs that, in part, boost the overall immune response in patients with hepatitis C in an attempt to clear the infection. The natural immune response to viral infection often employs the IFN family, which includes four naturally occurring cytokines that exhibit antiviral, antiproliferative, and immunomodulatory activities: IFN-alpha (α), IFN-beta (β), IFN-gamma (γ ), and IFN-omega (ω). These glycoproteins are produced predominantly by leukocytes in the blood in response to viral infection and other

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TABLE 11. Emerging Therapies in Development for Hepatitis C Virus, 2005 Compound

Development Phase

Marketing Company

Interferons Interferon-omega United States Europe Japan

II II —

Intarcia Intarcia —

Albuferon-alpha United States Europe Japan

II II —

Human Genome Sciences Human Genome Sciences Human Genome Sciences

III III II

SciClone Pharmaceuticals Sigma Tau Schering-Plough KK

Isatoribine United States Europe Japan

I — —

Anadys Anadys Anadys

ANA-975 United States Europe Japan

— I —

Anadys/Novartis Anadys/Novartis Anadys/Novartis

CPG-10101 United States Europe Japan

I — —

Coley Pharmaceutical Coley Pharmaceutical —

II II —

Vertex Pharmaceuticals Vertex Pharmaceuticals —

III III —

Valeant Valeant —

II II —

Idenix/Novartis Idenix/Novartis Idenix/Novartis

HCV-796 United States Europe Japan

I — —

ViroPharma/Wyeth Wyeth Wyeth

R-1626 United States

—

Roche

Non-interferon immunomodulators Thymosin-alpha-1 United States Europe Japan

Nucleoside analogues and IMPDH inhibitors Merimepodib United States Europe Japan Viramidine United States Europe Japan HCV polymerase inhibitors Valopicitabine (NM-283) United States Europe Japan

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TABLE 11. (continued) Compound Europe Japan

Development Phase

Marketing Company

I —

Roche Roche

PC — —

Pharmasset/Roche Pharmasset/Roche Pharmasset/Roche

II I —

Vertex Pharmaceuticals Vertex Pharmaceuticals Mitsubishi Pharma

II II —

Schering-Plough Schering-Plough —

I — —

Gilead Sciences — —

III III —

Endo, generics Generics —

Celgosivir United States Europe Japan

II — —

Migenix — —

Antibody therapies XTL-6865 United States Europe Japan

I — —

XTL Biopharmaceuticals XTL Biopharmaceuticals —

Civacir United States Europe Japan

I/II I —

Nabi Biopharmaceuticals Nabi Biopharmaceuticals —

HuMax-HepC United States Europe Japan

PC PC —

Genmab Genmab —

HCV vaccines IC-41 United States Europe Japan

— II —

— Intercell —

PSI-6130 United States Europe Japan HCV protease inhibitors VX-950 United States Europe Japan SCH-503034 United States Europe Japan GS-9132 United States Europe Japan Broad-spectrum antivirals Amantadine United States Europe Japan

HCV ISCOM

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TABLE 11. (continued) Compound United States Europe Japan HCV E1E2/MF59C.1 United States Europe Japan

Development Phase

Marketing Company

PC — —

Chiron/CSL Chiron/CSL —

I — —

Chiron Chiron —

PC = Preclinical (including discovery); S = Suspended. Note: Development phase is based on databases such as R&D Focus, R&D Insight, Pharmaprojects, and the Investigational Drugs Database (IDdb3); periodicals such as Scrip, the FDC’s Pink Sheet,and Marketletter; company reports and press releases; and industry contacts.

biological stimulants. Researchers are working on novel formulations of IFN-α as well as IFN-ω that they hope will have longer half-lives and/or greater efÞcacy than currently used IFN-αs. Mechanism of Action. Experts attribute the antiviral effects of immunomodulators currently used in HCV treatment, IFN-αs, to the induction of intracellular enzymes, such as 2
5
oligoadenylate synthetase (OAS) and protein kinase R (PKR), which ultimately cause degradation of viral RNA and inhibit protein synthesis. IFNs also enhance the expression of human leukocyte antigens (HLAs) on the surface of infected cells, resulting in increased recognition of the infected cells by cytotoxic T cells and natural killer cells. IFN-α, IFN-β, and IFN-ω have predominantly antiviral and antiproliferative effects, whereas IFN-γ acts as an immunomodulatory mediator. The only currently approved IFN for the treatment of hepatitis C is IFN-α, though IFN-β is used in Japan. Another naturally occurring immunomodulator is thymosin alpha-1 (thymalfasin), a 28-amino-acid peptide produced by the thymus gland. Thymosin alpha-1 is an immunomodulator with multiple biological activities directed primarily at enhancing the TH 1 (inßammatory) immune response; it is able to induce the release of immunomodulating cytokines, activating T cells and natural killer cells (Sjogren MH, 2004). Thymosin alpha-1 mediates this activity through interaction with tumor necrosis factor receptor-associated factor 6 (TRAF6), which regulates activation of the IκB kinase (IKK) signaling pathway, controlling induction of interleukin-6, an important immunoregulatory cytokine (Zhang P, 2005). Interferon-Omega. Intarcia’s IFN-ω * * is a natural human IFN produced, in culture, through recombinant DNA technology. Boehringer Ingelheim originated IFN-ω for the treatment of hepatitis B and C, but after deciding not to pursue its development, the company licensed the compound to BioMedicines—which eventually became Intarcia—in 1998. IFN-ω is in Phase II trials for treatment of HCV in the United States and Europe. Because IFN-ω interacts with cells

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TABLE 12. Companies with Emerging Hepatitis C Therapies by Class and Phase, 2005 Non-

Corporation

Interferons

Anadys

Nucleoside

Interferon

Analogues

Immuno-

and IMPDH

Polymerase

Protease

Spectrum

Broad-

modulators

Inhibitors

Inhibitors

Inhibitors

Antivirals

I, Ia

Chiron

I, PCb

Coley

I

CSL

PCb

Endo, generics

III

Genmab

PC

Gilead Human Genome Sciences

I II

Idenix

IIc

Innogenetics Intarcia

II II

Intercell InterMune

II I

PC, PC

Japan Tobacco Migenix

II

Nabi Biopharmaceuticals Novartis

I/II Ia

IIc

Pharmasset

PCd

Roche

I, PCd

Schering-Plough SciClone

II, PC IIIe

Valeant

III

Vertex

II

IIf

Viropharm

Ig

Wyeth

Ig

XTL Biopharmaceuticals

I

a This molecule is being codeveloped by Anadys/Novartis. b This molecule is being codeveloped by Chiron/CSL. c This molecule is being codeveloped by Idenix/Novartis. d This molecule is being codeveloped by Pharmasset/Roche. e This molecule is being developed by Sigmau Tau in Europe and Schering-Plough KK in Japan. f This molecule is being developed by Mitsubishi Pharma in Japan. g This molecule is being codeveloped by Viropharm/Wyeth in the United States and Wyeth in Europe and Japan. I = Phase I; II = Phase II; III = Phase III; PC = Preclinical. IMPDH = Inosine-5’-monophosphate dehydrogenase.

by binding to the IFN-receptor complex with a distinct afÞnity proÞle (Cutrone EC, 2001), researchers expect that IFN-ω will be helpful for patients who do not respond to IFN-α. Intarcia is collaborating with Alza to formulate IFN-ω for use in Alza’s proprietary implantable, subcutaneous drug-delivery system (Duros), a strategy that would eliminate the need for patients to self-inject the drug on a daily or weekly basis.

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Preliminary data from a Phase II clinical trial of IFN-ω with or without ribavirin cotherapy was presented at the 56th annual meeting of the American Association for the Study of Liver Disease (AASLD) (Gorbakov V, 2005). This ongoing open-label, multicenter, active-controlled study, conducted in Russia, has enrolled 102 treatment-naive, HCV genotype 1-infected patients, split into two groups: IFN-ω alone and IFN-ω/ribavirin. Patients are administered IFN-ω (25 µg, subcutaneously) daily, to approximate blood levels of IFN-ω anticipated from the IFN-ω-Duros device; ribavirin is dosed at conventional doses (1,000–1,200 mg/day). End points of the study include early virological response (EVR; a greater than 2 log10 [100-fold] reduction in HCV RNA after 12 weeks of treatment) and SVR (undetectable HCV RNA six months after the end of 48 weeks of treatment). Results at 12 weeks of treatment demonstrate that IFN-ω is effective and well tolerated, with 84% of the patients treated with IFN-ω/ribavirin combination therapy achieving an EVR, in comparison to 60% of patients treated with IFN-ω alone. Furthermore, of those achieving an EVR, 92% had undetectable HCV RNA, as determined by Roche’s AMPLICOR HCV assay (limit of quantitation: less than 600 IU/mL). These EVR data suggest that IFN-ω/ribavirin will be comparable to peg-IFN-α/ribavirin treatment in terms of the frequency of SVR. In addition, normalization of serum alanine aminotransferase (ALT) levels, a common marker for liver inßammation and damage, occurred in approximately 73% of patients treated with IFN-ω/ribavirin and in 53% of patients receiving IFN-ω alone. Previous data on the conventional formulation of IFN-ω reported, in late 2001 at the 52nd AASLD conference, included the results of two Phase II trials, one in the United States and a larger trial in Europe. The U.S. Phase II study was designed to measure the effect of escalating doses of IFN-ω in patients with genotype 1 HCV infection who did not respond to prior treatment with IFNα. This study included three groups of eight patients who had HCV RNA that was consistently detectable in blood despite treatment with IFN-α (given with or without ribavirin). Patients received 15, 30, or 60 micrograms (µg) of IFNω once daily for 14 days. HCV RNA levels fell sharply (some to undetectable levels) within 48 hours in all patients treated with IFN-ω (McHutchison JG, 2001). Further reductions were seen in two out of the three dosing groups at the 14-day end point. Researchers reported that adverse effects were similar to those associated with other IFNs; 6 of 18 patients required dose reductions. The European Phase II study was undertaken to evaluate the safety, tolerability, and antiviral activity of six different dose levels of IFN-ω in treatment-naive patients with chronic hepatitis C. Preliminary data suggest that IFN-ω can reduce HCV RNA to undetectable levels in IFN-naive patients who have any of the four major HCV genotypes. In this open-label study, 90 patients infected with HCV (greater than 1 million copies/mL) and elevated liver ALT enzymes were treated with IFN-ω at 15, 30, 45, 60, 90, or 120 µg doses three times a week for 12 weeks. After 12 weeks, HCV was undetectable in 59% of the patients and ALT levels were normalized in 52% of the patients (Jetschmann JU, 2002; Plauth M, 2001). In 45% of the patients infected with genotype 1, HCV RNA

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was undetectable at any dose (15–120 µg); researchers saw the greatest efÞcacy at the 60 and 90 µg doses. In patients with genotypes 2, 3, or 4, HCV RNA was undetectable in 78% of patients; the most efÞcacious doses were 45 and 60 mcg. IFN-ω ’s safety and tolerability were similar to those of other IFNs; dosedependent inßuenza-like illness and thrombocytopenia were reported most often. Dose reduction was necessary for three patients who developed leukopenia, a condition that may limit this therapy’s efÞcacy, according to some experts. The IFN-ω-impregnated, titanium Duros drug-delivery device will be implanted, subcutaneously, into a patient’s upper arm and is expected to administer a steady dose of IFN-ω for up to three months. IFN-ω-Duros will likely launch in the United States and Europe in 2009. Although it is not in development in Japan, development is projected to follow successful trials in the United States and Europe and that it will launch in Japan in 2011. The drug could represent a signiÞcant improvement over available therapies for treating patients who are nonresponsive to IFN-α-based therapies. However, as it is likely that numerous options for therapies will be available by the time IFN-ω launches, it is doubtful that IFN-ω will have a major impact on the market at that point. Albuferon-Alpha. Human Genome Sciences’ albuferon-α * * is a novel, longacting form of IFN-α developed by fusing human IFN-α to human albumin. Researchers hope albuferon-α’s longer half-life will prove a clinical advantage over current interferons by reducing dosing frequency as well as adverse side effects that become evident with long-term treatment (e.g., myelosuppression, depression) or frequent parenteral administration (injection-site infections). Albuferon-α is in Phase II clinical trials in North America, Australia, and Europe and is expected to be used as an alternative treatment for patients who have failed previous IFN-α treatment for hepatitis C. In November 2004, Human Genome Sciences began enrollment in a randomized, open-label dose-escalation Phase II clinical trial studying the effect of albuferon in combination with ribavirin in HCV patients who had failed to respond to previous IFN and peg-IFN/ribavirin therapies (Human Genome Sciences, 2004). This trial enrolled approximately 115 patients in the United States and randomized them to Þve albuferon dose groups (ranging from 900 to 1, 800 µg) receiving subcutaneous injections of albuferon either 14 or 28 days apart and treating all patients with weight-based dosing of ribavirin (1,000 mg/day or 1,200 mg/day). This ongoing study will evaluate the frequency of SVR and the safety and tolerability of albuferon in combination with ribavirin, at the end of 48 weeks of treatment and a 24-week follow-up period. Results from 71 patients (after 24 weeks of treatment) in three treatment groups (900 or 1, 200 µg albuferon every 14 days and 1, 200 µg albuferon every 28 days [all patients received weight-based ribavirin daily]) were presented at the 56th AASLD meeting in 2005 (Nelson DR, 2005). In this 71-patient cohort, approximately 64% had not responded to previous therapy with peg-IFN-α and 93% were infected with HCV genotype 1. Over 60% had previously been treated with more than one IFN-α-based regimen with an average duration of 15 months.

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After 24 weeks of therapy, approximately 35%, 29%, and 25% of patients (in the 900 µg/14 days, 1, 200 µg/14 days, and 1, 200 µg/28 days cohorts, respectively) had no detectable HCV RNA, as determined using the Quantasure taqman PCR assay (limit of detection: less than 10 IU/mL). Although the greatest reduction in HCV RNA was observed in the 900 µg/14-day cohort (greater than 3 log10 [1,000-fold] reduction, after 24 weeks therapy), all dosage cohorts achieved average reductions in HCV RNA of over 2 log10 after 24 weeks of treatment. Albuferon/ribavirin was well tolerated and no difference in antiviral activity appeared between 14-day and 28-day administrations of albuferon. The most common side effects of albuferon/ribavirin treatment were fatigue, headache, myalgia, and nausea and were similarly distributed among all treatment groups, with 11% of subjects receiving dose reductions owing to adverse events. Hematological abnormalities arising from treatment were managed with dose reductions, with 14% of patients receiving dose modiÞcations because of hematological reductions. In April 2005, at the 40th annual meeting of the European Association for the Study of the Liver (EASL), Human Genome Sciences reported results from a Phase II trial of albuferon in treatment-naive HCV genotype 1-infected patients (Human Genome Sciences, 2005b). This trial was conducted in Canada and enrolled 56 patients, randomized to Þve treatment groups (200, 450, 670, 900, or 1, 200 µg). Patients were given two doses of albuferon, administered subcutaneously, 14 days apart (days 0 and 14). Patients were followed for evaluation of safety, tolerance, and antiviral activity for six weeks. HCV viral load was assessed by PCR during the six-week study period. Although the goal of the study was to determine the appropriate dose for follow-up studies, the primary efÞcacy end point of virologic response at day 28 was achieved. The mean reduction in viral load was 3.2 log10 IU/mL at day 28 in the 900 µg and 1, 200 µg treatment groups. Of these two cohorts, 69% of patients showed a viral load reduction greater than 2 log10 , and at day 42 (28 days after the second therapeutic injection), 23% of the patients had undetectable levels of viral RNA. Furthermore, after two subcutaneous injections of albuferon administered 14 days apart, albuferon was detectable in the serum for up to four weeks after the second injection. The median half-life was 148 hours, which supports dosing intervals of two to four weeks. Adverse events were transient and mild to moderate in severity. One subject developed low-titer antibodies to human serum albumin (HSA). However, no correlation was observed between induction of HAS antibodies and adverse events, pharmacokinetics, or antiviral response. Human Genome Sciences announced, in October 2005, completion of enrollment for an open-label, randomized, multicenter, dose-escalating Phase IIb clinical trial of albuferon/ribavirin in treatment-naive HCV genotype 1-infected patients (Human Genome Sciences, 2005a). This trial, which will be conducted in treatment centers in Australia, Canada, France, Germany, Israel, Poland, Romania, and the Czech Republic, will enroll more than 440 patients randomized into four treatment groups (all of which will receive weight-based ribavirin dosing): 900 µg albuferon at 14-day intervals, 1, 200 µg albuferon at 14-day intervals,

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1, 200 µg albuferon at 28-day intervals, and 180 µg peg-IFN-α-2a at weekly intervals. Patients will be treated for 48 weeks and followed for an additional 24 weeks after treatment. The primary end point is an SVR 24 weeks post-treatment. Albuferon-α will likely launch in the United States and Europe in 2009 and in Japan in 2012. The drug appears to offer a more convenient administration schedule compared with currently available treatments and is likely to be as effective as other IFN-α products in treating patients who have not responded to previous therapy. Experts are optimistic that albuferon-α will provide a more convenient option for long-term IFN-α therapy. With a reduction in dosing frequency to once every two weeks or even once monthly and no loss in efÞcacy, in comparison to peg-IFN-α, albuferon-α will have a distinct advantage over the peg-IFN-αs owing to its convenience for patients and physicians. Non-Interferon Immunomodulators Although the IFN drug class has been shown to be effective in treating HCV, and many new formulations of IFN-α and novel IFN variants are under investigation for treatment of HCV, other novel, non-IFN immunomodulators are also being investigated for potential treatment of HCV, including thymosin alpha-1 and Tolllike receptor (TLR) agonists. Thymosin-α may have advantages over IFN-based therapies if it demonstrates both improved safety and efÞcacy in comparison to peg-IFN-α. Additionally, because its mechanism of action is distinct from that of IFN, thymosin-α could be used to treat patients who are nonresponsive to IFN-based therapy and could potentially be used to augment the effects of IFN-α therapy. Toll-like receptor agonists induce upregulation of IFN gene expression; they have the advantage of being oral antivirals that can replace the use of parenteral IFN-α therapies. Mechanism of Action. Thymosin alpha-1 (thymalfasin), a 28-amino-acid peptide produced by the thymus gland, is an immunomodulator with multiple biological activities directed primarily at enhancing the TH 1 immune response; it is able to potentiate the action of cytokines and it stimulates T cells and natural killer cells. Likewise, Toll-like receptors regulate the immune response, particularly the expression of several immunomodulating cytokines, including IFN-α, IFN-β, IFN-γ , and IFN-ω. TLR agonists bind to TLRs, activating the receptors and thus potentiating de novo synthesis of cytokines that play an important role in viral eradication. Thymosin Alpha-1. SciClone Pharmaceuticals’ Zadaxin is a parenterally administered synthetic version of thymosin alpha-1 *. Thymosin alpha-1 is registered in more than 20 countries, predominantly for the treatment of HCV and hepatitis B virus (HBV) and as a vaccine adjuvant. Results of clinical trials are pending in the major markets, but thymosin alpha-1 is already launched for the treatment of hepatitis C in Myanmar, Pakistan, the Philippines, Singapore, Mexico, Peru, Sri Lanka, Argentina, and Venezuela and is preregistered or registered

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for treatment of HCV in Brazil, Egypt, Turkey, and Cambodia. Thymosin alpha1 is in Phase III trials for chronic HCV in the United States and Europe and in Phase II trials in Japan. SciClone has all rights to thymosin alpha-1 in the United States but has licensed exclusive development and marketing rights for Europe to Sigma-Tau and exclusive development and marketing rights in Japan to Schering-Plough KK, the Japanese subsidiary of Schering-Plough. SciClone has initiated two Phase III clinical trials assessing the beneÞt of thymosin alpha-1 cotherapy with peg-IFN-α-2a (Roche’s Pegasys) in nonresponder patients. These two trials enrolled more than 500 patients each, with the Þrst trial including patients with no cirrhosis of the liver and the second trial including patients with early cirrhosis of the liver. Roche provided Pegasys free of charge and, in exchange, SciClone agreed to share the trial data at the completion of the studies. Patients in the studies are given a 48-week course of either thymosin alpha-1 (1.6 mg twice/week) and peg-IFN-α-2a (180 µg once/week), or placebo and peg-IFN-α-2a. After the 48-week treatment period, patients will be followed for 24 weeks, with end points of both trials being elimination of HCV from serum and evidence of histological improvement. These two studies are expected to be completed by the end of 2005 (SciClone Pharmaceuticals, 2005b). SciClone, in collaboration with its partner Sigma-Tau, is conducting an additional Phase III clinical trial testing the efÞcacy of the triple-combination therapy regimen of peg-IFN-α-2a/ribavirin/thymosin alpha-1 in nonresponder patients (SciClone Pharmaceuticals, 2004a; SciClone Pharmaceuticals, 2004b). The trial will be conducted by Sigma-Tau and is a multicenter, placebo-controlled, doubleblind study that will enroll 550 nonresponder patients, all of whom are infected with HCV genotype 1 and have failed to respond to prior treatment with pegIFN-α and ribavirin cotherapy (SciClone Pharmaceuticals, 2004a). All trial participants will receive peg-IFN-α-2a (180 µg, once per week)/ribavirin (1,000 to 1,200 mg daily) cotreatment and will be given either placebo or thymosin alpha-1 (1.6 mg, twice weekly) in addition. Patients will undergo 48 weeks of treatment and 24 weeks of post-treatment follow-up. The primary end point will be SVR at week 72; secondary end points include normalization of ALT at the end of weeks 48 and 72, absence of HCV RNA at week 48, and an improvement in liver biopsy. Results from this trial are expected in 2007. Preliminary results from an open-label pilot Phase III clinical trial conducted in Mexico were presented at the 2004 AASLD meeting (SciClone Pharmaceuticals, 2004b). Updated, but not complete, results from this trial were recently presented during the annual meeting of the Congress of the Mexican Association of Hepatology (CMAH) in June 2005 (SciClone Pharmaceuticals, 2005a). The trial enrolled 40 patients infected with HCV who had not responded to prior treatment with peg-IFN-α/ribavirin cotherapy. Results presented at the CMAH meeting demonstrated that after 48 weeks of treatment and 24 weeks of followup, 6 of the of 30 patients (20%) treated with peg-IFN-α/ribavirin/thymosin alpha-1 had achieved an SVR (analysis of intent-to-treat group) (SciClone Pharmaceuticals, 2005a); all 6 patients who achieved SVR had HCV genotype 1. SciClone has stated that this 20% SVR achievement is an improvement over

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the 9% SVR previously reported in a trial of peg-IFN-α/ribavirin re-treatment of peg-IFN-α/ribavirin nonresponders (PEG-Intron Study Group, 2003; SciClone Pharmaceuticals, 2005a). Thymosin alpha-1 is administered via subcutaneous injection and is generally well tolerated; most clinical trials have found only local irritation at the injection site. No data are available on its safety in pregnant or nursing women. According to SciClone, thymosin alpha-1 has been administered to more than 10,000 patients in both clinical and commercial use, alone and in combination with antiviral and anticancer drugs, without producing any signiÞcant observed side effects or toxicities. Thymosin alpha-1 is expected to launch for HCV treatment in most of the major markets in 2007. In Japan, thymosin alpha-1’s clinical development for hepatitis B is more advanced than for HCV, and although the former indication is likely to be approved Þrst, the agent will likely gain approval for HCV in 2007. Thymosin alpha-1’s apparent ability to reduce serum ALT levels and achieve HCV viral clearance in some chronically infected patients when combined with peg-IFN-α-2a should allow this agent to be used in the treatment of patients who do not respond to Þrst-line therapy. Also, although SciClone has not enrolled treatment-naive patients in its Phase III trials, some patients who are contraindicated for ribavirin or viramidine are expected to be prescribed a thymosin alpha-1/peg-IFN-α combination therapy. Experts anticipate that thymosin alpha-1 may be a valuable agent for use in treatment of nonresponders, likely as an adjunct to peg-IFN-α/ribavirin combination therapy. Isatoribine. Anadys is developing the nucleoside analogue isatoribine (ANA245) * for the treatment of hepatitis C. The intravenous formulation of isatoribine is in Phase I trials in the United States, and an oral pro-drug is in preclinical development. Anadys predicts that isatoribine will initially be used to increase the therapeutic potential of IFN-α but that it may be used without IFN-α if clinical data demonstrate its efÞcacy as monotherapy. Isatoribine belongs to a class of nucleoside analogues that confer immunomodulatory effects through interaction with Toll-like receptor 7 (TLR7), inducing the innate immune response, including cytokines (such as interferon α, β, and γ ) and natural killer cells. Anadys has completed placebo-controlled, double-blind, single-dose escalation (50, 100, 200, 400, and 750 mg) Phase Ia clinical trials of intravenously administered isatoribine in healthy volunteers. The results of this trial, presented at the Hep DART meeting in 2003, showed that although isatoribine was well tolerated, the plasma half-life was relatively short (1.5 to 2 hours), and clearance was dose-dependent, requiring more frequent and larger doses to mediate activity. No serious or moderate adverse events were reported during this trial. Anadys presented preliminary results from an ongoing Phase Ib trial evaluating the safety, tolerability, pharmacokinetics, and pharmacodynamics of isatoribine, at the 55th AASLD meeting in 2004. This open-label, dose-escalating study (200, 400, 600, or 800 mg per day in 32 chronic HCV treatment-naive patients), conducted in Belgium, showed a decline in viral load (averaging 0.76 log10

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copies/mL) after one week of treatment in the 12 HCV-infected patients treated with 800 mg isatoribine per day (Horsmans Y, 2005). Ten of the 12 patients in this cohort were infected with HCV genotype 1 virus. Clinical trials are ongoing, and the company has not released data on isatoribine’s long-term clinical performance. ANA-975. In June 2005, Anadys signed a global license and codevelopment agreement with Novartis on the development and potential commercialization of ANA-975 *, a pro-drug of isatoribine that can be formulated for oral administration. An investigational new drug application (IND) for ANA-975 was recently accepted by the FDA, and preliminary data, presented at the 56th AASLD meeting, from a dose-escalating Phase I clinical trial conducted with 36 subjects in the United Kingdom demonstrated that a single dose of ANA-975 has nearly complete bioavailability and that conversion to isatoribine is rapid and efÞcient, achieving clinically relevant levels of isatoribine (Kerr B, 2005). CPG-10101. Coley Pharmaceutical Group is developing CPG-10101 (actilon)∗ —a TLR9 agonist—for the treatment of HCV infection. Like other TLR agonists in development, CPG-10101 functions by stimulating the innate immune response through induction of interferons. Results of a Phase Ia clinical trial conducted in 40 healthy volunteers followed for 29 days showed that CPG-10101 is well tolerated over all doses tested (0.25, 1, 4, 10, or 20 mg administered twice, 14 days apart) and induces IFN-α. Volunteers had no serious adverse events or dose-limiting toxicities although mild injection site reactions and mild to moderate ßu-like symptoms were experienced by several volunteers and were consistent with expectations because of CPG-10101’s mechanism of action (Coley Pharmaceutical Group, 2005b). Results from a double-blind Phase Ib clinical trial studying the safety and efÞcacy of CPG-10101 in 42 nonresponder, predominantly HCV genotype 1-infected patients were presented at the DDW meeting in May 2005. Patients in this study were randomized to the same dosing cohorts as the previous study and received twice-weekly injections of CPG-10101 for four weeks. CPG-10101 was well tolerated, and patients in the 20 mg cohort achieved a mean 1.4 log10 reduction in viral load within four weeks; Þve of the six patients in this cohort achieved at least a 1 log10 reduction in viral load during the trial (Coley Pharmaceutical Group, 2005a). A Phase I study on the CPG-10101-induced immune response in 60 HCVinfected patients (primarily genotype 1) was presented at the 56th AASLD meeting in 2005. This study demonstrated that CPG-10101 activates numerous leukocytes, including NK, T, B, and dendritic cells in a dose-dependent manner (Vicari AP, 2005). Further clinical trials have not yet been announced. Nucleoside Analogues and IMPDH Inhibitors Overview. The nucleoside analogue ribavirin (Schering-Plough’s Rebetol, Roche’s Copegus) (Figure 3) is a broad-spectrum antiviral that was found to be effective in treating HCV. However, ribavirin therapy is often associated with severe side effects, including anemia, and is thus contraindicated for use

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in a sizable portion of patients. Thought leaders indicate that therapies that do not cause anemia would be potentially more efÞcacious because they could be used at higher doses and in a wider range of patients. Several second-generation ribavirin-like molecules are in development that improve on the safety and tolerability of ribavirin. However, because the anti-HCV mechanism of action of this drug class is poorly deÞned, subtle changes in structural chemistry of drug candidates can have a dramatic impact on their efÞcacy. Therefore, it is difÞcult to predict which of these molecules will be successful. Valeant’s viramidine is the most advanced in development and appears to have favorable safety and efÞcacy. Mechanism of Action. Ribavirin is a nucleoside analogue that is effective in treating HCV. Although its mechanism of action remains relatively unclear, it likely affects viral replication by impeding two important pathways in DNA and RNA replication. As a guanosine analogue, ribavirin may be incorporated into the elongating RNA strand, and owing to its variant chemical structure, it can be base-paired with either cytosine or uracil (guanosine usually basepairs with cytosine), causing “error catastrophe” mutagenesis of the HCV RNA genome that leads to production of noninfectious virions (Maag D, 2001). In addition, as a guanosine analogue, ribavirin may mediate antiviral properties through inhibition of inosine-5’-monophosphate dehydrogenase (IMPDH), an enzyme that plays a role in guanosine synthesis. Inhibition of IMPDH activity reduces the available intracellular pool of guanosine, augmenting ribavirin’s error-catastrophe-generating antiviral effect. Although it has been shown, using the in vitro replicon HCV model, that addition of exogenous guanosine reduces ribavirin’s antiviral effects, similar experiments have shown that ribavirin’s inhibition of IMPDH is not the major contributing factor conferring its overall antiviral properties (Zhou S, 2003). Merimepodib. Vertex Pharmaceuticals is developing merimepodib (VX-497) *—an oral, IMPDH inhibitor—for the treatment of HCV infection. Early Phase II trials for merimepodib alone and in combination with peg-IFN and ribavirin have been completed in the United States and Europe, and another Phase II trial of triple-combination therapy is ongoing in the United States. The company anticipates Þling a new drug application (NDA) for merimepodib as early as 2006–2007 for use in patients who do not respond to previous HCV treatment. Merimepodib is a potent non-nucleoside inhibitor of IMPDH and is thought to be potentially more effective than ribavirin for treating HCV. In vitro studies have demonstrated that merimepodib is more potent than ribavirin against several viruses, including HBV and inßuenza A, and that this antiviral activity is enhanced in the presence of IFN-α (Markland W, 2000). Researchers do believe that combination therapy of IMPDH inhibitors may increase ribavirin’s antiviral efÞcacy by reducing the pool of guanosine triphosphate (GTP) available for replication of HCV RNA, forcing the virus to incorporate triphosphate ribavirin more frequently into its genome, and increasing mutagenesis (Zhou S, 2003). Consequently, merimepodib may be most efÞcacious in triple-therapy regimens with peg-IFN-α and ribavirin.

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This assumption was tested in a European, placebo-controlled Phase IIa pilot clinical trial comparing merimepodib in combination with peg-IFN-α and ribavirin (Vertex Pharmaceuticals, 2004a). In April 2004, during the 39th annual meeting of the EASL, Vertex researchers presented data from the completed treatment arm of this Phase II trial. In the trial, 31 patients with genotype 1 HCV who did not respond to at least 12 weeks of prior interferon and ribavirin therapy received triple therapy including merimepodib. After six months of therapy, patients who responded to therapy had the option to extend the therapy an additional six months. Initial reports showed that the triple-therapy combination was well tolerated, meeting its primary end points of safety and tolerability, and was not associated with any serious adverse events. Merimepodib also demonstrated a statistically signiÞcant dose-dependent reduction of HCV viral load, which was the secondary trial end point (Vertex Pharmaceuticals, 2003). At the 2004 EASL meeting, in Berlin, Germany, the company presented data from ten patients who participated in the trial extension and completed 48 weeks of treatment; three of the patients were given placebo plus peg-IFN-/ribavirin therapy. After 48 weeks of treatment and 24 weeks of post-treatment follow-up, one of the three patients in the placebo group (peg-IFN-α/ribavirin alone) and three of the seven patients in the triple-combination treatment group had achieved an SVR (Vertex Pharmaceuticals, 2004a). Vertex has completed enrollment in a follow-up Phase II merimepodib triplecombination study (METRO), which aimed to enroll approximately 315 nonresponders and treat them with 50 mg merimepodib, 100 mg merimepodib, or placebo, twice per day, in combination with standard doses of peg-IFN-α-2a/ribavirin for 24 weeks (Vertex Pharmaceuticals, 2004b; Vertex Pharmaceuticals, 2005a). After 24 weeks, patients with undetectable viral load will continue on low-dose interferon for an additional 24 weeks, after which patients will be followed for another 24 weeks. The multicenter study, which is being conducted in the United States, has two main end points: assessment of merimepodib’s antiviral activity as measured by the proportion of treated nonresponders who achieve SVR, compared with placebo, and evaluation of merimepodib’s contribution to efÞcacy in a regimen with peg-IFN-α and ribavirin. Merimepodib is expected to launch in the United States and Europe in 2008, and in Japan in 2011. The agent is a potent in vitro inhibitor of viral replication, is well tolerated, and is able to signiÞcantly reduce ALT levels. Because merimepodib targets IMPDH, thought leaders expect that merimepodib may be used in triple combination with peg-IFN-α/ribavirin in treatment of patients who do not respond to an initial course of peg-IFN-α/ribavirin combination. Assuming that its use in triple combination with peg-IFN-α and ribavirin is safe and demonstrates enhanced efÞcacy over current combination therapies, and owing to competition from other emerging therapies and experts’ skepticism of the magnitude and signiÞcance of the potential boost in efÞcacy conferred by the triple-combination regimen, merimepodib will likely be used as a second-line adjunct therapy for treatment of nonresponders.

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Viramidine. Valeant (formerly ICN) Pharmaceuticals’ viramidine is undergoing Phase III clinical trials in the United States and Europe for the treatment of chronic HCV infection, and the company recently initiated clinical development of viramidine in Japan (Valeant Pharmaceuticals, 2005c). Viramidine exhibits antitumor and immunomodulatory activities and has shown some efÞcacy as a broad-spectrum antiviral, with reports of potential use in treatment of inßuenza (Sidwell RW, 2005; Smee DF, 2004). A pro-drug of ribavirin, viramidine is converted to ribavirin via the enzyme adenosine deaminase. Because the liver has signiÞcantly higher levels of adenosine deaminase than other organs, viramidine is most frequently processed in the liver, resulting in localization of the active drug within the liver (Lin CC, 2003), with relatively little drug accumulating in red blood cells (RBCs), where its toxicity can lead to hemolysis. Preclinical studies have shown that viramidine is targeted to the liver and is not signiÞcantly teratogenic in animal models (Barnard D, 2002; Lin CC, 2003). A pilot study of viramidine’s pharmacokinetics, conducted in healthy male volunteers, showed that orally administered viramidine was rapidly absorbed and converted to ribavirin (Lin CC, 2004). Results of an open-label study conducted in 24 patients with HCV infection determined that after multiple dosing, viramidine has a shorter half-life (66–76 hours in plasma; 200–240 hours in RBCs) than ribavirin (340–410 hours in plasma; 360–430 hours in RBCs) (Aora S, 2005). At the 2005 EASL meeting, researchers from Valeant presented data from a Phase II clinical trial comparing the efÞcacy of peg-IFN-α/viramidine compared with peg-IFN-α/ribavirin (Valeant Pharmaceuticals, 2005a). This study was conducted in the United States and enrolled 180 treatment-naive patients with chronic HCV. Patients were treated (400, 600, or 800 mg twice daily) based on the genotype of their HCV infection, with genotype 2- or 3-infected patients receiving 24 weeks of cotherapy and genotype 1-infected patients receiving 48 weeks of cotherapy. Both regimens received 24 weeks of post-treatment follow-up. Overall, viramidine-associated anemia (Hb less than 10 g/dL) occurred in only 4% of viramidine-treated patients compared with 27% of ribavirin-treated patients. The greatest efÞcacy and lowest frequency of adverse events were observed in the 600 mg treatment group, where only 2% of patients became anemic. Thirty-seven percent of patients dosed with 600 mg viramidine achieved an SVR compared with 44% of ribavirin-treated patients. More importantly, 27% of viramidine (600 mg) and 35% of ribavirin-treated patients with HCV genotype 1 had an SVR, while 62% of viramidine (600 mg) and 73% of ribavirin-treated patients with HCV genotype 2 or 3 had an SVR (Valeant Pharmaceuticals, 2005a). Except for anemia, no signiÞcant side effects were observed. The most common adverse effects included fatigue, headache, insomnia, depression, and myalgia. These side effects occurred with equal frequency in the viramidine-treated groups and the ribavirin-treated groups. Although no statistically signiÞcant difference was observed in terms of proportion of patients achieving an SVR with either viramidine or ribavirin treatment, some experts state that viramidine’s efÞcacy data are “underwhelming” and believe that the data may potentially force physicians

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to choose between improved safety (viramidine) and more robust efÞcacy (ribavirin) in treatment of HCV patients. However, other thought leaders express optimism for viramidine; they state that its beneÞts in reducing anemia, along with the reduction in the costs associated with both monitoring of patients for the occurrence of this common side effect of ribavirin treatment and treating anemic patients, boost its prospects for widespread use, potentially replacing ribavirin in treatment of HCV. Based on the results of the Phase II trial, Valeant has initiated two Phase III trials, Viramidine’s Safety and EfÞcacy vs. Ribavirin (VISER1 and VISER2), which will be conducted at approximately 100 sites worldwide and recruit approximately 1,000 patients in each study. Both studies will compare viramidine (600 mg twice daily) with ribavirin (1,000/1,200 mg daily) when used in combination with a peg-IFN-α. VISER1 will use Schering-Plough’s peg-IFN-α-2b and VISER2 will use Roche’s peg-IFN-α. Both studies will treat genotype 1 patients for 48 weeks and genotype 2 or 3 patients for 24 weeks, with a 24-week follow-up. The studies will measure end points of SVR and anemia. Viramidine will likely be launched in the United States and Europe in 2008. Though the agent is not in development in Japan, development is expect to follow successful trials in the United States and Europe and it may launch in Japan in 2011. Current treatments for hepatitis C rely on the use of ribavirin to enhance the efÞcacy of IFN-α; however, ribavirin’s use is limited by its adverse side effects, primarily hemolytic anemia. Although more data are needed to fully evaluate the efÞcacy of viramidine, the drug appears to be a less toxic alternative to ribavirin and is localized to the liver. If viramidine proves to be as effective as ribavirin in its ability to potentiate the activity of IFN-α but provides an improved safety proÞle over that of ribavirin, viramidine will likely replace ribavirin. However, the availability of generic ribavirin may constrain the market outlook for viramidine. Cost-conscious health care systems will have to weigh the cost of administering additional therapy to compensate for anemia in ribavirin-treated patients, and the stafÞng and labor costs of monitoring patients on ribavirin therapy against the costs of viramidine. Furthermore, given some experts’ skepticism of viramidine’s efÞcacy, physicians may be less willing to forgo ribavirin’s perceived advantages in efÞcacy in favor of viramidine’s improvements in safety. HCV Polymerase Inhibitors Overview. Owing to clinical successes in targeting viral RNA replication in other viral infections (HIV-1, HBV), the RNA-dependent RNA polymerase (RdRp) of HCV has been the focus of intense drug discovery activity. Unlike the IMPDH inhibitors, HCV polymerase inhibitors will speciÞcally inhibit the replication step of the HCV life cycle. Researchers hope that this speciÞcity will make this class of antivirals have fewer side effects and greater potency than broad-spectrum antivirals. Mechanism of Action. During viral infection, the HCV RdRp makes copies of the HCV genome for packaging into new virus particles. Interference with replication of the genome inhibits the production of new virions, limiting the spread

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of the virus. Because infected cells may have a long life span, it is unknown whether inhibition of HCV replication is sufÞcient to clear the infection. However, effective blocking of new virus production would certainly prevent further disease manifestations and possibly give the immune system time to eliminate persistently infected cells. Researchers have identiÞed both nucleoside and nonnucleoside inhibitors of the HCV RdRp, and they are the subject of ongoing research efforts. Some nucleoside analogues have dual effects: polymerase inhibition and IMPDH inhibition. The level to which each activity contributes to the overall antiviral properties of the nucleoside analogue is unclear. Valopicitabine. Novartis/Idenix Pharmaceuticals’ valopicitabine (formerly known as NM-283) * is an inhibitor of the HCV RdRp that was developed using structure-based drug design. In March 2003, Novartis acquired 51% of Idenix stock and has the option to jointly develop additional Idenix antiviral candidates, including valopicitabine. Valopicitabine is in Phase II clinical trials in the United States and Europe. At the 54th annual AASLD meeting (2003), clinical investigators at Idenix presented results of a Phase I/II trial on valopicitabine’s efÞcacy and tolerability in patients with HCV genotype 1 infection (Idenix Pharmaceuticals, 2004). Patients enrolled in this trial had several deÞning characteristics: they were either treatment-naive or nonresponders to interferon-based therapy, had viral load of greater than Þve log10 IU/mL, had ALT levels below Þve times the upper limit of normal (ULN), had not received interferon treatment in the past six months, and had compensated liver disease with no cirrhosis. Patients were randomized into eight groups, with each group comprising ten drug-treated and two placebotreated patients. Each group received a different dose, ranging from 50 mg to 800 mg valopicitabine once daily, for 15 days and two weeks of post-treatment follow-up. The trial demonstrated that valopicitabine, when administered at doses greater than 400 mg per day, is well tolerated, with the most common side effect being mild to moderate transient nausea in patients. Valopicitabine therapy did not cause any serious adverse events or abnormalities, nor did it reveal any doselimiting toxicities. During the treatment period, the greatest reduction in viral load was observed in patients receiving 800 mg/day, the highest dose administered in the trial. The log10 reduction in viral load in these patients ranged from 0.41 log10 below baseline to 1.79 log10 below baseline, with one patient achieving a reduction in viral load of 2.37 log10 below baseline. However, viral clearance was not achieved during the 15-day treatment period and viral load in most valopicitabine-treated patients began to return to baseline levels during the two-week post-treatment follow-up period (Idenix Pharmaceuticals, 2004). The response to valopicitabine treatment observed in patients with HCV genotype 1 infection was encouraging, and Idenix followed up the previous trial with an open-label Phase IIa trial to test whether the effects of valopicitabine treatment were synergistic with those of peg-IFN-α treatment. The data from this trial were presented in April 2005, at the 40th annual EASL meeting. This trial enrolled 30 patients with similar demographics to the previous Phase I trial. Twelve patients received valopicitabine monotherapy, while the remaining

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18 patients were treated with valopicitabine (step-up therapy of 400 to 600 to 800 mg/day until day eight, then 800 mg/day thereafter) and peg-IFN-α-2b (1.0 µg/kg weekly). Of the 12 patients receiving monotherapy, 2 discontinued, 9 did not achieve an EVR by 12 weeks of treatment, and 1 patient achieved EVR at 12 weeks and continued with treatment to 24 weeks. Of the combinationtherapy recipients, 3 were discontinued and another 3 did not achieve EVR by 12 weeks of treatment. Of the 12 patients who continued with therapy beyond 12 weeks, 9 of them completed therapy through 24 weeks. No serious adverse side effects or dose-limiting toxicities were observed during the study, and the most frequent adverse events were nausea, headache, vomiting, dyspepsia, and ßu-like symptoms. One grade 3 lab abnormality was observed in a patient receiving combination therapy: in this patient, absolute neutrophil count (ANC) approached 620 cells/mm3 at day 11 but rebounded to baseline levels by day 15 with no interruption or modiÞcation of the treatment regimen. This Phase IIa trial showed that by week 12 (day 85), combination therapy produced an average reduction in viral load of 3 log10 IU/mL, with four patients already PCR-negative (Idenix Pharmaceuticals, 2005). By week 24, the remaining nine patients receiving combination therapy had achieved an average viral load of 1.4 log10 , approximately 4.5 log10 below baseline. Eight of the nine patients had viral loads below both the lower limit of quantitation for the Amplicor HCV RNA viral load (below 600 IU/mL); seven of nine were below the lower limit of detection for this test (50 IU/mL), while six of nine had viral loads below the lower limit of detection of the TaqMan assay (10 IU/mL). Of the Þve combination therapy patients who were PCR-positive at week 12, two of them achieved PCRnegative status by week 24. Of the combination-therapy recipients, 50% of them achieved at least a 2 log10 reduction in viral load from baseline, with 33% of the 18 HCV genotype 1 patients achieving PCR-negative status after 24 weeks of combination therapy. This trial has been extended through 48 weeks of treatment, although results of the trial extension have not yet been published. The 24-week results demonstrate that valopicitabine improves efÞcacy in combination therapy with peg-IFN-α and may provide an alternative treatment regimen for patients with HCV genotype 1 infection- and for those who have not responded to previous therapy. At the 56th AASLD meeting, in 2005, Idenix presented the preliminary results (week 12) of a Phase IIb trial designed to compare efÞcacy of treatment using a valopicitabine/peg-IFN-α-2a regimen with ribavirin/peg-IFN-α-2a in patients with HCV genotype 1 infection who failed to respond after at least three months of conventional peg-IFN-/ribavirin treatment (O’Brien C, 2005). The trial enrolled 178 HCV genotype 1-infected patients who were nonresponsive to previous therapy, split into Þve treatment groups, all of which were treated for 48 weeks followed by a 24-week follow-up period: • •

Group A: 800 mg valopicitabine monotherapy throughout study. Group B: 400 mg valopicitabine monotherapy for one month, followed by cotherapy with peg-IFN-α-2a for 44 weeks.

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Group C: 400 to 800 mg valopicitabine (ramped-up dosing within Þrst week) monotherapy for one month, followed by cotherapy with peg-IFN-α-2a for 44 weeks. Group D: 800 mg valopicitabine monotherapy for one month, followed by cotherapy with peg-IFN-α-2a for 44 weeks. Group E: Peg-IFN-α-2a/ribavirin therapy started at one month and continued for 44 weeks.

HCV RNA was evaluated after 4, 12, and 24 weeks of therapy, and patients were continued on therapy if they had achieved a reduction in HCV RNA (from baseline) of greater than 0.5 log10 , 1 log10 , and 2 log10 , at each respective time point. By week 12, 21 patients had been discontinued and 157 remained on therapy. The mean reduction in HCV RNA, at week 12, in the peg-IFN-α-2a/ribavirin group was approximately 1.3 log10 while only two of the four treatment groups showed statistically signiÞcant improvements in reduction of viral load, in comparison to peg-IFN-α-2a/ribavirin: groups C (-2.51 log10 ) and D (-2.77 log10 ). Similarly, at week 12, although 41% of peg-IFN-α-2a/ribavirin-treated patients achieved an EVR, 71% (group C) and 63% (group D) of the patients in the valopicitabine-cotherapy groups achieved an EVR. Groups C and D, combined, outperformed group E (peg-IFN-α-2a/ribavirin) with 21% of patients in groups C and D experiencing a reduction in HCV RNA of over 4 log10 , while only 6% of peg-IFN-α-2a/ribavirin treated patients achieved a similar result (O’Brien C, 2005). Idenix anticipates commencement of Phase III clinical trials of valopicitabine in early 2006. With Novartis’s involvement, valopicitabine will likely launch in 2009 in the United States and Europe and in 2011 in Japan. Initially, valopicitabine will not take share from peg-IFN-α/ribavirin. Rather, experts anticipate, valopicitabine will Þrst be used as part of a triple-therapy combination for difÞcult-to-cure (e.g., genotype 1, African-Americans) treatment-naive patients and, more frequently, in combination with a peg-IFN-α (or third-generation IFNbased therapies such as Albuferon-α) for treatment nonresponders. By 2014, valopicitabine likely will be used in both Þrst-line and second-line therapies and could potentially replace nucleoside analogues in combination with peg-IFNα (or albuferon-α). Furthermore, with the launch of novel therapies in other drug classes, such as the protease inhibitors, it is projected that physicians will investigate the use of triple-combination therapies consisting of valopicitabine, peg-IFN-α, and a protease inhibitor and it is likely that several regimen variations that include valopicitabine will be under investigation at the end of the 2004–2014 study period. HCV-796. ViroPharma and Wyeth are codeveloping a novel, orally administered RNA polymerase inhibitor, HCV-796, for treatment of HCV. The drug is undergoing clinical trials; if launched, it will be copromoted by the two companies in the United States and Canada, with Wyeth holding commercial rights for the rest of the world. HCV-796 is in Phase I clinical trials in the United States.

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In vitro assays conducted using the HCV-replicon in human cell lines have shown that HCV-796 has an effective concentration for a 50% reduction in viral replication (EC50) of 5 nM (against genotype 1a) and 9 nM (against genotype 1b), which is a 128-fold and 22-fold, respectively, increase over the potency of a predecessor antiviral, HCV-086 (for which development has been suspended) (ViroPharma, 2005a). Results from studies conducted in chimeric mouse models have shown that HCV-796 has signiÞcant antiviral activity as a single agent. Preliminary data from a single-dose, placebo-controlled, dose-escalation Phase I clinical trial have shown that HCV-796 is well tolerated and has a favorable pharmacokinetic proÞle at all doses tested (ViroPharma, 2005b). In May 2005, ViroPharma announced plans for a randomized, double-blind, multiple ascending-dose Phase Ib clinical trial to be conducted in the United States involving 96 treatment-naive HCV-infected patients. Patients in this study will be given multiple ascending doses of HCV-796 for 14 days, and the safety, antiviral activity, and pharmacokinetic proÞle of the drug will be evaluated in comparison to placebo. Preliminary results of this trial demonstrated that maximal reduction in viral load occurred after four days of therapy, with 83% of patients receiving 1,000 mg HCV-796 twice daily achieving an average reduction of 1.4 log10 HCV RNA (ViroPharma, 2005b). R-1626 and PSI-6130. Roche is expanding its HCV pipeline with several candidates being developed in-house as well as through collaborations. The products in Roche’s pipeline include R-1626, an HCV RdRp inhibitor in Phase I trials, and PSI-6130, a polymerase inhibitor being developed by Pharmasset for use in combination with peg-IFN-α-2a/ribavirin in treatment of HCV genotype 1-infected patients. HCV Protease Inhibitors Overview. HCV proteases play a critical role in the life cycle of the virus and are high-proÞle targets for antiviral development. Researchers are developing HCV-speciÞc protease inhibitors with the hope that these novel therapies, in combination with existing therapies, will boost efÞcacy, especially against difÞcult-to-treat HCV genotypes (1 and 4) and in patients who have not responded to current therapies or who have relapsed. Although several companies have programs developing novel HCV-speciÞc protease inhibitors, including Boehringer Ingelheim, Bristol-Myers Squibb, InterMune, Chiron, and Medivir, only the leading candidates that have been publicly disclosed are discussed here. Mechanism of Action. The life cycle of HCV depends on the activity of speciÞc proteases that are encoded within the RNA genome of the virus. The open reading frame of the HCV genome is translated in a single process, resulting in a lengthy polyprotein that is subsequently spliced to produce the various enzymes required for propagation and replication of the virus. NS3-4A, a serine protease and promiscuous helicase, plays an important role in both the splicing of the HCV polyprotein and in RNA replication. NS3-4A is an induced-Þt protease requiring both substrate, NS3, and NS4A, a cofactor encoded

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downstream from NS3 in the HSV operon, for full activation of its catalytic properties. NS4A binds NS3, with the β-sheets of NS4A becoming encapsulated within the structure of NS3, causing a conformational change in NS3’s structure that augments catalytic activity at the active site (Yan Y, 1998). The active site of the NS3-4A protease is susceptible to inhibition by its own Nterminal cleavage products (corresponding to the NS4A/NS4B, NS4B/NS5A, and NS5A/NS5B junctions). This feature of product inhibition has prompted many investigators to focus on developing peptide inhibitors of the HCV NS34A protease, though many of the drugs currently in clinical trials are small molecules. Formulation. Most protease inhibitors are competitive inhibitors designed from peptide substrates or cleavage products. Others have been generated through extensive screening of peptide and combinatorial libraries. The active site of the NS3-4A protease is fairly smooth and shallow, characteristics that make the NS3-4A protease particularly challenging for small-molecule design (Lamarre D, 2003; Lu L, 2004). However, several small, orally formulated peptidomimetic compounds that inhibit NS3-4A, such as BILN-2061 and VX-950, have been developed (Lamarre D, 2003; Lin C, 2004). Modeled after tetrapeptide and tripeptide mimics of the protease-binding domain, BILN-2061 is a potent and speciÞc protease inhibitor with in vitro activity toward the NS3-4A serine proteases of HCV genotypes 1a and 1b (Lamarre D, 2003). Although safety or tolerance issues did not arise during the short treatment course in the Phase I pilot studies, BILN-2061 was recently withdrawn from Phase II studies in Europe because of potentially adverse supratherapeutic effects observed during animal studies. Experts are disappointed but not surprised by this development and speculate that although several of the leading HCV-speciÞc antivirals in development would likely fail, they would pave the way for more-successful second-generation agents. VX-950. Vertex Pharmaceuticals is developing VX-950, an orally available small-molecule peptidomimetic inhibitor of the NS3-4A protease, for the treatment of HCV. VX-950 has demonstrated encouraging efÞcacy in Phase I clinical trials and is entering Phase II clinical trials. VX-950 was originally developed in collaboration with Eli Lilly; Vertex was responsible for discovery and precommercial manufacturing, and Lilly was responsible for clinical trials, formulation, and marketing. However, in January 2003, Vertex announced that it had regained all rights to VX-950 from Lilly, including global marketing rights, though Lilly retains some royalty interest in future sales. In June 2004, Vertex signed an agreement with Mitsubishi Pharma, giving Mitsubishi rights to develop and commercialize VX-950 in Japan and certain Far East countries. Mitsubishi will make precommercial payments to Vertex, in support of VX-950’s preclinical development, and will pay royalties on sales of VX-950. Vertex retains exclusive development and marketing rights to VX-950 in the rest of the world. In October 2003, preclinical data for VX-950 were presented at the 54th AASLD conference. In an in vitro cell-based replicon system that models HCV

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infection, nine days of treatment with VX-950 reduced HCV viral loads by almost 4 log10 . In June 2004, a Phase I placebo-controlled trial conducted in 35 European patients tested safety and tolerability of single doses ranging from 25 mg to 1,250 mg VX-950 (Vertex Pharmaceuticals, 2004c). The study found that VX950 was well tolerated and not associated with any serious adverse events and that increasing the dose did not lead to an increased frequency of adverse events. In addition, VX-950 is orally bioavailable, and blood concentrations of VX-950 reached or exceeded desired antiviral active levels for doses at or above the middle of the range tested. Clinical and preclinical data predict that average liver concentrations of VX-950 exceeded the in vitro 90% inhibitory concentration (IC90) for the drug by 57-fold and the IC50 by 113-fold and that these levels could be sustained for over 12 hours for some doses. Vertex conducted a Phase Ia clinical trial to investigate safety and tolerability of VX-950 and presented data from this study at the Digestive Disease Week conference in May 2005. This double-blind, placebo-controlled study enrolled 24 patients with chronic genotype 1 HCV infection; each patient received placebo or one of three doses of VX-950 (450 mg every eight hours, 750 mg every eight hours, or 1,250 mg every 12 hours) for 14 days (Vertex Pharmaceuticals, 2005b; Vertex Pharmaceuticals, 2005c). After three days of treatment, the median reduction in viral load was approximately 3 log10 in all dose groups. In the treatment group receiving 750 mg per eight hours, a further reduction in viral load was observed by the end of the 14-day treatment period, with median reduction in viral load of 4.4 log10 . At day 14, four of the eight patients in this treatment group had viral loads below the lower limit of quantitation in the Roche COBAS Taqman assay (below 30 IU/mL) and two of these four patients had undetectable virus (below the 10 IU/mL lower limit of detection in the Taqman assay). Overall, Þve patients in the study had viral loads below the lower limit of quantitation of the Taqman assay after 14 days of treatment. However, after completion of treatment, viral load levels generally increased over the 28-day post-treatment observatory period, and 28 days after receiving the Þnal dose of VX-950, two patients had viral loads within 1 log10 of their pretreatment baseline level (Vertex Pharmaceuticals, 2005c). The second phase of this study was presented, along with several additional studies on pharmacokinetics and viral resistance, at the 56th AASLD meeting in 2005 (Reesink HW, 2005). In this Phase 1b study, 34 genotype 1-infected patients (including 5 treatment-naive patients) were divided into the same three treatment groups (each patient receiving either placebo or one of three doses of VX-950: 450 mg every eight hours [10 patients], 750 mg every eight hours [8 patients], or 1,250 mg every 12 hours [10 patients]), with six patients receiving placebo. Patients were treated for 14 days, with common side effects being headache, abdominal pain, nausea, and diarrhea. Those treated with 750 mg VX-950 every eight hours experienced the largest declines in HCV RNA; all 8 of them dropped over 3 log10 HCV RNA within the Þrst two to three days of treatment. In this treatment group, the drop in viral load was sustained throughout the 14-day study period. Across all three doses, 26 of the 28 patients had an HCV RNA decline

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of over 3 log10 and all had an HCV RNA decline of over 2 log10 . Reductions in ALT levels were observed in all treatment groups, with over one-half of patients achieving normalization of ALT by the end of treatment. RNA (cDNA) sequencing of the HCV NS3 protease isolated from patients in this Phase 1b clinical trial prior to and following treatment revealed that appearance of VX-950-resistant HCV correlates with the type of treatment response observed: continuous HCV RNA decline; viral rebound (increase in HCV RNA after the nadir of decline and while on therapy); plateau (sustained, diminished HCV RNA after nadir) (Sarrazin C, 2005). Although resistant HCV was more frequently present in patients with treatment responses similar to either the viral rebound or plateau response, most of the resulting resistant viruses were less Þt and the variant with the highest VX-950 resistance, A156V/T, is both less Þt than wild-type virus and susceptible to interferon therapy (Lin C, 2005; Sarrazin C, 2005). SCH-503034. Schering-Plough is conducting research into HCV NS3 protease inhibitors, and its lead compound in this class, SCH-503034, is entering Phase II clinical trials. Data on SCH-503034 were recently presented at the 56th AASLD meeting. In a Phase I trial, 48 patients who were infected with HCV genotype 1 virus and were nonresponsive to prior therapy were split into four treatment groups, with 10 to 12 patients in each group receiving oral SCH-503034 (doses: 100, 200, or 400 mg twice daily, or 400 mg thrice daily) and the remaining 4 patients receiving placebo for 14 days (Zeuzem S, 2005a). The drug was rapidly absorbed and was well tolerated, with no dose-related side effects. Although the greatest reductions in HCV RNA and ALT were observed in patients receiving the maximal dose (400 mg, thrice daily), the average reduction of HCV RNA in this treatment cohort was approximately 1.5 log10 at the end of the 14 days of treatment, and the mean change in ALT was over 50 U/L (Zeuzem S, 2005a). Both reductions in HCV RNA and ALT were less dramatic for other doses of SCH-503034. A drug-resistant HCV variant, V170A, was identiÞed in only one patient, although it became undetectable during follow-up, indicating its relative lack of replicative Þtness. A Phase I crossover-design trial studying the effects of SCH-503034 in combination with peg-IFN-α-2b versus the effects of either agent alone in HCV genotype 1-infected patients who were nonresponsive to prior peg-IFN-α-2bbased therapy was also reported during the 56th AASLD meeting (Zeuzem S, 2005b). In this study, patients were randomized to receive treatment in random sequence in a three-period crossover design with a three-week washout period between treatments: • • •

Group A: SCH-503034 (200 or 400 mg thrice daily) as monotherapy for seven days. Group B: Peg-IFN-α-2b (1.5 µg/kg weekly) as monotherapy for 14 days. Group C: combination of the group A and B protocols for 14 days.

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Results from this study demonstrate that SCH-503034/peg-IFN-α-2b combination therapy is effective in reducing HCV RNA, with four of the ten patients in the 400 mg SCH-503034 thrice-daily combination-therapy cohort achieving undetectable HCV RNA levels during the 14-day course of therapy. The average reduction in HCV RNA was greatest in this cohort, reaching over 2 log10 , while peg-IFN-α-2b and SCH-503034 monotherapies reduced viral load by only 1.1 log10 and 0.5—2.5 log10 , respectively (Zeuzem S, 2005b). Although SCH503034 appears likely to be used in combination therapy with peg-IFN-α, it remains to be seen what impact viral resistance will have on drug development and usage. GS-9132. In November 2004, Gilead Sciences established an agreement with Achillion Pharmaceuticals for the development of GS-9132, a small-molecule inhibitor of the HCV protease. The compound was discovered by researchers at Achillion, where initial preclinical studies were conducted. The agreement grants Gilead the worldwide rights to development and commercialization of GS9132. In August 2005, the companies announced the commencement of the Þrst Phase I clinical trial for GS-9132, currently under way in the United States. This double-blind, randomized, placebo-controlled dose-escalation study will enroll approximately 20 healthy volunteers to evaluate pharmacokinetics, safety, and tolerability of a single escalating doses of GS-9132. Broad-Spectrum Antivirals Overview. Interferons have been the cornerstone of hepatitis C therapy, but their efÞcacy is greatly boosted when an IMPDH inhibitor (namely, ribavirin) is added to the therapeutic regimen. However, ribavirin use is limited by its toxicity. Researchers are investigating antivirals that are effective against HCV via alternative mechanisms of action and that have reduced side effects or contraindications, in comparison with ribavirin. Companies hope that these new antivirals will replace ribavirin as Þrst-line therapy for hepatitis C in combination with immunomodulators. Mechanism of Action. Broad-spectrum antivirals are drugs that work to inhibit the replication of a virus but do not act directly on the virus itself. Celgosivir is a broad-spectrum antiviral that functions by inhibiting α-glucosidase, a mammalian enzyme that facilitates the early-stage glycosylation processes that enveloped virus, such as HCV and HIV, require for assembly, maturation, and infectivity. Amantadine is a broad-spectrum antiviral that has found use in multiple indications, including inßuenza and Parkinson’s disease. Because these drugs are not highly speciÞc for HCV and, in some cases, target host components in the viral life cycle, it is difÞcult to predict the effect these drugs will have against HCV. Amantadine. Amantadine (Endo Pharmaceuticals’ Symmetrel, generics) (Figure 4) is an oral antiviral agent widely marketed for the treatment of infection caused by various strains of inßuenza A. The drug is also indicated in the treatment of Parkinson’s disease and for the treatment of side effects caused by

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NH2 • HCl

FIGURE 4. Structure of amantadine.

certain psychiatric drugs. Although no company is directly developing amantadine for the treatment of chronic hepatitis C, several academic-based clinical trials of amantadine administered in combination with IFN-α (with or without ribavirin) are under way in Europe and the United States. These trials are searching for an effective alternative therapy for patients intolerant of or unresponsive to current recommended therapies. Researchers do not fully understand how amantadine exerts its antiviral effects. It appears that the drug prevents the release of infectious viral nucleic acid into the host cell; it may also inhibit viral replication by preventing viral assembly. In vitro, amantadine has been shown to reduce HCV RNA concentrations, and in vivo the drug is observed to potentiate the early decline in viral titer. In clinical trials, however, the drug has not been useful in eliminating the virus when used as monotherapy. Treatments combining amantadine with peg-IFN-α or conventional IFN-α plus ribavirin can be effective. A study of treatment in nonresponder patients showed that adding amantadine to a conventional IFN-α plus ribavirin regimen boosted response rates (AdinolÞ LE, 2003). Patients receiving the regimen of conventional IFN-α plus ribavirin had an end-of-treatment response of 29%, with only 4% achieving SVR after one year of follow-up. In contrast, 68% of patients who received amantadine, 200 mg/day, in addition to this regimen had a response by the end of treatment and 25% of them achieved an SVR by the end of follow-up (AdinolÞ LE, 2003). A meta-analysis of 31 randomized, controlled studies of amantadine/IFN-α/ribavirin combination therapy in treatment of HCV patients showed that the addition of amantadine raised the overall percentage of nonresponder patients achieving an SVR by 8% but had a negligible impact in treatment of relapsed or treatment-naive patients (Deltenre P, 2004). A study of the outcome of peg-IFN-α/ribavirin/amantadine triple therapy in treatmentnaive patients resulted in SVR rates of 55.2% in patients with HCV genotype 1 infection, a higher SVR percentage than dual or triple therapy with conventional IFN-α (Mangia A, 2005) but within the range of efÞcacy for the peg-IFN-α plus ribavirin regimen in HCV genotype 1-infected patients. In summary, amantadine seems to enhance the effects of the combination of peg-IFN-α and ribavirin in proportion to the patient’s sensitivity to IFN-α, although amantadine is barely effective by itself. It is doubtful that amantadine will be approved for hepatitis C therapy in Europe or the United States because the additive beneÞt of the drug is minimal, at best. Given the drug’s inconsistent efÞcacy when used in triple combination

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FIGURE 5. Structure of celgosivir.

with IFN-α and ribavirin and its marginally improved efÞcacy when used in triple combination with peg-IFN-α and ribavirin, most experts do not see amantadine as the solution for treating relapsers and nonresponders. Celgosivir. In 2004, Migenix acquired the rights to develop celgosivir (MX3253) (Figure 5), a broad-spectrum glycosylation dysregulator, from Virogen. Celgosivir, a pro-drug of castanosperimine (derived from the Australian black bean chestnut tree), is formulated for oral administration and is under investigation for treatment of HCV. In 2005, Migenix reached an agreement with Schering-Plough, wherein Schering-Plough will provide technical and material support (including peg-IFN-α-2b and ribavirin) for Migenix’s combination Phase II clinical trials, in return for limited opportunities for exclusive access to the clinical trial data and options for licensing. In November 2005, Migenix announced initiation of recruitment for a randomized, multicenter, controlled Phase IIb clinical trial to asses the efÞcacy of celgosivir/peg-IFN-α-2b/ribavirin, celgosivir/peg-IFN-α-2b, and placebo/pegIFN-α-2b/ribavirin. The trial will enroll HCV genotype 1-infected nonresponder patients and will last 12 weeks (Migenix, 2005). Antibody Therapies Overview. Several antibody-based therapies are in early stages of development but will likely be positioned in the market as second-line therapy for nonresponders. Antibody therapies may establish a role as part of combination therapy for HCV, either with other antivirals to increase efÞcacy and limit resistance or with current therapeutic regimens to give speciÞcity to treatment. Mechanism of Action. Most of the antibody therapies in development are puriÞed antibodies against the HCV envelope. These antibodies can potentially have two mechanisms of action against HCV infection. First, by binding to the envelope of the virus, antibodies are able to inhibit virus binding to cellular receptors and thereby prohibit entry of the virus into cells. Secondly, antibodybound virus particles can be quickly recognized and cleared by the host’s immune system and may stimulate an immune response against HCV.

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XTL-6865. XTL Biopharmaceuticals’ XTL-6865 (formerly HepeX-C; XTL002) is a combination of two fully humanized monoclonal antibodies against the HCV E2 protein. XTL-6865 had shown some efÞcacy in preclinical models of HCV infection as both a therapeutic and prophylactic agent, and XTL Biopharmaceuticals recently applied for an IND with the FDA to begin Phase Ia/Ib clinical trials of XTL-6865 for protection from reinfection of HCV in liver transplant recipients (XTL Biopharmaceuticals, 2005). Recent data on one of the antibodies, XTL-68, were presented at the 56th AASLD meeting and showed that daily administration of the antibody could effectively reduce viral load during the Þrst week post-transplantation (Schiano T, 2005). Civacir. Nabi Biopharmaceuticals is developing Civacir (HCV immune globulin) for postexposure prophylaxis against HCV infection in liver transplant patients. Civacir consists of puriÞed antibodies against HCV that are isolated from the plasma of HCV-infected blood donors. Nabi collects plasma at its nine collection centers and fractionates it to concentrate neutralizing HCV antibodies. In December 2002, the FDA granted Civacir orphan drug status, a designation that gives Nabi seven years of marketing exclusivity upon FDA approval. Sponsored by the National Institute of Allergy and Infectious Diseases (NIAID), Nabi began a Phase I/II study in April 2002 to study Civacir’s safety and pharmacokinetics in patients who received a liver transplant as a result of HCV infection. Nabi enrolled 18 patients scheduled for liver transplantation in this randomized trial testing two doses of Civacir. The study design called for 17 infusions of Civacir to be given to these patients over 14 weeks after liver transplantation, followed by 8 weeks of follow-up observation. Detailed results from the trial have not been released, although Nabi has reported that general trends in declining ALT levels and viral load in liver tissue were observed (Nabi Pharmaceuticals, 2004). Civacir has since gained orphan medicinal product status in the European Union, a designation that gives it ten years of market exclusivity after launch. HuMax-HepC. Genmab’s HuMax-HepC is in preclinical development for the treatment of HCV. Genmab licensed HuMax-HepC from Connex and INSERM, the French National Institute for Health and Medical Research. Both Connex and INSERM will receive royalty payments on the future sales of HuMax-HepC. HuMax-HepC is a human monoclonal antibody against the E2 envelope protein of HCV. This antibody was identiÞed from a patient with mild chronic hepatitis resulting from HCV infection. HuMax-HepC is reported to cross-react with a wide range of HCV genotypes, including the most common ones, but data demonstrating this activity have not been released. In preclinical studies, HuMax-HepC greatly inhibited binding of the E2 protein to HCV-susceptible cells. No further clinical development has been announced. HCV Vaccines Overview. The development of an HCV vaccine poses a great challenge to the scientiÞc community. The genetic heterogeneity of the virus and its ability to

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establish chronic infection—by evading the immune response or by an inadequate host response—are just some of the difÞculties associated with vaccine development (Forns X, 2002). However, several therapeutic vaccines in development aim to boost speciÞc immune responses against HCV; they may be effective alone or in combination with other therapeutics. Experts express great interest in a prophylactic vaccine that could prevent infection with HCV. They say that a vaccine that protected against the most prevalent and hard-to-treat genotypes of HCV (i.e., genotypes 1 and 4) would quickly be included in pediatric vaccine schedules, much like current HBV vaccines. However, experts are unanimously skeptical of the potential of the prophylactic vaccines in development. Many are concerned that data published to date have not demonstrated the desired efÞcacy, and even more doubt the ability of any prophylactic vaccine to protect against the numerous genotypes of HCV. Mechanism of Action. Therapeutic vaccines aim to induce an immune response against HCV-infected cells. In this manner, their therapeutic aim is similar to that of immunomodulators. However, the key difference between the two classes is the speciÞcity of a therapeutic vaccine’s immune response, targeting it against speciÞc viral proteins. In this manner, a vaccine should be more efÞcacious and less toxic than immunomodulators. An additional advantage of therapeutic vaccines is that the therapeutic effect is delivered by the patient’s immune system; thus, few doses of vaccine are needed to stimulate the process, in stark contrast to the up to 48 weeks of therapy that current immunomodulators require. In chronic HCV infection, the patient’s immune response against the virus is ineffective; otherwise, the virus would have been cleared at the acute stage of infection. Vaccines target the host immune response to boost its recognition and clearance of speciÞc viral proteins that previously were unrecognized by the host. Therapeutic vaccines often consist of puriÞed viral proteins, or peptides from viral proteins, that are likely to be recognized and bound by cytotoxic CD8+ T cells, which play a critical role in viral clearance. HCV vaccines comprise viral epitopes (antigens) delivered, with an immunostimulatory adjuvant, to the patient. The patient’s immune system recognizes the viral proteins and peptides as foreign and, boosted by the adjuvant, produces a robust immune response against them. From this activation, the immune response then recognizes virally infected cells and clears them from the host. If the individual is later exposed to the virus, a potent immune response should be able to block infection. Prophylactic vaccines work almost identically to therapeutic vaccines, but because they are given to very large numbers of healthy individuals, their safety and toxicity proÞles must be extraordinary. According to experts, a prophylactic vaccine with safety concerns would not be approved when reasonably effective therapeutics are available. IC-41. Intercell is developing a peptide vaccine, IC-41, as a treatment for HCV infection. This vaccine consists of Þve HCV peptides (obtained from patients

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who generated a natural protective immune response against HCV) with polyL-arginine as an adjuvant. In November 2002, Intercell initiated Phase II trials of its peptide vaccine in 11 institutions across Europe. The vaccine is being given as six injections over Þve months to patients with chronic HCV who did not respond to IFN-α/ribavirin treatment. Clinical end points determine the vaccine’s safety, efÞcacy, and most viable dosing. The trial was completed in 2004, and IC-41 was shown to be safe and well tolerated. In addition, a TH 1 immune response was observed in some patients and a subset of these patients had a transient reduction in HCV viral load. A Phase II clinical trial including standard peg-IFN-α/ribavirin cotherapy in combination with IC-41 is expected to be completed by early 2006. An additional Phase II clinical trial has begun, enrolling 50 subjects and administering up to 16 injections per patient at weekly intervals. Intercell anticipates Phase III clinical trials to commence in 2008, with a target launch in the European Union in 2011 (Intercell, 2005). HCV ISCOM. Chiron, in collaboration with CSL, is developing a therapeutic HCV vaccine using CSL’s immunostimulatory complex (ISCOM) technology. This adjuvant system is able to prime CD8+ cytotoxic T cells and CD4+ Thelper cells. The vaccine comprises the ISCOM adjuvant with a yeast-derived fusion protein of HCV nonstructural proteins NS3, 4, and 5. In preclinical studies, this vaccine was given to Þve chimpanzees in three vaccinations. Four of Þve animals became positive for cytotoxic T-cell function against HCV proteins, and all Þve had functional CD4+ T-cell responses, as measured by IFN-γ . In preclinical studies, this vaccine was given to Þve chimpanzees in three vaccinations. Four of Þve animals became positive for cytotoxic T-cell function against HCV proteins, and all Þve had functional CD4+ T-cell responses, as measured by IFN-γ secretion, against HCV. CSL is conducting a Phase I trial of this vaccine candidate in Australia. Limited interim clinical reports show that treatment has so far been well tolerated, but full results are not yet available. HCV E1E2/MF59C.1. In addition to the therapeutic HCV vaccine it is developing, Chiron is researching a prophylactic vaccine against the hepatitis C virus. In conjunction with St. Louis University and the National Institutes of Health (NIH), the company has obtained approval to begin clinical trials with an E1E2 (the two HCV envelope proteins) vaccine. This vaccine includes Chiron’s novel adjuvant MF59, which is currently used in the company’s Europe-approved inßuenza vaccine Fluad. Results from a blinded Phase I clinical trial conducted in 60 healthy, HCV-negative adults were presented at the 56th annual AASLD meeting (Di Bisceglie AM, 2005). In this study, 48 patients received four subcutaneous injections of the vaccine at weeks 0, 4, 24, and 48. Three doses of the vaccine were administered (4, 20, or 100 mg), with 16 vaccinated and 4 placebo patients in each dose cohort. Fifty-Þve, 52, and 38 patients received the second, third, and fourth vaccinations, respectively. Immunogenicity was determined by measuring serum titers of anti-E1E2, lymphocyte proliferation, and neutralization of vesicular stomatitis virus (VSV) pseudotype (a research model for HCV that expresses

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HCV-E1E2) by the E1E2 antibodies in serum. All doses of the vaccine produced adequate antibody after the Þrst injection, and binding antibody titers reached similar ranges for all doses by the third injection. Neutralizing antibodies were present in all three dose cohorts by the third injection. All subjects experienced some treatment-related pain, which increased with dosage. REFERENCES Clincaltrials.gov web site. 11-1-2005. www.clinicaltrials.gov. Accessed November 1, 2005. AdinolÞ LE, et al. Effects of alpha interferon induction plus ribavirin with or without amantadine in the treatment of interferon non-responsive chronic hepatitis C: a randomised trial. Gut.2003;52:701–705. AFDHAL N, et al. Colchicine versus PEG-Intron Long-Term (COPILOT) Trial: Interim analysis of clinical outcomes at year 2. 55th AASLD; November 1, 2004; BOSTON, MA. Abstract # 171. Alberti A, et al. Antibody to hepatitis C virus and liver disease in volunteer blood donors. Ann Intern Med.1991;114:1010–1012. Alberti A, et al. Natural history of hepatitis C. J Hepatol.1999;31 Suppl 1:17–24. Alter HJ, Seeff LB. Recovery, persistence, and sequelae in hepatitis C virus infection: a perspective on long-term outcome. Semin Liver Dis.2000;20:17–35. Alter MJ. Epidemiology of hepatitis C. Hepatology.1997;26 Suppl 1:62–65. Alter MJ. Recommendations for prevention and control of hepatitis C virus (HCV) infection and HCV-related chronic disease. Centers for Disease Control and Prevention. MMWR Recomm Rep.1998;47:1–39. Alter MJ, et al. The prevalence of hepatitis C virus infection in the United States, 1988 through 1994. N Engl J Med.1999;341:556–562. Angelico M, et al. Recombinant interferon-alpha and ursodeoxycholic acid versus interferon-alpha alone in the treatment of chronic hepatitis C: a randomized clinical trial with long-term follow-up. Am J Gastroenterol.1995;90:263–269. Ansaldi F, et al. Different seroprevalence and molecular epidemiology patterns of hepatitis C virus infection in Italy. J Med Virol.2005;76:327–332. Aora S, et al. Ascending multiple-dose pharmacokinetics of viramidine, a prodrug of ribavirin, in adult subjects with compensated hepatitis C infection. J Clin Pharmacol.2005;45:275–285. Appel N, et al. Mutational analysis of hepatitis C virus nonstructural protein 5A: potential role of differential phosphorylation in RNA replication and identiÞcation of a genetically ßexible domain. J Virol.2005;79:3187–3194. Armstrong GL, et al. The past incidence of hepatitis C virus infection: implications for the future burden of chronic liver disease in the United States. Hepatology.2000;31:777–782. Bacon BR. Treatment of patients with hepatitis C and normal serum aminotransferase levels. Hepatology.2002;36:S179-S184. Bailon P, et al. Rational design of a potent, long-lasting form of interferon: a 40 kDa branched polyethylene glycol-conjugated interferon alpha-2a for the treatment of hepatitis C. Bioconjug Chem.2001;12:195–202.

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Methicillin-Resistant Staphylococcus aureus ETIOLOGY AND PATHOPHYSIOLOGY Staphylococci are one of the most common bacterial pathogens in humans. They cause a wide spectrum of infections, ranging from minor skin infections to life-threatening bacteremia (bloodstream infection). Methicillin-resistant Staphylococcus aureus (MRSA) is among the most troubling staphylococcal infections. MRSA infection occurs predominantly in the hospital setting, and, over the past several decades, it has become one of the most serious hospital-acquired infections worldwide (Raynor D, 2003). MRSA is endemic in many hospitals and is one of the leading causes of hospital-acquired (HA) pneumonia and surgical site infection and the second-leading cause of HA-bloodstream infections (bacteremia) (Blanc DS, 2002; Boyce JM, 1994). Strains of S. aureus that are resistant to methicillin are typically also resistant to many other Þrst-line antibiotics, including aminoglycosides, macrolides, tetracyclines, ßuoroquinolones, and beta-lactam (β-lactam) antibiotics. Infections caused by MRSA are therefore difÞcult to treat and lead to prolonged hospitalization, increased morbidity and mortality, and higher cost (Plowman R, 2000; Raynor D, 2003; Rubin RJ, 1999). MRSA infections are more prevalent in large teaching hospitals than in smaller community hospitals and are more commonly isolated from patients in intensive care units (ICUs) than in general wards (Blanc DS, 2002). In hospitals, MRSA is also more commonly isolated in certain settings and patient groups such as from patients presenting to dialysis centers and infusion clinics. In addition to the hospital setting, MRSA is found in long-term-care facilities such as nursing homes (Sista RR, 2004). MRSA rates have increased signiÞcantly over the past 15 years—current estimates from regional studies of MRSA incidence range from 29% to more than Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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ETIOLOGY AND PATHOPHYSIOLOGY

721

50% of S. aureus isolated from U.S. hospitals being methicillin-resistant (National Nosocomial Infections Surveillance [NNIS], 2003). MRSA rates differ geographically; MRSA prevalence varies from less than 1% in northern Europe to more than 40% in southern and western Europe. Studies have found that the highest proportions are present in southern and parts of western Europe (Spain, Italy, United Kingdom) and the lowest proportions are in northern Europe (the Scandinavian countries) (Tiemersma EW, 2004). In Japan, more than 50% of S. aureus isolated in hospitals are estimated to be methicillin-resistant (Takeda S, 2000). Although HA-MRSA is by far the more common type of MRSA infection, community-acquired MRSA (CA-MRSA) is increasingly of concern following a recent rise in its frequency. Several outbreaks of MRSA infections have been reported in young, healthy individuals in the community (Stevens D, 2003). A brief review of the history and evolution of staphylococci treatment is warranted to provide context. Prior to the introduction of penicillins in the 1940s, patients with staphylococcal disease had a high mortality rate. The introduction of penicillins brought about a remarkable improvement in outcome in these patients (Daum RS, 1988). However, S. aureus quickly became resistant to penicillin. Subsequently, new antistaphylococcal penicillin derivatives such as methicillin, cloxacillin, and ßucloxacillin were introduced. However, soon after their introduction, S. aureus that was resistant to methicillin and the penicillin derivatives was reported. It is important to note that methicillin is not used in clinical practice; rather, in the past, it was used in laboratory antibiotic susceptibility testing to determine bacterial resistance to antibiotics. Strains that are resistant to methicillin are typically resistant to all beta-lactam agents, including cephalosporins (Sista RR, 2004). When resistance was Þrst described in 1961, methicillin was used to test and treat infections caused by S. aureus; however, methicillin is no longer commercially available in the United States. The antistaphylococcal penicillin oxacillin, which is in the same class of drugs as methicillin, was chosen for testing staphylococci in the early 1990s. Oxacillin (Bristol-Myer’s Squibb’s Prostaphlin, generics) is used because it maintains its activity during storage better than methicillin does. The acronym MRSA, however, is still used to describe the isolates because of its historic role. MRSA is deÞned as an isolate having a minimum inhibitory concentration (MIC)—the concentration of antibiotic required to inhibit growth of the pathogen—of more than 4 µg of oxacillin/mL. Methicillin-susceptible S. aureus (MSSA) is deÞned as an isolate having a MIC of < 2 µg of oxacillin/ mL. Methicillin resistance is important because MRSA strains are typically resistant to all other beta-lactam antibiotics and penicillin derivatives such as cloxacillin and ßucloxacillin (Alborzi A, 2000; Greenwood D, 2000). This resistance limits the use of an entire class of antibiotics (the beta-lactams), a problem that severely conÞnes physicians’ options in treating staphylococcal infections. Etiology S. aureus is an organism that is part of the normal ßora in humans (Lowy FD, 1998). Staphylococci are members of the Micrococcaceae family and appear

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as gram-positive cocci in clusters on a gram stain. The staphylococcal genome consists of a circular chromosome and extrachromosomal elements such as transposons and plasmids. Genes governing virulence and resistance to antibiotics are found on the chromosome as well as on the extrachromosomal elements. These extrachromosomal elements can be transferred from one strain of bacteria to another, resulting in the spread of virulence factors and resistance to antibiotics that provide the organism with a high potential for human pathogenicity. Staphylococci. Staphylococci are among the most common nosocomial pathogens. They are usually found on the skin and mucous membranes of the body (particularly the perianal area, nasal membranes, and hair follicles). Researchers can easily distinguish staphylococci from other clinically relevant gram-positive bacteria, such as streptococci, by using a catalase test. This test measures the presence or absence of the enzyme catalase, which bacteria use as a defense mechanism to convert harmful products into nonharmful products. Staphylococci are catalase-positive, while streptococci are catalase-negative. Two clinically relevant species of staphylococci can be differentiated on the basis of the enzyme coagulase; S. aureus is coagulase-positive, while other strains such as S. epidermis are coagulase-negative. Both coagulase-negative staphylococci (CoNS) and S. aureus can exist as part of the normal human ßora (i.e., exist as commensal organisms). These bacteria can colonize individuals, but they generally cause disease only when the body’s defense systems are compromised. S. aureus colonization is widespread in the general population and the most common sites of colonization are the nares (nostrils). Researchers estimate that 30–50% of healthy adults are colonized with S. aureus and 10–20% are chronically colonized (Sista RR, 2004). Although individual strains of S. aureus differ in their ability to invade and establish infection, infection essentially relies on the capacity of this bacterium to penetrate the skin or mucous membranes (Day NP, 2001; Peacock SJ, 2002). It is able to do so when the skin surface is broken (e.g., in postsurgical or trauma patients), when the mucosal surface is damaged (e.g., chemotherapy-induced mucositis), or when it is introduced into the host via an external source (e.g., catheterized patients). Methicillin-Resistant Staphylococcus aureus. MRSA is a formidable pathogen associated with hospital-acquired infections. As discussed previously, methicillin resistance in the hospital setting often suggests cross-resistance to most beta-lactam antibiotics (such as penicillins), as well as the macrolide, tetracycline, ßuoroquinolone, and aminoglycoside classes of antibacterials (BergerB¨achi B, 1997; Jones RN, 1999). MRSA isolated in the community is generally resistant only to beta-lactams. MRSA leads to signiÞcantly higher morbidity, mortality, and cost than MSSA. (Table 1 highlights the important outcome parameters of MRSA and MSSA.) Mechanisms of Methicillin-Resistance. There are several known mechanisms by which S. aureus becomes resistant to methicillin (Sista RR, 2004). Many strains of S. aureus produce enzymes (beta-lactamase) that prevent the binding of

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TABLE 1. Key Outcome Parameters of Methicillin-Resistant and Methicillin-Susceptible Staphylococcus aureus Outcome Parameters Mortality (in-hospital mortality rate)a Average cost/patientb Antibiotic susceptibility/resistance

Median hospital stayc

MRSA

MSSA

63.8%

23.7%

$92,363 Vancomycin-susceptible

$52,791 Penicillin-derivative-(fluoxacillin-) susceptible Cephalosporin-susceptible

Multidrug-resistant (beta-lactams, penicillin derivatives, cephalosporins) 15.5 days

11 days

a Blot SI, 2002. b Engemann JJ, 2003. c Kopp BJ, 2004.

Full source citations appear in ‘‘References.’’

the beta-lactam antibiotic to the drug target in the cell membrane, the penicillinbinding proteins (PBPs). One mechanism of methicillin resistance in S. aureus is caused by an acquired PBP known as PBP 2a (encoded by the gene termed mecA), which has a low afÞnity for all beta-lactam antibiotics (i.e., penicillin derivatives and cephalosporins [Sista RR, 2004]). Bacterial strains that harbor the PBP 2a are less susceptible to beta-lactam antibiotics than are the bacteria that lack the protein. The mecA gene is not present in methicillin-susceptible strains of S. aureus. The gene is carried on a genetic element called the staphylococcal cassette chromosome mec (SCCmec). Four forms of the SCCmec have been described that differ in size, and their presence varies in CA-MRSA strains versus HA-MRSA strains. Polymerase chain reactions have revealed that SCCmec types I, II, and III are found in HA-MRSA. Unlike hospital strains of MRSA, strains of CA-MRSA carry a different form of SCCmec: type IV. The SCCmec type IV is smaller than types I, II, and III, suggesting the possibility of increased mobility and easier transfer to a methicillinsusceptible strain, which is common in CA-MRSA infections. This characteristic increases the risk of drug-resistant strains developing in the community. Hospital- and Community-Acquired MRSA. The most common deÞnition of CA-MRSA is the presence of MRSA in a sample culture of an outpatient or isolation of MRSA within 72 hours of a patient’s hospitalization (MRSA isolated 72 hours after hospitalization is considered HA-MRSA). However, there are some important caveats and ambiguity concerning this deÞnition of CA-MRSA (discussed further on). In general, CA-MRSA and HA-MRSA differ substantially in clinical severity, genetic makeup, and resistance spectrum (Table 2). MRSA Trends in the Hospital Setting. The prevalence of MRSA in the hospital setting has been rising over the past 15 years, as recorded by the U.S. National Nosocomial Infections Surveillance Survey (NNIS System Report, 2003). More recently, the NNIS data showed that from January 1998 through June 2003,

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METHICILLIN-RESISTANT STAPHYLOCOCCUS AUREUS

26% of all S. aureus isolates from outpatients were methicillin-resistant, 42% of all S. aureus isolates from non-intensive care unit (ICU) inpatients were methicillin-resistant, and 52% of all S. aureus isolates from ICU patients were methicillin-resistant. The incidence of MRSA infections in ICU patients has dramatically increased in recent years. Between 1989 and 2002 in the United States, the proportion of S. aureus infections due to MRSA in ICU patients increased from 29% to more than 60% (www.cdc.gov/ncidod/hip/ARESIST/mrsa.htm). In Western Europe, 30–50% of S. aureus isolates obtained from hospital-based infections are resistant to methicillin. In Japan, the percentage of MRSA among hospital-acquired S. aureus strains is estimated to be 60–70% (Kusachi S, 1999; Takeda S, 2000). MRSA Trends in the Community Setting. Although not the focus of this report, CA-MRSA cases have been reported in recent years. S. aureus infections in the community are mostly caused by MSSA, but several cases of MRSA in the community have recently been reported. However, the deÞnition of CAMRSA is somewhat ambiguous. There is conßict in the literature and among experts interviewed in deÞning CA-MRSA relative to the time of acquisition of MRSA. Some researchers deÞne CA-MRSA as MRSA acquired within 48 hours of hospitalization, whereas others deÞne CA-MRSA as MRSA acquired within 72 hours of hospitalization. In addition, some researchers deÞne CA-MRSA as MRSA acquired by a patient who has not had contact with a hospital setting in the last three months, while some researchers deÞne it as MRSA acquired by a patient without hospital contact for one year. Regardless of the time-based deÞnition, MRSA acquired in the community may be present in patients without predisposing risk factors (as described further on) and in patients with risk factors for MRSA acquisition (immunocompromised patients). Genetic analyses of community-acquired MRSA have revealed that there are predominant clones found in the community that are different from the hospital clones. (Note, as indicated in Table 2, CA-MRSA do not exhibit the same level of multiclass resistance as HA-MRSA.) Pathophysiology As mentioned previously, S. aureus is a commensal organism (an organism that coexists with its host without damaging or beneÞting the host) that is part of TABLE 2. Differences Between Hospital- and Community-Acquired MRSA Attributes Drug resistance Genetic makeup Infection sites Toxins

Hospital-Acquired MRSAa

Community-Acquired MRSAa

Multiple antibiotic classes SCCmec types I, II, III Multiple (skin, wounds, blood, bone) Multiple

Reduced beta-lactam susceptibility SCCmec type IV Most commonly skin infections Enterotoxins, PVL

a Sista RR, 2004.

PVL = Panton-Valentine leukocidin; SCC = Staphylococcal chromosomal cassette.

ETIOLOGY AND PATHOPHYSIOLOGY

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the normal human ßora and colonizes the nares, axillae, pharynx, and skin surfaces. Infections are initiated when a breach of the skin or mucosal barrier occurs, allowing the bacteria to access adjoining tissues or the bloodstream. The containment or spread of an infection depends on the interaction between the S. aureus virulence determinants (discussed further on) and the host’s defense mechanisms and immune system. S. aureus has a diverse arsenal of components and products that contribute to its virulence. Virulence Mechanisms of MRSA. Numerous staphylococcal virulence factors have been identiÞed, including factors that promote colonization, protect the bacteria from the host’s defense systems, and facilitate the disease process. These virulence factors are potential (though not yet exploited) targets for future drug development because they play an important role in advancing disease. Their precise mechanisms and the extent to which these virulence factors contribute to the pathogenesis of various staphylococcal diseases are largely unknown. Virulence factors can generally be separated into two main classes: surfaceassociated factors and secreted factors. Surface-associated factors provide the bacteria with mechanisms for adherence, attachment, and immune evasion. Secreted factors (such as exotoxins) cause tissue destruction and can counteract host cell responses. Adherence of S. aureus to host tissues is a critical step in colonization and subsequent establishment of infection. Investigations into factors that facilitate tissue adherence and intracellular invasion have implicated Þbronectin-binding proteins and other cell-wall adhesins of the MSCRAMM (microbial surface components recognizing adhesive matrix molecule) family (Cucarella C, 2001; Menzies BE, 2002). Also, nearly all clinical isolates of S. aureus have a polysaccharide capsule that likely plays an important role in the pathogenesis of S. aureus infections by inhibiting phagocytosis. In vitro studies have demonstrated that bacterial components such as adhesins and capsules elicit an immune response in the form of antibodies directed against the capsule and activation of phagocytosis by the complement system (Lee JC, 1996). However, the role of antibodies in vivo is poorly understood. Researchers speculate that S. aureus surface-associated antigens have the potential to induce immunity to staphylococcal infections. However, the multiple (eight) serotypes of S. aureus capsules and the poorly understood immune response elicited by them have proven difÞcult for vaccine development. Serotyping studies of staphylococcal isolates from diverse strain collections representing several geographic regions have revealed that, of the eight serotypes, serotypes 5 and 8 together account for 75% of the isolates recovered from humans (Arbeit RD, 1984). Moreover, these two serotypes are prevalent among isolates from clinical infections. As a result, capsular antigens have become target antigens to elicit immune response against and protect against staphylococcal infections. Several antibodies and a vaccine are in development that target the cell surface capsular antigens of S. aureus. A noteworthy toxin-producing virulence factor is the Panton-Valentine leukocidin gene (pvl ), which has been identiÞed speciÞcally in CA-MRSA and has

726

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been associated with necrotic lesions involving the skin and with severe necrotizing pneumonia. The pvl gene is carried on a bacteriophage (a virus that infects bacteria) and produces the cytotoxin PVL. Strains of CA-MRSA carrying the pvl gene are relevant because they are unique to CA-MRSA strains, they are globally distributed, and they are resistant to all beta-lactam antibiotics (Lina G, 1999). Although the roles of several virulence factors in the pathogenesis of MRSA have been elucidated, many more play an unknown role in the infectious process. Elaboration of these virulence determinants occurs in a preprogrammed fashion in vitro by an extensive regulatory network. These regulatory factors integrate environmental parameters to coordinate the expression of several, if not all, virulence genes. Risk Factors. Several risk factors predispose patients to MRSA colonization and infection in the hospital (Table 3). In the community, cases of MRSA have been reported among patients who may or may not have predisposing risk factors. In the next paragraphs, some of the important risk factors for MRSA acquisition are highlighted. Prolonged Hospital/ICU Stay. Risk factors for acquiring MRSA in the hospital setting include prolonged hospital stay and repeated hospitalization (Sista RR, 2004). Hospitalized patients are at a higher risk of being infected with MRSA if

TABLE 3. Risk Factors for Methicillin-Resistant Staphylococcus aureus Carriage and Infection in the Hospital Setting Factor

Comments

Age

• Patients older than 60 years with declining immune systems and multiple comorbid illnesses

Previous colonization

• Previous nasal or cutaneous colonization by MRSA predisposes a patient to developing an MRSA infection

Host factors

• History of hospitalization in the past five years • • • •

Prolonged hospital stay Residence in nursing home Intravenous drug use Invasive medical devices and surgical procedures

• Previous use of antibiotics Chronic medical illness

• Diabetes mellitus • Patient undergoing hemodialysis

Impaired immune function

• Intensive care unit • AIDS • Cancer

ETIOLOGY AND PATHOPHYSIOLOGY

727

they have a preexisting chronic disease (such as diabetes), are undergoing invasive treatment for a chronic condition (such as patients undergoing hemodialysis for kidney failure), have an impaired immune function, or have had prolonged exposure to health care facilities. Patients in a long-term-care facility (such as a nursing home) or ICU are at higher risk for MRSA because of the use of multiple invasive devices and frequent use of broad-spectrum antimicrobials (which promote colonization with hospital pathogens). In addition, patients in ICUs are in direct contact with staff and medical devices that could harbor pathogens, thus putting the patients at risk of acquiring infections for a prolonged period of time. Furthermore, frequent antibiotic use in ICUs selects for drug-resistant pathogens (such as MRSA), and these pathogens are found more often in ICUs than in general wards or the community. Invasive Medical Devices or Procedures. The use of invasive medical devices and surgical procedures increases the risk of colonization with MRSA (Sista RR, 2004). Almost every type of in-dwelling device (e.g., central venous catheter, in-dwelling bladder catheter, endotracheal tube) may become colonized with bacteria, making in-dwelling devices the single most important environmental risk factor for hospital-acquired infections (Crnich CJ, 2001; Kunin CM, 2001). Invasive devices enable the organism to adhere and further facilitate colonization. In-dwelling devices also hinder the host’s defenses, usually by compromising the external skin or mucosal barrier and providing an access point for microbes. In some patients, infection with MRSA could lead to more severe infections such as bacteremia, endocarditis (infection of the endocardium—the innermost layer of the heart’s chambers and valves), disseminated infection, systemic inßammatory response syndrome (sepsis), or multiple organ dysfunction syndromes. Age. Patients older than 65 with a declining immune function, multiple chronic underlying diseases, and prolonged hospital stay are at high risk of MRSA acquisition and colonization. Previous Antibiotics Use. Studies have shown that antimicrobial pressures exerted from the previous use of antibiotics are a risk factor for colonization and infection by antibiotic-resistant pathogens (Graffunder EM, 2002). High usage of antibiotics, along with inappropriate use of antibiotics (e.g., inappropriate dosing, incorrect spectrum of antibiotic) selects for and results in antibiotic-resistant pathogens. Researchers have identiÞed consistent associations and dose-effect relationships that support a causal relationship between MRSA and antimicrobial drug use (Monnet D, 2001). Studies have also shown that patients with MRSA infection had a signiÞcantly longer length of stay in hospitals and were likely to have received prior antimicrobial therapy (Hershow RC, 1992). MRSA Clinical Infections. Infections caused by S. aureus range from skin and soft tissue infections such as acne, boils, carbuncles (infections affecting the skin and subcutaneous tissues), and wound infections to pneumonia, catheter-related

728

METHICILLIN-RESISTANT STAPHYLOCOCCUS AUREUS

bloodstream infections, and endocarditis. If a patient develops a bloodstream infection, such as staphylococcal bacteremia, it could lead to severe infections such as osteomyelitis (infection of the bone) and sepsis (systemic inßammatory response to the bacteria). Infections caused by staphylococcus resistant to antibiotics (such as MRSA and CoNS) have serious clinical consequences and are associated with increased mortality in patients (see Table 1). The distribution of clinical infections differs between HA-MRSA and CA-MRSA. The most common infections caused by HA-MRSA include surgical site infections (SSIs), bacteremia, other skin and soft tissue infections, and lower respiratory tract infections. Urinary tract infections, suppurative arthritis, and osteomyelitis are less common. Most HA-MRSA is resistant to multiple classes of antimicrobial agents such as the aminoglycosides, macrolides, tetracyclines, and beta-lactams. Often, this multiple resistance is indicative of an MRSA infection (Greenwood D, 2000). CA-MRSA infections usually present with skin and soft tissue infections such as impetigo, cellulitis, and boils. Community-acquired MRSA is usually resistant to only the beta-lactam antibiotics and is susceptible to several other low-cost classes of antibiotics. In the following sections, the most common MRSA-associated hospital infections are highlighted. Because HA-MRSA is by far the most common and commercially relevant type of MRSA infection, this discussion is limited to MRSA-associated hospital infections. MRSA Pneumonia. MRSA is increasingly a common cause of ventilator-associated pneumonia (VAP) in ICU patients. A study showed that from 1997 to 1998, 11.5% of all mechanically ventilated ICU patients developed VAP, and 19% of these cases were due to MRSA (Ibrahim EH, 2000). Studies assessing the impact of methicillin resistance on clinical outcomes in staphylococcal pneumonia have shown mixed results. Some studies Þnd increased mortality associated with VAP resulting from MRSA versus MSSA infection whereas other studies show no differences between MRSA- and MSSA-associated pneumonia in ICU patients (Gonzalez C, 1999; Rello J, 1994). Irrespective of the results, hospitals have established evidence-based preventive measures to reduce the risk of VAP. The most common mechanism responsible for hospital-acquired pneumonia is direct aspiration of bacteria that have colonized the mucosal surfaces of the upper respiratory tract (Kollef MH, 1999). Pneumonia that develops within 48 hours of intubation (early onset of VAP) is often a result of aspiration complicating the intubation process and is therefore frequently associated with common endogenous organisms such as S. pneumoniae and H. inßuenzae. In contrast, late-onset VAP is more frequently associated with antibiotic-resistant hospital pathogens, such as MRSA, that colonize patients during prolonged hospitalization. In addition, prior antibiotic therapy predisposes patients to infection with difÞcult-to-treat and drug-resistant bacteria such as MRSA. Late-onset VAP and pneumonia caused by difÞcult-to-treat bacteria such as MRSA are associated with high mortality rates (Kollef MH, 1999).

ETIOLOGY AND PATHOPHYSIOLOGY

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MRSA Bacteremia. Colonization of an in-dwelling intravascular device (IVD) by organisms found on the skin (such as S. aureus) is the major cause of bacteremia. Most colonizing microbes originate from the skin at the insertion site; other colonizers may originate from remote infection sites (carried via the blood to the catheter tip). Contamination of the infusate can occur and is the cause of most epidemic bloodstream infections (Crnich CJ, 2001). Although most IVDs are placed using appropriate sterile techniques, errors can occur, and maintaining sterility for the duration of catheterization is difÞcult, especially in the ICU. For example, when effective bathing is difÞcult, patients are colonized with a higher than normal concentration of skin ßora and other hospital bacteria. SigniÞcantly, most primary bloodstream infections occur as a result of a lapse in patient care procedures rather than as a result of host or environmental factors. S. aureus accounts for 16% of all nosocomial bacteremias and is associated with a high rate of mortality (Edmond MB, 1999). Studies have shown that ICU patients with MRSA bacteremia were associated with more than twice the risk of death compared with patients with MSSA bacteremia (Cosgrove SE, 2003; Whitby M, 2001). Factors associated with increased mortality resulting from S. aureus bacteremia include age above 50, presence of endocarditis, and serious underlying cardiac, neurologic, or respiratory disease. Risk factors associated with the development of bacteremia include the use of invasive devices such as central venous and urinary catheters and nasal colonization of S. aureus (Sista RR, 2004). S. aureus bacteremia is associated with a high frequency of complications, the most serious being endocarditis. One study found that 13% of patients with S. aureus bacteremia had endocarditis (Chang FY, 2003). As they do for MRSA-associated pneumonia, hospitals have evidence-based measures to reduce the risk of catheter-related bloodstream infections. MRSA Surgical Site Infections. Most SSIs occur at the time of operation. Operations involving the peritoneum, peritoneal dialysis, cardiac systems, burns, or eyes provide conditions that promote bacterial colonization and subsequent infection, and are, therefore, more likely to be associated with a postoperative wound infection. S. aureus is a common pathogen in SSIs; an increasing proportion of SSIs are caused by MRSA (NNIS, 2003). Studies have shown that, in the United States, 29% of culture-positive SSIs were caused by MRSA (Manian FA, 2003). Patient-speciÞc and postoperative risk factors for the development of MRSA SSIs include postoperative antibiotic treatment for more than one day and discharge of patients to a long-term-care facility. SSIs caused by MRSA are associated with a higher morbidity and mortality rate than are SSIs caused by MSSA (Mekontso-Dessap A, 2001). Although most SSIs are self-limited and easily treated with local measures (e.g., drainage) and/or antibiotics, some are life-threatening, requiring aggressive surgical debridement to limit tissue loss and preserve life (Nichols RL, 2001). SSIs resulting from

730

METHICILLIN-RESISTANT STAPHYLOCOCCUS AUREUS

MRSA put patients at risk for the development of kidney infections, endocarditis, pneumonia, and osteomyelitis (inßammation of bone) because the infecting bacteria may disseminate to remote sites. Prevention and Control of MRSA. There is considerable debate regarding the effectiveness and practicality of different control measures for MRSA (and infection control in general). In May 2003, the Society for Healthcare Epidemiology of America (SHEA) published evidence-based guidelines designed to reduce the incidence of MRSA in hospitalized patients by focusing on reducing the transmission of MRSA between patients and reducing antibiotic use (Muto CA, 2003). Prevention control measures include surveillance culturing of all patients for the presence of MRSA, rigorous hand washing practices, contact isolation precautions, rational use of antibiotics, and topical microbicides. The Centers for Disease Control and Prevention (CDC) advocates the use of contact isolation precautions to prevent the spread of MRSA. Requirements for contact isolation are a private room for the patient, gloves when touching infective material, masks for coming in close contact with the patient, and gowns if soiling is likely. However, several published reports debated the effectiveness of contact isolation in controlling MRSA transmission (Jernigan JA, 1996; Zafar AB, 1995). The preventive measures of infection control for MRSA include hand washing, the use of gloves, careful linen handling, and environmental cleaning. Fomites (e.g., bed linens, towels, pajamas, dishes) have not been implicated as vectors in the transmission of MRSA, and environmental surfaces, in most instances, are not considered important vectors of MRSA. Effective strategies for curtailing MRSA spread also include limiting the use of antibiotics in the ICU by using evidence-based antimicrobial treatment guidelines and placing formulary restrictions on certain antibiotics (Feucht CL, 2003; Imahara SD, 2003). However, reducing antibiotic use alone has not been shown to reduce the incidence of MRSA infections in hospitalized patients (Wilton P, 2002). New strategies such as the cycling or rotation of antibiotics for empirical therapy have been examined as a method for preventing the development of antimicrobial resistance (Merz LR, 2004). Antimicrobial switching is the alternating of antibiotic classes to minimize the continued exposure and selection pressure of one class. Data suggest that patterns of antibiotic use inßuence the development of resistance (Merz LR, 2004). The concept of switching or cycling seems to have the greatest likelihood of success in the acute care hospital setting, especially in the ICU, where several antibiotics are prescribed for a variety of illnesses. Some studies demonstrate that cycling or switching of antibiotics may affect antibiotic resistance patterns within the ICU and reduce the incidence of antibiotic-resistant infections and infection-related mortality (Moss WJ, 2002). However, evaluation of this approach is hindered by ambiguities in the deÞnition of what is considered switching or cycling, as well as confusion about the drugs to be used and the duration of their use. In addition, the emergence of new resistant organisms during the period of one cycle is likely to reduce therapeutic success in the

CURRENT THERAPIES

731

next cycle (Burke JP, 1998). Evaluating the result of switching drugs is difÞcult because a single antimicrobial agent may select for resistance to many different classes of agents (Friedrich LV, 1999). Physicians believe additional data analysis is warranted to determine the implications and outcomes of this cycling intervention.

CURRENT THERAPIES Overview Methicillin-resistant Staphylococcus aureus (MRSA) is an increasingly prevalent pathogen in hospitals and has become one of the most notorious hospital-acquired infections (HAIs). The incidence of MRSA infections has risen substantially since the 1980s and is a major concern in the intensive care units (ICUs) of many hospitals. Hospital-acquired MRSA (HA-MRSA) infection tends to be more common among particular risk groups, such as patients with chronic illnesses (e.g., diabetes), patients undergoing hemodialysis, and patients with an impaired immune function. MRSA infections are of special concern because they are associated with increased morbidity and mortality, prolonged hospitalization, and increased hospital costs. The vast majority of S. aureus are no longer susceptible to Þrst-line antibiotics, such as ampicillin, because they produce beta-lactamase, which renders the antibiotic ineffective against the pathogen. Antistaphylococcal penicillin derivatives such as methicillin (Bristol-Myers Squibb’s Staphcillin, generics), oxacillin (Bristol-Myers Squibb’s Prostaphlin, generics), and nafcillin (BristolMyers Squibb’s Nafcil, generics), which have been widely available for decades, were developed with activity against S. aureus, but soon after their introduction, S. aureus developed resistance to them. MRSA strains are now resistant to all classes of beta-lactam antibiotics (including early-generation cephalosporins) (Greenwood D, 2000). In addition, MRSA is often cross-resistant to other classes of antibiotics, such as ßuoroquinolones, macrolides, and sulfonamides (Barrett JF, 2004). This problem has prompted physicians to use agents such as the glycopeptides, which were generally reserved as a last line of therapy, to combat staphylococcal infections. One of the most worrisome consequences of rising MRSA rates, and subsequent increases in glycopeptide use, is the development of glycopeptide resistance in other pathogens (e.g., vancomycin-resistant enterococci). Thus, the implications of rising MRSA rates go beyond limited treatment options for the infection itself. Table 4 summarizes the most common antibiotics used to treat MRSA infections. MRSA infection also occurs in the outpatient setting (community-acquired MRSA) and has been reported to be on the rise in the community. Unlike HA-MRSA, community-acquired MRSA (CA-MRSA) is typically susceptible to several classes of non-beta-lactam antibiotics (such as lincosamides). Moreover, most pharmaceutical development has been aimed at the hospital infections, and

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METHICILLIN-RESISTANT STAPHYLOCOCCUS AUREUS

TABLE 4. Current Therapies Used for Methicillin-Resistant Staphylococcus aureus Infections, 2005 Agent Glycopeptides Vancomycin Teicoplanin

Lipopeptides Daptomycin

Oxazolidinones Linezolid

Company/Brand

Daily Dose

Availability

Lilly’s Vancocin, generics Sanofi-Aventis’s Targocid

1 g q12h (IV)

US, F, G, I, S, UK, J

400 mg loading, then 200–400 mg q24h (IV)

F, G, I, S, UK, J

Cubist Pharmaceuticals’ Cubicin

4 mg/kg q24h (IV)

US

Pfizer’s Zyvox/Zyvoxid

600 mg bid (PO); 600 mg q12h (IV)

US, F, G, I, S, UK, J

7.5 mg/kg q12h (IV)

US, F, G, I, S, UK, J

300 mg tid (PO); 600 mg q8h (IV)

US, F, G, I, S, UK, J

150 mg and 300 mg bid (PO); 600 mg q12h (IV)

US, F, G, I, S, UK, J

15 mg/kg q12h (IV)

US, F, G, I, S, UK, J

Streptogramin combinations Quinupristin/ King Pharmaceutidalfopristin cals/Nordic Pharma/Fujisawa’s Synercid Lincosamides Clindamycin Pfizer’s Cleocin/Dalacin/ Sobelin, generics Rifamycins Rifampin Sanofi-Aventis’s Rifadin, generics Dihydrofolate reductase inhibitors Trimethoprim/ Roche’s Bactrim, sulfamethoxazole Monarch’s Septra, (TMP/SMX) generics

Bid = Twice daily; IV = Intravenous; kg = Kilogram; PO = Oral; Qd = Once daily; Qid = Four times daily; Tid = Three times daily. IV dosing: q4h = Every four hours; q6h = Every six hours; q8h = Every eight hours; q12h = Every 12 hours; q24h = Every 24 hours. US = United States; F = France; G = Germany; I = Italy; S = Spain; UK = United Kingdom; J = Japan.

the treatment of HA-MRSA will be the area of highest growth over the forecast period because of the pathogen’s severity and receptivity to novel agents. Preventive Measures to Control MRSA. Because of S. aureus’s declining susceptibility to many antibiotics and the increasing emergence of multidrugresistant strains, many hospitals have introduced preventive measures to curtail the spread of MRSA infections. These hospitals employ antibiotic cycling programs and robust infection control policies. Antibiotic cycling involves switching or cycling of the most commonly used classes/antibiotics to reduce selection pressure and resistance. Infection control programs encourage health care workers to

CURRENT THERAPIES

733

wash their hands often and use contact isolation procedures (isolation of patients with MRSA infections). In addition, hospitals are attempting to prevent or treat MRSA colonization in patients to reduce the risk of MRSA infection. Studies have shown that patients colonized with MRSA are at increased risk for subsequently developing an infection with the organism (Davis KA, 2004). Therefore, reducing colonization (e.g., by treating nasal carriage of the pathogen) is one approach to control outbreaks of MRSA infections in the health care setting and to prevent infection in highly susceptible patients. Likewise, reducing colonization can prevent recurrent infections. Several agents are available to eliminate nasal carriage, including oral and topical antibiotics and topical disinfectants. Oral agents include rifampin and TMP/SMX, which have shown activity in vitro against MRSA colonization, but few of these agents reach sufÞcient concentrations in nasal secretions to be clinically useful (Mulligan ME, 1993). Topical agents deliver higher concentrations of the antibiotic to the colonized site than do oral agents and so are more commonly used. They are used in patients who are colonized with MRSA without an infection. A commonly used topical agent is mupirocin (GlaxoSmithKline’s Bactroban) ointment, which is applied to the nares (nostrils) of the patient and has been shown to clear MRSA from the patient at the end of the treatment course (three times a day for Þve days) with no relapses (Mulligan ME, 1993). In clinical practice, this treatment is reserved for patients who are at high-risk for MRSA acquisition (e.g., patients with recurrent MRSA-associated skin infections, immunosuppressed patients). Topical disinfectant agents such as iodine are also used to eradicate MRSA colonization from invasive devices such as catheters. The use of antibioticimpregnated catheters has reduced the incidence of bloodstream infections and is approved by the FDA. The most extensively studied and used are catheters impregnated with chlorhexidine and silver sulfadiazine (ARROWgard; Arrow International; Reading, PA) and minocycline and rifampin (Cook Bio-Guard Spectrum; Cook Surgical; Bloomington, IN) (Marciante KD, 2003). There is concern, however, regarding the possibility of development of resistance to the antibiotics. Choice of Antibiotic Therapy. Physicians consider several important factors when choosing an antibiotic for MRSA, including speciÞc activity against MRSA and availability of the product in an intravenous formulation (for use in the hospital setting, which is the focus of this report). Patients who are infected with MRSA have limited therapeutic options because MRSA is a multidrug- and often multi-drug-class-resistant organism. Therefore, the susceptibility of MRSA to the agent and the agent’s MRSA-speciÞc activity are important attributes of a drug. Most patients with MRSA infections are hospitalized patients with serious infections. These patients require an antibiotic that will rapidly achieve therapeutic concentrations at the site of infection. For this reason, physicians prefer antibiotics with high bioavailability and availability in an intravenous (IV) formulation. IV antibiotics ensure that patients

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METHICILLIN-RESISTANT STAPHYLOCOCCUS AUREUS

achieve a high serum concentration of the antibiotic, and IV antibiotics are necessary for ventilated patients in the ICU because oral therapy is not practical for these patients. In addition, antibiotics with a long half-life and minimal dosing frequency reduce administration costs in the hospital setting—an issue increasingly important because of the intense pressure to cut costs. Among hospitalized patients who are started on IV antibiotics, agents with high bioavailability can facilitate the switch from IV to oral therapy and enable a shorter hospitalization. In addition, products with bactericidal rather than bacteriostatic activity are considered more desirable because they destroy the pathogen rather than just inhibit its growth (i.e., static activity). In the community setting, physicians prefer oral antibiotics with high bioavailability and tissue penetration, and low dosing frequency because they ensure adequate attainment of serum and tissue concentration, and czencourage compliance. Glycopeptides Overview. Glycopeptides have been the proverbial “safety net” for patients infected with beta-lactam-resistant pathogens such as MRSA. The glycopeptides are active only against gram-positive bacteria; they have no activity against gramnegative organisms. These agents are available only in IV formulation for use in systemic infections. (The oral form is used for gastrointestinal [GI] infections and is not absorbed systemically.) Glycopeptides are generally reserved for patients who have HAIs. Previously, they had been reserved as the last line of defense for treating pathogens that were not susceptible to traditional Þrst-line agents such as the penicillins and beta-lactam antibiotics, but their use has increased in tandem with resistance rates. In recent years, the increased use of glycopeptides has resulted in major consequences, namely the development of highly problematic pathogens, such as vancomycin-resistant enterococci (VRE). The increased use of vancomycin has in part contributed to the spread of resistance to other bacterial species, causing collateral resistance. This spread of resistance has changed antibiotic-prescribing patterns in many hospitals. As a result, vancomycin use in hospitals is restricted to infections caused speciÞcally by multi-drug-class-resistant pathogens such as MRSA. Experts anticipate that the new lipopeptide class will help address this problem because lipopeptides have enhanced efÞcacy against these resistant bacteria. Glycopeptides are relatively well tolerated; the most severe adverse reactions involve the inner ear (dizziness and vertigo), hearing loss, and the kidneys. Studies report hypersensitivity (of which skin rash is the most common allergic reaction) in 5–10% of patients. Other possible side effects of glycopeptides include GI disturbances, muscle pain, blood disorders (e.g., eosinophilia), and disturbances in liver enzymes. Mechanism of Action. Glycopeptides inhibit synthesis of the bacterial cell wall by blocking glycopeptide polymerization. Other mechanisms include altering the permeability of this membrane and directly inhibiting RNA synthesis.

CURRENT THERAPIES

H3N

HO

Me

OH

HO Me

O

735

O

O

O Cl O

O H

HO

OH

Cl H O O H NH −O

HN H

O

O H

H N

Me

N N

H

O H

O

H

N H

H

+

NH2 H

O

2C

H2N

OH HO

OH FIGURE 1. Structure of vancomycin

However, the impact of these effects on bacterial viability is largely unknown. Glycopeptides have a concentration-independent bactericidal activity resulting primarily from the inhibition of cell-wall synthesis. They are considered slowly bactericidal against staphylococci. Pathogens achieve glycopeptide resistance by modifying their bacterial cell wall. Vancomycin. Vancomycin (Lilly’s Vancocin, generics) (Figure 1) is a narrowspectrum, bactericidal glycopeptide antibiotic. Originally developed in the 1950s, vancomycin has been available worldwide since the 1960s and is available in oral and IV formulations. However, oral vancomycin is poorly bioavailable to treat systemic infections and so is limited to some GI infections (pseudomembranous colitis due to Clostridium difÞcile or Staphylococcus aureus-associated enterocolitis). Physicians employ only IV formulations of vancomycin for systemic infections. It requires administration every 12 hours. Vancomycin has been the cornerstone of therapy for serious MRSA infections acquired in the hospital. Among the range of MRSA infections it treats are complicated skin and soft tissue infections (cSSTIs), ventilator-associated pneumonia (VAP), surgical site infections, and bloodstream infections (BSIs). Despite being the most commonly used agent against MRSA, vancomycin’s slow bactericidal activity against S. aureus and its toxicities remain concerns for physicians. Physicians also report requisite monitoring of serum levels in some patients because of variable pharmacokinetics/pharmacodynamic parameters that can lead to toxicity (at high serum concentrations) or serum concentrations lower than the minimum

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METHICILLIN-RESISTANT STAPHYLOCOCCUS AUREUS

inhibitory concentration (MIC). The additional monitoring adds to hospital costs, thereby increasing treatment costs. In addition, vancomycin has shown inadequate penetration into tissues including that of the lung, which is a problem for physicians treating MRSA-associated pneumonia (Cruciani M, 1996). In a randomized, open-label trial, 460 hospitalized adults with MRSA infections were treated with vancomycin (1 g twice daily) or the oxazolidinone linezolid (600 mg twice daily) for 7–28 days (Stevens DL, 2002). Among the patients, cSSTI was the most common diagnosis, followed by pneumonia and urinary tract infection (UTI). At the test-of-cure visit (15–21 days after the end of therapy), clinical cure rates were similar between the two groups; 73.2% of patients in the linezolid group and 73.1% in the vancomycin group. Microbiological success rates were as follows: 58.9% in the linezolid group and 63.2% in the vancomycin group. Both regimens were well tolerated, with similar rates of adverse events. Vancomycin has been associated with several commonly observed adverse reactions. Patients receiving vancomycin have had transient and permanent ototoxic reactions (inner ear disturbances), but specialists attribute most of these cases to excessively high IV doses or to patients with a history of hearing complications. Studies have also associated vancomycin with nephrotoxic (kidney) effects, and experts note that physicians should screen patients for a history of renal complications. Clinical experience shows that vancomycin can cause anaphylactoid reactions in allergic patients, resulting in a condition known as “red man syndrome.” Teicoplanin. Teicoplanin (SanoÞ-Aventis’s Targocid) is a narrow-spectrum, bactericidal glycopeptide similar in structure to vancomycin. First launched in France and Italy in 1988, teicoplanin was available in the other European markets by 1997. Aventis partnered with Fujisawa for distribution and promotion to launch teicoplanin in Japan in 1997. The agent has never been available in the United States, where development was suspended in 1999. Teicoplanin is available in parenteral formulations and has a signiÞcantly longer half-life than vancomycin, allowing once-daily administration. Unlike vancomycin, teicoplanin is available in both intramuscular (IM) and IV formulations. Many thought leaders consider teicoplanin a valuable alternative to vancomycin because of its lower toxicity compared with that of vancomycin (Bucaneve G, 1999). Teicoplanin’s spectrum of activity includes grampositive organisms, including MRSA. A randomized, prospective study was performed to compare the oxazolidinone linezolid with teicoplanin for the treatment of suspected or proven gram-positive infections in an ICU population (Cepeda JA, 2004). In the double-blind, prospective design, 200 patients were randomized to receive IV linezolid (600 mg/12 h) or teicoplanin (400 mg/12 h for three doses then 400 mg/24 h IV). Clinical and microbiological assessments were made daily in the Þrst week and at 8 and 21 days after treatment. Results showed that, at the end of treatment, clinical success was 78.9% in the linezolid group versus 72.8% in the teicoplanin group. Microbiological success rates (70.0% in the linezolid group versus 66.2% in the

CURRENT THERAPIES

737

teicoplanin group), adverse effects, and ICU mortality rates were similar between the two groups. Teicoplanin has a favorable side-effect proÞle, especially when compared with vancomycin; it is less likely to cause nephrotoxicity, ototoxicity, and anaphylactoid reactions (Wood MJ, 2000). Indeed, side effects are relatively uncommon. Allergic reactions, which occur in 2.6% of patients, are the most common side effects associated with teicoplanin. Nevertheless, there remain risks for these adverse reactions, and physicians should carefully weigh the beneÞts of therapy against the potential for complications when considering glycopeptide therapy. Lipopeptides Overview. Similar to the glycopeptides, the newly introduced class of lipopeptides is used to treat severe infections caused by gram-positive organisms such as Staphylococci. The one drug approved within the lipopeptide class, daptomycin (Cubist Pharmaceuticals’ Cubicin), is approved only for cSSTIs caused by susceptible strains of S. aureus and by MRSA. It offers enhanced activity against resistant hospital pathogens such as MRSA. Mechanism of Action. Lipopeptides disrupt the cell membrane of bacteria. The lipopeptide daptomycin inserts itself into the cell membrane, which disrupts membrane integrity. This disruption of the membrane is dose-dependent and clearly correlates with reduced bacterial viability. Unlike glycopeptides, lipopeptides do not inhibit the synthesis of any cell-wall components (Cubist, press release, 2004). Daptomycin. Daptomycin (Cubist Pharmaceuticals’ Cubicin) is the Þrst agent to be introduced in the lipopeptide class. Eli Lilly discovered daptomycin in the early 1980s and licensed the worldwide rights to Cubist in 1997. The agent reached the U.S. market in 2003 and is available only in parenteral formulation. Cubist continues to develop daptomycin for the European and Japanese markets. Daptomycin is indicated for use in cSSTIs. It is in Phase III trials to treat infections such as endocarditis, vancomycin-resistant enterococci infections, UTIs, community-acquired pneumonia, and BSIs. Daptomycin has a novel mechanism of action that rapidly kills gram-positive bacteria by compromising bacterial membrane function. It has activity against most clinically relevant gram-positive bacteria and, most importantly, has excellent efÞcacy against bacteria resistant to methicillin, vancomycin, and linezolid. Being a fairly new drug with limited usage experience among physicians, daptomycin is used as a second-line therapy for patients with MRSA-associated cSSTIs. Nevertheless, its bactericidal activity and new mechanism of action are signiÞcant advantages in treating critically ill patients and may keep resistance to the agent at bay. Two randomized Phase III trials evaluated daptomycin in 1,092 adult patients with gram-positive cSSTIs (Arbeit RD, 2004). The studies compared daptomycin

738

METHICILLIN-RESISTANT STAPHYLOCOCCUS AUREUS

(4 mg/kg IV once daily) with vancomycin (1 g IV twice daily) or a semisynthetic penicillin (4–12 g IV four times daily). The efÞcacy end point for daptomycin was clinical equivalence to the comparator in each trial. Daptomycin met this primary end point in both studies; it had a clinical success rate of 83.4%, compared with 84.2% for the comparators among 902 clinically evaluable subjects. The clinical success rate by bacteriological eradication indicated a success rate of 75% compared with 69.4% for the comparators. According to the Cubicin package insert, some patients in clinical trials experienced elevations in serum creatine phosphokinase (CPK), an event that may be associated with skeletal muscle effects; the package insert states that physicians should monitor CPK levels for abnormalities and observe patients for symptoms of myopathy. During clinical trials, other adverse reactions associated with daptomycin included constipation (6.2%), nausea (5.8%), local injection site reactions (5.8%), and diarrhea (5.2%). Bacteriologic investigations in clinical trials isolated very few daptomycin-resistant bacteria (2 of >1,000 patients or less than 0.2%). The mechanism of resistance to daptomycin remains unknown and no study has determined whether resistance is transferable. Oxazolidinones Overview. The oxazolidinones are a novel antimicrobial class on the market. The one marketed oxazolidinone, linezolid (PÞzer’s Zyvox/Zyvoxid), is available both orally and parenterally and has a bioavailability of 100%. The development of oxazolidinones helped address the need for antibiotics effective against resistant gram-positive bacteria, such as MRSA. It also helped address physician concern about the toxicity and resistance patterns of glycopeptides. The oxazolidinone class offers physicians an alternative to glycopeptide therapy, thereby reserving glycopeptide use and lowering the risk of the development of glycopeptide-resistant pathogens. Furthermore, the fact that linezolid is available in oral formulation facilitates step-down therapy, enables the discharge of patients earlier, and potentially reduces health care costs. Oxazolidinones are of particular importance because of their activity against MRSA. They also demonstrate signiÞcant activity against all gram-positive cocci, including strains resistant to methicillin, vancomycin, macrolides, tetracyclines, aminoglycosides, ßuoroquinolones, and sulfonamides. However, experts view their bacteriostatic activity against staphylococci as a disadvantage for serious MRSA infections, which warrant bactericidal activity. Mechanism of Action. Oxazolidinones bind to a site on the bacterial 23S ribosomal RNA of the 50S subunit, preventing the formation of a functional 70S initiation complex, an essential component of the bacterial translation process. Existing studies have not demonstrated any direct action on DNA or RNA synthesis. Linezolid. Linezolid (PÞzer’s Zyvox/Zyvoxid) (Figure 2) was launched in the United States in 2000 and is now available in all seven countries under study

CURRENT THERAPIES

739

O O

N

N

O

O HN

F

CH3

FIGURE 2. Structure of linezolid

(United States, France, Germany, Italy, Spain, United Kingdom, and Japan). The product was launched by Pharmacia, prior to the company’s acquisition by PÞzer. Available in IV, injection, tablet, and oral suspension formulations, linezolid is approved for MRSA infections such as VAP and cSSTIs (such as diabetic foot infections). Linezolid’s high cost has somewhat limited its potential use, and some physicians state that it is reserved for second-line therapy in MRSA infections—particularly in the European markets. A retrospective analysis was conducted on data from two prospective, randomized, double-blind studies from 134 sites involving a total of 1,019 patients with S. aureus nosocomial pneumonia and documented MRSA pneumonia (MRSA sub set) (Wunderink RG, 2003). The patients received 600 mg linezolid or 1 g vancomycin twice daily for 7–21 days, each with the antibiotic aztreonam (BristolMyers Squibb’s Azactam). Outcome was measured by survival and clinical cure rates (assessed 12–28 days after the end of therapy). Clinical cure rates for the MRSA subset were 59.0% for linezolid/aztreonam compared with 35.5% for vancomycin/aztreonam. Another study evaluated the efÞcacy of linezolid and vancomycin in the treatment of MRSA-associated cSSTIs (Stevens DL, 2002). One group of 240 patients received 600 mg of linezolid twice daily (Þrst intravenously, then orally), while another group of 220 patients received 1 g of vancomycin twice daily. Among clinically evaluable patients with proven MRSA, 73.2% of the linezolidtreated group and 73.1% of the vancomycin-treated patients achieved clinical cure. Clinical studies have revealed some hematological abnormalities (e.g., thrombocytopenia, anemia) that are consistent with mild, reversible, duration-dependent myelosuppression in patients receiving linezolid (Gerson SL, 2002). This suppression of bone marrow activity limits prolonged use of linezolid. In particular, physicians must monitor blood chemistries and platelets in patients who have received more than two weeks of linezolid therapy. This side effect could be an issue in MRSA infections because patients can be treated for two weeks or longer depending on the severity of the underlying infection—particularly considering that these patients are already high-risk and often immunosuppressed. According to the Zyvox package insert, other adverse reactions to linezolid include diarrhea (8.3%), headache (6.5%), and nausea (6.2%). Physicians must monitor patients who have recurrent nausea or vomiting for lactic acidosis. Other side effects are relatively benign and are similar to those reported for other antibiotics.

740

METHICILLIN-RESISTANT STAPHYLOCOCCUS AUREUS

Lincosamides Overview. The lincosamide class of drugs includes clindamycin, an antibiotic commonly used in the treatment of skin infections. Lincosamides are active against gram-positive cocci, including MRSA. Physicians use lincosamides in combination with other antibiotics to provide gram-positive coverage. They are primarily used to treat less severe skin infections associated with MRSA that are identiÞed in the community. Mechanism of Action. Lincosamides bind to the 23S ribosomal RNA (rRNA) in the 50S subunit of the ribosome. This binding inhibits the translocation of RNA during protein synthesis and blocks initiation of polypeptide formation. Lincosamides are bacteriostatic but can be bactericidal against highly susceptible bacteria or at high concentrations. They have a long post-antibiotic effect that is sufÞcient to allow therapeutic levels to fall below the MIC. Clindamycin is also used to treat serious infections because it blocks protein production. Physicians often add the agent to a therapeutic regimen for patients at risk of toxic shock to curtail the production of bacterial toxin. Clindamycin. Clindamycin HCl (PÞzer’s Cleocin/Dalacin/Sobelin, generics) (Figure 3) has been available in all the major markets since 1970. Today, many companies offer generic versions of the agent. Clindamycin is indicated for skin infections caused by staphylococci, although physicians usually reserve it to treat suspected gram-positive bacteria in patients allergic to penicillins. Clindamycin is also often the treatment of choice for patients with hypersensitivity to penicillins in the community. Suspected infections with S. aureus or MRSA in the outpatient setting are usually localized skin infections and are treated with the convenient oral forms of clindamycin. In general, MRSA strains isolated in the community setting are susceptible to clindamycin, but increasing reports of pathogen resistance to clindamycin have limited its use. The constitutive expression of the macrolidelincosamide-streptogramin B (MLSB) erm resistance phenotype in S. aureus can limit the antibiotic’s bactericidal effect by conferring resistance to the agent. Strains resistant to erythromycin could impart resistance to clindamycin, thus limiting its use for the treatment of infections caused by MRSA, most of which are resistant to erythromycin (Clarebout G, 2001; Hamilton-Miller JM, 2000).

CH3 CH3 N

HCCl CONHCH H HO

CH3CH2CH2

O OH

H

SCH3 OH

FIGURE 3. Structure of clindamycin

CURRENT THERAPIES

741

Clindamycin has good bioavailability (90%), and the drug is rapidly absorbed when taken orally. It is available in both oral and parenteral forms. Clindamycin is widely distributed in body ßuids and tissues; serum levels exceed the MIC for most indicated organisms, and excretion is primarily via the kidneys. A retrospective study of hospitals in Minnesota identiÞed a total of 354 patients (median age, 16 years) with community-acquired MRSA infection (Naimi TS, 2001). Most case patients (299 [84%]) had skin infections, and 103 (29%) were hospitalized. The researchers determined that 93% of isolates were susceptible to clindamycin, suggesting a role for clindamycin in the treatment of MRSA infections in the community setting. GI disturbances and generalized hypersensitivity reactions (e.g., maculopapular skin rash) are the most frequently reported adverse reactions to clindamycin. Despite the mild nature of most clindamycin side effects, physicians must monitor patients for symptoms of pseudomembranous colitis, a GI complication associated with many antibiotics that can range in severity from mild to life-threatening. Rifamycins Overview. The rifamycins are a family of antibacterial antibiotics produced by the soil bacterium Streptomyces mediterranei. After being chemically modiÞed to make them orally active, they have found widespread clinical use in the treatment of tuberculosis and AIDS-associated mycobacterial infections. However, rifamycins are not commonly used as an antibacterial class because of their metabolic interactions with numerous other drugs. Mechanism of Action. Rifamycins inhibit transcription and translation processes. Their mechanism of antibacterial action involves the inhibition of a single target enzyme—the b subunit of RNA polymerase. The formation of a drugenzyme complex inhibits the initiation of chain formation in the process of RNA synthesis. Interestingly, the existence of rifamycin-inactivating enzymes has not been reported, and antibiotic resistance mechanisms rely solely on mutagenesis of the unique target. Rifamycin-related mutations, insertions, and deletions in bacterial RNA polymerase commonly occur. As a result, resistance to antibiotics in this family (such as rifampin) has developed at a faster pace than for other antibiotic families (Spratt BG, 1994). Rifampin. Rifampin (SanoÞ-Aventis’s Rifadin, generics) (Figure 4) is a semisynthetic antibiotic derivative of rifamycin that has been available in the major markets since the 1970s. It binds to the RNA polymerase and prevents translation of proteins. Rifampin is used primarily as adjunctive (i.e., combination) therapy in difÞcult-to-treat staphylococcal infections, including MRSA infections. It is not commonly used for MRSA and is generally not used alone because of the risk of resistance developing against it. When they do use rifampin, physicians indicate that they combine it with at least two other drugs, such as gentamicin and vancomycin. Rifampin is occasionally used in the treatment of serious MRSA infections in combination with vancomycin to obtain better tissue penetration. It is also used

742

METHICILLIN-RESISTANT STAPHYLOCOCCUS AUREUS

CH3 CH3 HO CH3COO H3C CH3O H3C

CH3

OH O OH OH

O O CH3

CH3 NH CH

O

OH

N N N CH3

FIGURE 4. Structure of rifampin

in combination with other antibiotics to prevent colonization of MRSA, thereby reducing the occurrence of MRSA-associated infections. However, because of the development of resistance against the agent and its numerous drug interactions, its use in MRSA treatment is limited. Rifampin exhibits in vitro activity against S. aureus and MRSA; however, its safety and effectiveness in treating clinical infections due to these microorganisms have not been established in adequate and well-controlled trials. Likewise, the use of rifampin in the treatment of serious staphylococcal infections such as endocarditis remains an unsettled issue (Sande MA, 1983). One small randomized trial compared vancomycin monotherapy with vancomycin plus rifampin for the treatment of MRSA-associated endocarditis (Levine DP, 1991). The researchers found slow clearance of bacteremia in both groups and no trend toward beneÞt for the combination. A randomized, double-blind, multicenter comparative trial compared the combination of the antibiotic novobiocin plus rifampin with trimethoprimsulfamethoxazole (TMP/SMX) plus rifampin to determine the efÞcacy of each regimen in eradicating MRSA colonization (Walsh TJ, 1993). The two treatment arms consisted of 500 mg novobiocin given orally twice daily, plus one 300 mg rifampin capsule twice daily, for seven days; or TMP/SMX, 160 and 800 mg, respectively, twice daily, plus one 300 mg rifampin capsule twice-daily, for seven days. Of the 126 individuals enrolled in the study, 94 (80 patients; 14 hospital personnel) were evaluable. Of this group, 67% of those receiving novobiocin plus rifampin experienced successful clearance of the colonizing MRSA strains compared with 53% of subjects treated with TMP/SMX plus rifampin. Results suggest that rifampin is effective in eradicating MRSA colonization. The most frequent adverse effects of rifampin are GI in nature (nausea, vomiting) and discoloration of body ßuids. Because rifampin is a potent enzyme inducer of the cytochrome P-450 oxidase system, its administration is associated with numerous drug interactions.

CURRENT THERAPIES

743

FIGURE 5. Structure of trimethoprim

Dihydrofolate Reductase Inhibitors Overview. The dihydrofolate reductase inhibitors are primarily used to treat less serious MRSA infections isolated in the community setting. A combination of the classes sulfonamides and diaminopyrimidines, these agents are bactericidal against S. aureus. The most common agent in this class is the widely available generic agent trimethoprim/sulfamethoxazole (TMP/SMX). The agent is not recommended for the treatment of serious infections associated with MRSA (such as endocarditis) because of its poor efÞcacy compared with that of vancomycin. Rather, it is used in limited cases for MRSA-associated UTIs. Mechanism of Action. Sulfonamides and diaminopyrimidines indirectly interfere with both bacterial replication and protein synthesis. These drugs inhibit two consecutive steps in the synthesis of bacterial folic acid (an essential coenzyme in the biosynthesis of certain nucleic acids and amino acids, among others). Trimethoprim/sulfamethoxazole. TMP/SMX (Roche’s Bactrim, Monarch’s Septra, generics) is a combination of trimethoprim (Figure 5), a diaminopyrimidine, and sulfamethoxazole (Figure 6), a sulfonamide. It is widely used as an inexpensive antibacterial agent for the treatment of community-acquired nonserious MRSA infections as well as other infections such as UTIs. It is also used in combination with agents such as rifampin and the topical antibiotic mupirocin to eradicate MRSA colonization in patients. TMP/SMX is bactericidal against S. aureus and inhibits bacterial replication. Clinicians have used TMP/SMX in the past as an alternative to penicillins or vancomycin in the treatment of S. aureus infection and colonization. Although

FIGURE 6. Structure of sulfamethoxazole

744

METHICILLIN-RESISTANT STAPHYLOCOCCUS AUREUS

TMP-SMX has demonstrated fair clinical efÞcacy against MRSA, it is not commonly used to treat serious MRSA infections. In a randomized double-blind comparative trial of 228 IV drug users, 101 had S. aureus-associated skin infections and were included in the efÞcacy analysis (43 received TMP/SMX and 58 received vancomycin) (Markowitz N, 1992). Infections were cured in 98% of vancomycin recipients and in 86% of TMP/SMX recipients. The side-effect rates were similar for TMP/SMX (23%) and vancomycin (20%) recipients; TMP/SMX was associated with nausea and vomiting and vancomycin with inßammation at the IV site. Forty-four percent of TMP/SMX recipients and 29% of vancomycin recipients experienced side effects in the efÞcacy cohort. TMP/SMX is generally well tolerated, but sulfonamides can produce a variety of untoward effects that are due partly to allergy and party to direct toxicity. These adverse reactions, such as nausea, vomiting, anorexia, rash, urticaria, and photosensitivity, occur in 10% or more of patients. EMERGING THERAPIES The treatment of methicillin-resistant Staphylococcus aureus (MRSA) infections represents an area of relatively high unmet medical need and signiÞcant opportunity for drug development. MRSA infection is a relatively common occurrence in both health care and community settings. Vancomycin is the mainstay of treatment for MRSA, but the drug has several drawbacks (e.g., suboptimal activity against staphylococci, variable pharmacokinetics). The introduction of other antibiotics that are active against MRSA (e.g., linezolid [PÞzer’s Zyvox], daptomycin [Cubist’s Cubicin]) has expanded physicians’ options, but these agents also have important drawbacks related to side effects (e.g., myelosuppression). Finally, the recent identiÞcation of vancomycin-resistant Staphylococcus aureus (Chang S, 2003) and vancomycin-intermediate-resistant Staphylococcus aureus (Witte W, 2004) poses an additional threat. Thus, the MRSA treatment market is one characterized by signiÞcant need for additional therapeutics. Developing drugs for infections caused by drug-resistant organisms such as MRSA is very challenging. Antibiotic products are not typically developed against a pathogen(s) but rather are developed for a clinical indication (e.g., hospitalacquired pneumonia). Therefore, companies are required to conduct multiple trials in several indications to attain a broad and commercially viable label. Attaining a label for resistant infections is made more complex by the need for companies to enroll sufÞcient numbers of patients with resistant organisms—translating into potentially large and prolonged clinical trials. An additional market challenge for drug developers in this Þeld is that, historically, hospitals have restricted the use of new products with novel mechanisms of action and/or activity against resistant pathogens (e.g., quinupristin/dalfopristin [Synercid], linezolid [Zyvox]) to prevent the development of resistance and reserve their utility. This practice has limited their uptake to niche segments and reduced their overall commercial potential. Several attributes enhance a product’s clinical utility and potential success in this market. First, it must have high activity against staphylococci (both MRSA

EMERGING THERAPIES

745

and methicillin-susceptible S. aureus [MSSA]) and gram-positive pathogens in general because empiric therapy is commonplace. A new agent must be available in intravenous (IV) formulation because the majority of patients are hospitalized and many MRSA-infected patients are mechanically ventilated, but to facilitate step-down therapy and accelerate discharge of patients with less severe MRSA infections, physicians would also like an oral formulation. Because MRSAassociated infections are serious, clinicians prefer that a new agent have bactericidal (it destroys the pathogen) rather than bacteriostatic (it just inhibits its growth [i.e., static activity]) activity. In addition, because the majority of MRSA patients are already on multiple medications, agents with minimal drug interactions are at an advantage. Even if emerging therapies achieve some or all of these characteristics, they still face a challenge in replacing current therapies because of the latter’s low cost and physician familiarity. The majority of antibacterial agents in late-stage development for the treatment of MRSA are agents or derivatives from currently marketed classes. Some drugs in development, such as the novel glycopeptides, are narrow-spectrum drugs that will likely be used to treat serious infections caused by MRSA in deÞned at-risk patient populations (e.g., patients in the ICU setting, patients undergoing surgery, patients with pneumonia). Other drugs in development are broad-spectrum agents with activity against MRSA; they have the potential for much broader use across the hospital setting. These agents offer enhanced activity against MRSA, improved formulation and dosing, and better penetration into tissues such as the lung. An additional novel class of emerging therapies that may have an important impact on the treatment of MRSA is the antibody therapies. Antistaphylococcal antibodies are likely to be used in combination with other classes of drugs and have the potential to improve survival in cases of severe, life-threatening infections. Finally, one vaccine in late-stage development has the potential to prevent S. aureus and/or reduce the severity of infection. Table 5 summarizes the drug therapies in development for MRSA. Glycopeptides Overview. Glycopeptides such as vancomycin are the mainstay of treatment for MRSA infections. However, physicians note, vancomycin has suboptimal activity against staphylococci, demonstrates poor tissue penetration into the lung (Cruciani M, 1996), and requires careful monitoring in patients with impaired renal function. In addition, researchers have identiÞed S. aureus isolates that are resistant to vancomycin, raising concerns about the emergence of vancomycin-intermediate-resistant S. aureus and vancomycin-resistant S. aureus. Another drawback of vancomycin is the development of resistance in other hospital pathogens such as the enterococci (i.e., vancomycin-resistant enterococci [VRE]). All of these limitations represent areas for potential improvement. The glycopeptides that are in late-stage development demonstrate increased activity against MRSA compared with vancomycin and broad activity against other resistant hospital pathogens. These agents also have better dosing and side-effect

746

METHICILLIN-RESISTANT STAPHYLOCOCCUS AUREUS

TABLE 5. Emerging Therapies in Development for Methicillin-Resistant Staphylococcus aureus Infections, 2005 Compound Glycopeptides Dalbavancin United States Europe Japan

Development Phase

Marketing Company

PR II —

Vicuron Pharmaceuticals Vicuron Pharmaceuticals —

Oritavancin (LY-333328) United States Europe Japan

III II —

Targanta Targanta —

Telavancin United States Europe Japan

III — —

Theravance — —

Cephalosporins BAL-5788 United States Europe Japan

III III —

Basilea Pharmaceutica/Johnson & Johnson Basilea Pharmaceutica/Johnson & Johnson —

Dihydrofolate reductase inhibitors Iclaprim United States — Europe II Japan —

— Arpida —

Topoisomerase inhibitors NXL-101 United States Europe Japan

— Novexel —

— PC —

a Tigecycline is in Phase III trials for hospital-acquired pneumonia. b An acquisition of Peninsula Pharmaceuticals by Ortho-McNeil Pharmaceutical is pending. Prior to comple-

tion of the acquisition, Peninsula plans to spin out PPI-0903 into Cerexa, Inc. c An oral formulation of ramoplanin is in Phase III trials for the treatment of infections such as vancomycin-

resistant enterococci (VRE) infections and Clostridium difficile-associated diarrhea (CDAD). PC = Preclinical (including discovery); PR = Preregistration. Note: Development phase is based on databases such as R&D Focus, R&D Insight, Pharmaprojects, and the Investigational Drugs Database (IDdb3); periodicals such as Scrip,the FDC’s Pink Sheet,and Marketletter; company reports and press releases; and industry contacts.

proÞles than vancomycin. However, all of the agents are available in parenteral form only. Mechanism of Action. Glycopeptides inhibit biosynthesis of the bacterial cell wall by blocking glycopeptide polymerization. This effect immediately inhibits cell-wall synthesis and causes secondary damage to the cytoplasmic membrane of the bacterial cell. Glycopeptides also alter the permeability of the cytoplasmic membrane and directly inhibit RNA synthesis. However, the

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impact of these effects on bacterial viability is largely unknown. Glycopeptides have a concentration-independent bactericidal action resulting primarily from inhibition of cell-wall synthesis. They are considered slowly bactericidal against staphylococci. Dalbavancin. PÞzer (formerly Vicuron Pharmaceuticals) is developing dalbavancin *, a second-generation injectable glycopeptide for gram-positive infections. Dalbavancin’s mechanism of action is the same as that of the glycopeptide class—inhibition of bacterial cell-wall synthesis—but it is a chemically modiÞed version of a naturally occurring glycopeptide that is speciÞcally designed to have better activity against MRSA than either vancomycin or teicoplanin. Dalbavancin’s primary advantage over current glycopeptide antibiotics is its long half-life, which enables once-weekly dosing. In vitro studies have shown that dalbavancin is more potent than vancomycin and teicoplanin against gram-positive bacteria, including MRSA (Jones RN, 1999; Malabarba A, 1998). Dalbavancin is also more potent than Q/D against most gram-positive bacteria, and it is as potent as Q/D against methicillinsusceptible S. aureus (MSSA). In animal models of infection, a single dose of dalbavancin is reported to be as effective as four doses of vancomycin for the treatment of MRSA (Candiani GP, 2001; Jabes D, 2001). Dalbavancin also proved to be more potent than teicoplanin or vancomycin in murine models of staphylococcal septicemia and staphylococcal endocarditis (Malabarba A, 1998). Vicuron began two randomized, double-blind, Phase III clinical trials in December 2002 and announced their results in August 2004 (Versicor, press release, August 12, 2004). In a double-blind trial, 854 patients with complicated skin and soft tissue infections were randomly assigned to receive dalbavancin (1 g IV on day 1 and 500 mg IV on day 8) or standard dosing of linezolid for 14 days; 88.9% of dalbavancin patients showed a clinical response at follow-up visit versus 91.2% of patients taking linezolid. Dalbavancin was reportedly well tolerated. The second trial was an open-label study involving 156 patients with skin infections suspected or conÞrmed to be due to MRSA. Patients receiving dalbavancin had an 89.9% rate of clinical response compared with 86.7% for patients receiving vancomycin. Dalbavancin fared even better in the intent-to-treat group, with a lower discontinuation rate: 86% of patients receiving dalbavancin showed a clinical response compared with only 65.3% of patients receiving vancomycin. In September 2002, Versicor announced positive Phase II clinical trial results for dalbavancin in the treatment of skin and soft tissue infections (Versicor, press release, September 5, 2002). The randomized clinical trial enrolled 62 patients with infections that involved deep skin structures or required surgical intervention, such as abscesses, infected ulcers, burns, and cellulitis. Dalbavancin administered once weekly for two weeks had higher clinical and microbiological response rates than did other standard care regimens, including vancomycin, which was given for a mean duration of 15 days (Seltzer E, 2003). In a Phase I clinical trial involving 12 patients, dalbavancin administered as a once-daily, 30-minute IV infusion demonstrated linear kinetics with low

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interindividual variability and good tolerability (Leighton A, 2001[a]; White RJ, 2000). The drug had a serum half-life of 6.6–7.8 days; signiÞcant drug accumulation was observed following multiple doses. Patients who received 360 mg of dalbavancin exhibited bactericidal activity even when their serum was diluted eightfold 24 hours after dosing. Indeed, seven days after a single dose, plasma levels remained above the minimum bactericidal concentration (MBC 90) required for eradication of 90% of the MRSA isolates. These data support the use of dalbavancin in a wide range of dosing regimens, including once-weekly therapy (Versicor, press release, November 28, 2001). Researchers found once-weekly dalbavancin was superior to twice-daily vancomycin in the treatment of catheter-related bloodstream infections (Seltzer E, 2004). Conducted in North America, the comparative, open-label Phase II trial enrolled 67 patients with catheter-related bloodstream infections due to a suspected or identiÞed gram-positive organism. Patients were randomized to receive two doses of dalbavancin one week apart or vancomycin twice daily for 14 days. The overall response rate was higher with dalbavancin (87%) than with vancomycin (50%). Clinical and microbiological response rates were similar to the overall response rates in both treatment groups. In all clinical studies to date, dalbavancin has proved well tolerated whether dosed daily or weekly. No adverse events were considered clinically signiÞcant, although fever and headache were reported most frequently. Notably, no inner ear toxicities (either auditory or vestibular) have been detected for any dose of dalbavancin (Leighton A, 2001[b]). If approved, dalbavancin will be the Þrst once-weekly, injectable antibiotic on the market and could become the standard of care for serious skin and bloodstream infections caused by staphylococci such as MRSA. This possibility is strengthened by dalbavancin’s superior antistaphylococcal potency and improved tissue penetration, as well as its longer serum half-life compared with that of vancomycin. The availability of a once-weekly antibiotic not only will enable more ßexible and convenient dosing regimens but also will encourage shorter hospital stays. Experts caution, however, that a potential problem with this long-acting drug is hypersensitivity and/or toxicity; its extended half-life means that the drug persists in a reactive patient’s system for an extended period of time. In addition, because its mode of action is similar to that of vancomycin, formulary committees may restrict the use of dalbavancin for fear of the development of resistance. Oritavancin. Targanta Therapeutics is developing oritavancin (LY-333328), a parenteral derivative of the semisynthetic glycopeptide antibiotic LY-264826 is being developed for the treatment of gram-positive bacterial infections, primarily vancomycin-resistant pathogens (staphylococci and enterococci). Oritavancin’s mechanism of action is the same as that described in the “Mechanism of Action” section for the glycopeptide class. Oritavancin is in Phase III clinical trials for the treatment of complicated skin and soft tissue infections in the United States and in Phase II trials for nosocomial pneumonia and bloodstream infections in Europe.

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In vitro studies demonstrated that oritavancin has rapid bactericidal activity against all staphylococci, including MRSA (Jones RN, 1997). Its potency against these organisms was comparable or superior to that of vancomycin, teicoplanin, some ßuoroquinolone agents (e.g., moxißoxacin, ciproßoxacin), the carbapenem imipenem, linezolid, and Q/D (Fasola E, 1996; Schwalbe RS, 1996). The drug is also active against bacteria that are not susceptible to vancomycin or teicoplanin (Biavasco F, 1997). Oritavancin in combination with ampicillin-sulbactam displayed bactericidal activity against several vancomycinintermediate-resistant S. aureus strains (Hershberger E, 1999). In animal infection models, derivatives of LY-264826 were effective against lethal S. aureus infections at doses 1.5–3 times lower than effective vancomycin doses (Nicas TI, 1996). Other animal studies show that oritavancin is effective in the treatment of various S. aureus infections, including endocarditis due to MRSA (Kaatz GW, 1998). Resistant variants have not been observed after prolonged exposure to the drug. A double-blind, randomized Phase III clinical trial involving 517 patients with skin and soft tissue infections demonstrated that oritavancin can halve treatment time—from 11.5 days to approximately 5.5 days—compared with vancomycin followed by the cephalosporin cephalexin (Wasilewski MW, 2001[b]). This reduction in treatment time stems from the elimination of the oral cephalexin step-down therapy. However, because step-down therapy is inexpensive and convenient, the impact of this reduced treatment time could be negligible. Of the 517 patients, 33 had an MRSA-associated infection. Among these 33 patients, clinical success was achieved at Þrst follow-up in 74% of those given oritavancin and 80% of those given vancomycin/cephalexin. Among 256 bacteriologically evaluable patients, bacterial eradication was achieved in 74% and 76% of patients given oritavancin and vancomycin/cephalexin, respectively. In a small Phase II clinical trial involving 27 patients with gram-positive bacteremia, oritavancin demonstrated microbiological and clinical response of 85% and 78%, respectively (Wasilewski MW, 2001[a]). Bacterial eradication was conÞrmed in patients with vancomycin-resistant enterococcus and staphylococcus species. Phase I clinical trials have demonstrated that oritavancin (once-daily IV infusion over 30 minutes) has favorable pharmacokinetics and a serum half-life of approximately 10.5 days and low interindividual variability (Chien J, 1998; Thomasson HR, 1999). The long half-life reduces the dosing frequency. Safety data for oritavancin are limited, but preliminary data from clinical trials suggest that the drug is generally well tolerated. However, side effects that require additional examination include phlebitis, edema, anemia, tremor, elevated liver enzyme, and renal function abnormalities. Telavancin. Theravance (formerly Advanced Medicine) is developing its oncedaily injectable glycopeptide telavancin * for gram-positive infections. Telavancin has an extended mechanism of action; it inhibits both peptidoglycan synthesis and phospholipid synthesis in cell-wall production. The company is hoping such

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activity will reduce the level of spontaneous resistance against the drug. Telavancin is expected to be effective against MRSA. Theravance initiated Phase III trials for complicated skin and soft tissue infections in late 2004. The company also stated that it has conducted preclinical studies of telavancin using animal models of hospital-acquired pneumonia. In March 2005, the FDA granted the drug fast-track status for the treatment of hospital-acquired pneumonia and complicated skin and soft tissue infections. In January 2005, Theravance announced that it had initiated a multinational, multicenter, double-blind, active-control, Phase III trial called ATTAIN (Assessment of Telavancin for the Treatment of MRSA Pneumonia). Approximately 1,500 patients will be enrolled in the study. Primary end points will be noninferiority to vancomycin for all gram-positive infections and superiority to vancomycin in patients with MRSA. In December 2004, researchers reported the results of a Phase II clinical study of telavancin (Theravance, press release, December 8, 2004). The double-blind, multinational study compared IV telavancin dosed once a day with standard therapy (vancomycin dosed twice a day or antistaphylococcal penicillins dosed four times a day) for the treatment of complicated gram-positive skin and skin structure infections. The researchers randomized 201 patients at 18 clinical sites in the United States and South Africa to either standard therapy or 10 mg/kg telavancin. The clinical cure rates in the clinically evaluable population were 96.1% and 93.5% for patients treated with telavancin and standard therapy, respectively. Notably, in the group of microbiologically evaluable patients with MRSA infection, eradication was achieved in 92.3% of the telavancin-treated group versus 68.4% of the vancomycin-treated group. The overall incidence and severity of adverse events were similar in the telavancin-treated and standard therapy groups. In September 2004, researchers reported the results of a second Phase II study of telavancin (Stryjewski M, 2004). A total of 167 patients with suspected or conÞrmed gram-positive serious skin infections were randomized to either telavancin or standard therapy (a penicillin or vancomycin). For the 102 patients with S. aureus, telavancin cured 80% of the patients, while standard therapy cured 77%. Patients with MRSA (44 total) who received telavancin were cured 82% of the time, compared with only 69% of MRSA patients receiving standard therapy. It is not clear from available data whether standard therapy for these patients was solely vancomycin or if some patients received only standard penicillin, which would normally be ineffective against MRSA. Approximately 5% of patients withdrew from each arm because of adverse events, though fewer serious adverse events were reported for patients taking telavancin (four adverse events) than for those on standard therapy (nine adverse events). Phase I studies involving 119 healthy volunteers who received a 7.5 mg/kg or 15 mg/kg dose of telavancin as a 60-minute infusion over three days demonstrated linear pharmacokinetics, which were equal in men and women (Barriere S, 2003). This study also showed that at the higher dose, 33% of patients had nausea; 21% had headache; 15% had vomiting, dizziness, and rash; and 10% had abdominal pain/cramps. These adverse effects were classiÞed as mild. More concerning,

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at the higher dose, 10% of patients discontinued treatment because of infusionrelated “red man syndrome,” a serious allergic response. This response is one of the chief complaints about vancomycin, and it may limit the uptake of telavancin. Telavancin has the potential to be approved for complicated skin infections by 2009. However, experts state, its side-effect proÞle (e.g., red man syndrome, injection-site reactions in one-third of telavancin-treated patients) is a disadvantage. In fact, currently available information does not point to any advantage of telavancin over vancomycin or the other glycopeptide drugs that will precede it to market (i.e., dalbavancin and oritavancin). Cephalosporins Overview. The cephalosporin class is very mature and competitive; many branded and generic drugs are available. Antibiotics in this class are used to treat a variety of staphylococcal infections, and physicians are familiar with them and very comfortable prescribing them. The cephalosporins as a class are generally very safe. Although current cephalosporins are not used for MRSA because of the cross-resistance to the pathogen, new cephalosporins that are active against MRSA may beneÞt from physicians’ familiarity with the class in general. Emerging broad-spectrum cephalosporins have the potential to be very successful hospital antibiotics if they can demonstrate clinical activity, safety, and dosing similar to that of the leading cephalosporin, ceftriaxone. Cephalosporins active against MRSA would compete with the glycopeptides and the oxazolidinone linezolid. Mechanism of Action. Like all beta-lactams, cephalosporins bind to penicillinbinding proteins (PBPs) in bacteria and prevent bacterial cell-wall formation. PBPs have numerous roles within the bacterial cell, including maintenance of structural integrity, determination of cell shape, cell division, induction of capsule synthesis, and regulation of autolysis. Cephalosporins inhibit these bacterial processes, leading to bacterial cell death. BAL-5788. Basilea Pharmaceutica is developing a novel next-generation, parenteral cephalosporin called BAL-5788 (ceftobiprole, a pro-drug of BAL-9141) ∗ . It has broad-spectrum activity with potency against MRSA and penicillin-resistant S. pneumoniae and has shown a low potential to induce resistance in vitro. In February 2005, Basilea announced that it had licensed BAL-5788 to Cilag International, part of Johnson & Johnson (J&J). J&J’s Ortho-McNeil unit will market the product in the United States, while its Janssen-Cilag unit will commercialize it elsewhere. Basilea has retained the option to copromote the agent in the United States and major European countries. The FDA granted BAL-5788 two fast-track designations: the Þrst, in March 2003, is for the treatment of complicated skin and soft tissue infections due to MRSA; and the second, in June 2004, is for hospital-acquired pneumonia due to MRSA. Roche had an option to license BAL-5788 but turned it down in May

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2004, leaving Basilea with the global development, marketing, and manufacturing rights. Basilea initiated Phase III trials in November 2004. The company hopes to complete these trials by the end of 2005, a move that would enable an NDA submission in the United States in 2006. BAL-5788 has a broad-spectrum of coverage against both gram-positive and gram-negative pathogens. Like other beta-lactams, it binds to bacterial PBPs to inhibit cell-wall production, but unlike many other beta-lactams, it provides very good potency against MRSA. MRSA is resistant to most beta-lactams in two ways. The Þrst is the expression of beta-lactamases that cleave antibiotics such as methicillin and other beta-lactams. The second resistance mechanism is that MRSA acquires a new PBP from the chromosomal genetic determinant mecA. This protein, dubbed PBP2a, has a much lower binding afÞnity for methicillin and similar compounds, and these drugs are therefore very inefÞcient at blocking the protein’s activity. BAL-5788 is effective against MRSA because it is 10,000 times more resistant than penicillin G to hydrolysis by beta-lactamases, and it has a more than 100 times greater afÞnity than methicillin for PBP2a. Basilea reported positive results from a Phase II trial of BAL-5788 (Basilea, press release, March 4, 2004). Forty hospitalized patients with complicated skin infections requiring surgical intervention were enrolled. Most had deep infections caused by streptococci or staphylococci, and some patients were infected with MRSA. Of the 35 patients who were clinically evaluable, Basilea reported that all were cured. No major organ toxicity was reported, and the most frequent side effect was mild-to-moderate nausea. Further details were not disclosed. A Phase I trial involving 40 healthy male volunteers evaluated 30-minute infusions of BAL-5788 equivalent to 150, 250, 500, 750, and 1,000 mg of the active compound BAL-9141 (Schmitt-Hoffmann A, 2002). No adverse effects were reported, and analysis of the data suggested that the 750 mg dose infused twice a day would be effective against MRSA. If BAL-5788 continues to prove that it is safe and effective against MRSA, it may be used as Þrst-line broad-spectrum antibiotic therapy for hospitalized patients. BAL-5788 provides physicians with a beta-lactam option for the treatment of MRSA—an option that experts believe represents a competitive advantage because of the class’s perceived safety and efÞcacy (i.e., its bactericidal activity). However, BAL-5788’s uptake will be signiÞcantly limited by what will likely be twice-daily dosing and a higher price than that of generic cephalosporins. It will also face strong competition from the new glycopeptides—the class more commonly used for MRSA. Dihydrofolate Reductase Inhibitors Overview. Researchers believe that dihydrofolate reductase (DHFR) inhibitors will be less susceptible to resistance. Trimethoprim/sulfamethoxazole (TMP/ SMX) is currently used to treat mild MRSA infections (mainly urinary tract infections and some respiratory tract infections), but it is subject to high levels of resistance. If this resistance does not cross over to new DHFR inhibitors, then these drugs may be effective in treating MRSA while limiting resistance.

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Mechanism of Action. DHFR plays an essential role in the synthesis of DNA and RNA. By inhibiting synthesis of thymine, DHFR inhibitors target DNA and RNA synthesis, thereby causing bacterial cell death (Goodsell DS, 1999). Bacterial DHFR is distinct enough from human DHFR that drugs targeting the bacterial DHFR protein are generally not toxic to human cells. Iclaprim. Arpida has licensed all commercialization rights to the DHFR inhibitor iclaprim (AR-100) from Roche. In October 2004, Arpida announced the acquisition of Combio A/S, a Danish company with innovative chemistry technologies. The deal represents a strategic move for Arpida to rapidly expand its candidate portfolio and continue development of iclaprim. Arpida has successfully completed Phase II trials of iclaprim in the treatment of complicated skin and soft tissue infections in Europe; vancomycin was the reference compound. The intravenous formulation of the agent is advancing into Phase III trials. In October 2004, Arpida announced that it had completed a Phase I trial of oral iclaprim and that the agent will progress into later-stage trials. Because there is only one other antibiotic in the DHFR inhibitor class (TMP/SMX), Arpida hopes that resistant pathogens such as MRSA will prove sensitive to iclaprim. The company also reports that the compound has good tissue penetration, a critical asset for treating MRSA in wound infections. In January 2004, Arpida announced the results of a Phase II trial in which iclaprim was tested for efÞcacy and tolerability in patients hospitalized with infected burns, diabetic foot ulcers, cellulitis, and abscesses (Arpida, press release, January 9, 2004). Involving 92 patients, the double-blind trial compared 1 or 2 mg/kg of iclaprim with a standard dose of vancomycin. Both drugs were given intravenously twice a day for ten days. Twenty-six of the 28 patients (92.9%) taking 1 mg/kg iclaprim were clinically cured, compared with 29 of 31 patients (93.5%) taking 2 mg/kg iclaprim and 26 of 28 patients (92.9%) taking vancomycin. Microbiological success rates were similar for 1 mg/kg iclaprim and vancomycin (57.1% and 50%, respectively) and were superior for 2 mg/kg iclaprim (76.7%). Iclaprim was well tolerated, and adverse events were similar for all three groups. Preclinical studies showed that iclaprim was effective in prolonging survival in a murine model of septicemia caused by MRSA (Fischer RL, 2003). Following IV administration of iclaprim, the 50% effective dose (ED50) values were 4.3 and 5.1 mg/kg for once- and twice-daily doses, respectively. Following oral iclaprim administration, ED50 values were 17.4 and 21.7 mg/kg for once- and twice-daily doses, respectively. The corresponding ED50 values for the comparator, vancomycin, were 0.9 mg/kg after both IV and oral dosing. Iclaprim exhibited potent in vitro activity against MRSA, including many TMP/SMXresistant strains. Iclaprim will likely launch in 2009 for complicated skin and soft tissue infections. Experts suggest that the compound will likely compete for second-line status with the novel glycopeptides and other narrow-spectrum drugs. If it is launched only in an IV formulation, it will not be able to compete with linezolid

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for step-down therapy. The development of an oral formulation would be a signiÞcant advantage, especially if linezolid-resistant bacterial strains evolve by the time of iclaprim’s launch. Topoisomerase Inhibitors Overview. The topoisomerase inhibitors are a new class of antibiotics that target bacterial replication. Drugs in this class will compete with other broad-spectrum agents such as the cephalosporins. Mechanism of Action. Along with DNA gyrase (topoisomerase II), topoisomerase IV is a protein involved in bacterial DNA replication. Topoisomerase IV is responsible for decatenation and segregation of the bacterial chromosomes after replication. Inhibition of this activity stops the replication process and ultimately kills the bacteria. NXL-101. Novexel, a new pharmaceutical company formed in 2004 as the result of the spin-off of the anti-infectives unit of Aventis Pharma, is developing NXL101 * as an antibiotic with the potential to treat pathogens that are resistant to currently available drugs. NXL-101 is in preclinical development to treat gram-positive infections, including nosocomial pneumonia, skin and soft tissue infections, and VRE infections. Based on its target infections, NXL-101 will likely be used to treat hospital-acquired infections.

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Chang FY. A prospective multicenter study of Staphylococcus aureus bacteremia: incidence of endocarditis, risk factors for mortality, and clinical impact of methicillin resistance. Medicine. 2003;82:322–332. Chang S, et al. Infection with vancomycin-resistant Staphylococcus aureus containing the vanA resistance gene. New England Journal of Medicine. 2003;348:1342–1347. Charlebois ED, et al. Origins of community strains of methicillin-resistant Staphylococcus aureus. Clinical Infectious Diseases. 2004;39:47–54. Chastre J, Fagon J. Ventilator-associated pneumonia. American Journal of Respiratory and Critical Care Medicine. 2002;165:867–903. Chien J, et al. Safety and pharmacokinetics of single intravenous doses of LY333328 diphosphate (glycopeptide) in healthy men. Abstracts of the 38th Interscience Conference on Antimicrobial Agents and Chemotherapy. September 1998; San Diego, CA. Abstract A-55. Chotani RA, et al. Nosocomial infections. In: Nelson KE, et al. eds. Infectious Disease Epidemiology. Gaithersburg, MD: Aspen Publishers Inc; 2001. Clarebout G, et al. Unusual inducible cross-resistance to macrolides, lincosamides, and streptogramins B by methylase production in clinical isolates of Staphylococcus aureus. Microbial Drug Resistance. 2001;7:317–322. Coello R, et al. Surveillance of hospital-acquired infection in England, Germany, and the Netherlands: will international comparison of rates be possible? Infection Control and Hospital Epidemiology. 2001;22:393–397. Cooper BS, et al. Isolation measures in the hospital management of methicillin resistant Staphylococcus aureus (MRSA): systematic review of the literature. British Medical Journal. 2004;329:533. Cosgrove SE, et al. Comparison of mortality associated with methicillin-resistant and methicillin-susceptible Staphylococcus aureus bacteremia: a meta-analysis. Clinical Infectious Diseases. 2003;36:53–59. Critchley IA, et al. REP8839: a novel methionyl tRNA synthetase inhibitor with potent activity against Staphylococcus aureus and Streptococcus pyogenes. 44th Interscience Conference on Antimicrobial Agents and Chemotherapy. October 2004. Washington, D.C. Abstract F-728. Crnich CJ, Maki DG. The role of intravascular devices in sepsis. Current Infectious Disease Reports. 2001;3:496–506. Cross JT Jr., Campbell GD Jr. Therapy of nosocomial pneumonia. Medical Clinics of North America. 2001;85(6):1583–1594. Crowe M, et al. Bacteremia in the adult intensive care unit of a teaching hospital in Nottingham, UK, 1985–1996. European Journal of Clinical Microbiology and Infectious Diseases. 1998;17:377–384. Cruciani M, et al. Penetration of vancomycin into human lung tissue. Journal of Antimicrobial Chemotherapy. 1996;38(5):865–869. Cucarella C, et al. Bap, a Staphylococcus aureus surface protein involved in bioÞlm formation. Journal of Bacteriology. 2001;183:2888–2896. Dartois N. Results of a Phase III, double-blind, safety and efÞcacy study comparing tigecycline with vancomycin/aztreonam to treat complicated skin and skin structure infections. 44th Interscience Conference on Antimicrobial Agents and Chemotherapy. October 2004. Washington. Abstract L-986.

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White RJ, et al. V-Glycopeptide: Phase I single- and multiple-dose placebo-controlled intravenous safety, pharmacokinetic, and pharmacodynamic study in healthy subjects. Abstracts of the 40th Interscience Conference on Antimicrobial Agents and Chemotherapy. September 2000; Toronto, Canada. Abstract 2196. Wilcox MH. EfÞcacy of linezolid versus comparator therapies in Gram-positive infections. Journal of Antimicrobial Chemotherapy. 2003;51(suppl 2):ii27-ii35. Wilton P, et al. Strategies to contain the emergence of antimicrobial resistance: a systematic review of effectiveness and cost-effectiveness. Journal of Health Service Research Policy. 2002;7:111–117. Witte W. Glycopeptide-resistant Staphylococcus.Journal of Veterinary Medicine Series B. Infectious Diseases and Veterinary Public Health. 2004;51:370–373. Wood MJ. Comparative safety of teicoplanin and vancomycin. Journal of Chemotherapy. 2000;12(suppl 5):21–25. World Health Organization (WHO), Department of Communicable Disease, Surveillance and Response. Prevention of hospital-acquired infections. A practical guide, 2nd ed. 2002. www.who.int/drugresistance/en/. Accessed February 16, 2005. Wu JA, et al. Lysostaphin disrupts Staphylococcus aureus and Staphylococcus epidermidis bioÞlms on artiÞcial surfaces. Antimicrobial Agents and Chemotherapy. 2003;47:11:3407–3414. Wunderink RG, et al. Linezolid versus vancomycin: analysis of two double-blind studies of patients with methicillin-resistant Staphylococcus aureus nosocomial pneumonia. Chest. 2003;124(5):1789–1797. Yano M, et al. Preoperative intranasal mupirocin ointment signiÞcantly reduces postoperative infection with Staphylococcus aureus in patients undergoing upper gastrointestinal surgery. Surgery Today. 2000;30:16–21. Zafar AB, et al. Use of 0.3% triclosan (Bacti-Stat) to eradicate an outbreak of methicillinresistant Staphylococcus aureus in a neonatal nursery. American Journal of Infection Control. 1995;23:200–208.

Data and Information Integration in Pharmaceutical R&D

SUMMARY EfÞcient use of information will ultimately accelerate drug development and reduce its cost. To this end, pharmaceutical companies have invested heavily in high-throughput data-generation technologies that are producing enormous quantities of data and a rapidly growing fund of information—but so far, that investment has not yielded an increase in R&D productivity: the annual rate of introductions of new chemical entities remains constant. A probable reason is that researchers cannot efÞciently and effectively exploit the data and information they are amassing. What they need is better computational tools for accessing and integrating data and for identifying and integrating information. Innovative software companies are responding to that need, developing tools that will accelerate data generation, integration, and analysis and facilitate identiÞcation and retrieval of information from “unstructured” sources—text and images. In this article, we present a detailed discussion of current technologies and then examine some new and emerging technologies that will help pharmaceutical and biotechnology companies to realize maximum beneÞt from their R&D investments. BUSINESS IMPLICATIONS •

ToreducethecostandincreasetheefÞciencyofdataacquisition,pharmaceutical and biotechnology companies are using genomic, proteomic, and other highthroughput technologies for drug discovery. The widespread implementation of these industrialized technologies has yielded an explosion of data and contributed to a precipitous rise in R&D spending; however, corporate investment in data production has not yet increased pharmaceutical R&D productivity.

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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•

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To reap the beneÞts of R&D investments, drug companies need better computational tools for Þnding relevant data from heterogeneous information sources. Several companies and organizations—including IBM, geneticXchange, and The National Center for Genome Resources—have developed software that automates data access and integration from different sources. The current trend is toward use of federated database systems that dramatically accelerate access to new data. More effective text-based information retrieval and extraction technologies will provide an important catalyst for knowledge generation. Several laboratories and companies—including Inxight Software, SPSS, TheBrain Technologies, and Groxis—are developing new computational tools to enable automated access to relevant information contained in documents. Development and widespread adoption of standards for terminology and concepts that underlie database organization will be critical to efÞcient access and retrieval of needed information. Standardization will also help to increase R&D productivity by driving down the cost of information identiÞcation and access.

INTRODUCTION Developing a new pharmaceutical product requires the assembly of many separate and distinct processes, including drug target identiÞcation and validation, lead compound identiÞcation and optimization, drug manufacturing, preclinical testing, and human clinical trials. Most of these processes involve extensive research and experimentation and generate and use vast amounts of data. To reduce the cost and increase the efÞciency of data acquisition, pharmaceutical researchers are applying genomic, proteomic, and other high-throughput technologies to the selection of disease targets and therapeutic candidates. The widespread implementation of these industrialized technologies has yielded an explosion of data and an unwieldy and rapidly growing pool of information. Corporate investment in high-throughput, data-intensive technologies has contributed to a precipitous rise in R&D spending; however, the annual rate of introduction of new chemical entities (NCEs) has remained approximately constant. This analysis suggests that investment in more efÞcient data production has not yet increased pharmaceutical R&D productivity, and—as measured by NCE introduction—the current trend is arguably one of decreasing productivity. The efÞcient use of information will ultimately accelerate and reduce the cost of drug development. According to one estimate, the application of computational technologies to increase access to and use information across the R&D process could save up to $200 million and two to three years in the development of a new drug (IBM Business Consulting Services, 1999). Computational technologies excel at analyzing vast pools of data; however, these tools will increase productivity only if they help researchers to derive and synthesize new knowledge that can be used to improve experimental productivity and research program decision making. The current decrease in R&D productivity noted here probably

EXPLOITING THE KNOWLEDGE VALUE CHAIN

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reßects the inability of organizations to efÞciently access, analyze, and use the enormous quantities of data and information that are rapidly becoming available. To reap the potential beneÞts of their investments in new data-generating technologies, pharmaceutical and biotechnology companies need better computational tools for Þnding relevant data and more efÞcient methods for integrating information. In this article, we discuss how data and information integration technologies can improve the knowledge value chain in pharmaceutical R&D. EXPLOITING THE KNOWLEDGE VALUE CHAIN Drug discovery and development is rapidly becoming an information business whose success depends on the ability of researchers to derive new knowledge from data and information efÞciently (see the sidebar entitled “Data Terminology”). Figure 1 depicts the relationship between data, information, and knowledge. In this relationship, data, information, and knowledge form a “knowledge value chain” of assets of increasing value. At the beginning of this chain, researchers analyze data (such as experimental results) to identify relationships that help translate the data into information. Then, in the information derived from various types of data, researchers attempt to recognize patterns that will yield new knowledge. For example, relationships between data points from a

FIGURE 1. The knowledge value chain.

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high-throughput drug screen translate into information about the effects of a particular test compound on a drug target. This information—together with, perhaps, information from a gene expression proÞling experiment and a genetic knockout test—may yield new knowledge about a compound’s physiological effects. DATA TERMINOLOGY Data: The plural of the word datum. Database: A compilation of data and metadata. Database Management System (DBMS): A series of programs that access and manipulate information in a database. Database Schema: The organization of the entire collection of data in the database, including the data type, data structure, and metadata. Data Cleaning: The elimination or correction (by scientists, database administrators, and programmers) of data errors, including missing data, data entry errors, incorrect classiÞcation of data, and measurement errors. Data Structure: The way in which the data are organized in a database. Typical data structures include records, lists, arrays, and tables. Data Type: A class of data (e.g., percentage, time, concentration). Datum: A single piece of information, such as a measurement or value derived from an analysis, experiment, or survey. Metadata: Information about data. Researchers use new knowledge to make decisions, such as whether to move a compound forward in the development process. To ensure that these decisions are correct, scientists are seeking ways to generate more complete data and information about biological processes and potential drug compounds. Therefore, pharmaceutical and biotechnology companies have invested heavily in data-intensive technologies in an effort to increase the success rate of drug development programs—but companies have yet to reap the beneÞts from these investments. The inability to adequately and efÞciently access data, to integrate heterogeneous information, and to identify relevant pieces contributes to this failure. Although high-throughput technologies have dramatically increased the number and size of data “pools,” the “pipes” (primarily computer networks and software programs) required to access and interpret these data have not improved at the same rate. Clearly, better tools for accessing and integrating data and information are needed. In the following sections, we will discuss computational technologies that will help researchers to automate data access and integration. We will also explore strategies and tools that help researchers to identify and integrate new information.

A TYPICAL COMPUTER SYSTEM ARCHITECTURE

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UNIVERSAL DATA ACCESS AND INTEGRATION Data useful to drug developers—such as gene sequences, protein expression proÞles, functional information, and chemical information—are dispersed throughout hundreds of computer systems. Different organizations, using different types of software and hardware, have generated numerous distinct databases that contain many different types of data. For example, more than 500 databases of biological information are available in the public sector alone, many of them incompatible with other databases. Incompatible systems exist even within single companies, a result of the disconnected activities of different research groups and of corporate mergers and acquisitions. This dispersion of data creates a signiÞcant problem for drug developers. During the research process, scientists would like to be able to make a single request of, or query, several databases at once. For example, a researcher may want to look for a particular gene sequence but must query several databases to ensure a comprehensive search of all known gene sequences. One approach is to search different databases individually and then manually combine the results into a single set. However, as the research community’s appetite for data has grown, the manual compilation of individual queries from multiple systems into one database has become an unacceptably inefÞcient method of data integration. In response, researchers have developed various methods of automating this process. The two main approaches to automating multiple searches are data warehouses and federated databases. In data warehousing, data from different databases are combined into a single master database that can be searched efÞciently; in federated databases, special software programs enable users to search multiple databases separately and obtain a single set of results. We discuss these approaches in more detail in the following sections. For a basic description of the architecture underlying these systems, see the sidebar entitled “A Typical Computer System Architecture.” A TYPICAL COMPUTER SYSTEM ARCHITECTURE Typical computer system architectures consist of three layers (see Figure 2): client machines (the user interface), servers (computers that allocate applications and system resources), and a data storage layer (the database itself). Two types of software interact with data in this system: the database management systems (DBMS$) that identify and retrieve data, and the data analysis applications that yield information. The objective of computational tools for data integration is to automate access to as much data as possible through a single user interface. Data Warehouses An early strategy for automating searches and integrating data was to consolidate data from multiple sources into one master database known as a data warehouse (see Figure 3). Data-intensive industries such as the pharmaceutical, Þnancial,

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FIGURE 2. A typical computer system architecture.

and retail services industries have all employed this strategy. In data warehousing, data from different databases are restructured to a common format and then physically moved into a single database. This approach circumvents the problem of database incompatibilities. Data warehouse systems perform well: their implementation is straightforward for users and the quality of the data is typically high due to validation during data transfer. The Lion Bioscience (Heidelberg, Germany) SRS database system is an example of a data warehouse solution for the life sciences industry. A signiÞcant disadvantage of data warehouses is that they typically require considerable investments of time and effort by database administrators and

A TYPICAL COMPUTER SYSTEM ARCHITECTURE

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FIGURE 3. Comparison of the data warehouse and federated database strategies.

programmers—especially in the pharmaceutical industry. The pharmaceutical research environment generates and uses an ever-changing assortment of data types that are generated by new technologies for studying biological systems. Consequently, importing and restructuring data to build and maintain a data warehouse requires ongoing programming and database maintenance. In addition, research in the competitive pharmaceutical environment requires rapid access to new data in order to maximize the data’s value; the lag time to restructure and validate data diminishes their value. These disadvantages limit the utility of data warehouses for drug discovery and development research. Although data warehouses do not appear to be the general solution for pharmaceutical data integration, they are appropriate for certain applications. For example, if a company wishes to conceal its pattern of database searches from competitors to avoid revealing information on proprietary projects, it may prefer a data warehousing strategy: by downloading batch data from external genomic databases into an internal data warehouse, the company can perform detailed analysis entirely on its own secure internal computer system. Federated Database Systems The current high rate of data generation and the rapid evolution of data-generating technologies place an extraordinary demand on data integration technology. To keep pace with the ßood of new data, access automation and data integration at the

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source have become necessities. By linking source databases into “federations,” drug developers can automate and dramatically accelerate access to new data. Database federations use software programs to automate searches of the separate databases that form the federation (see Figure 3). These databases may be geographically distributed and operate independently. In a federated database, a software layer (called middleware) between the users and the databases links the databases to a single interface, thus causing multiple databases to appear to users as a single data source. These systems leave source data in their original locations and structures. Middleware performs several functions required for one-step access to multiple databases. The databases in a federation may use different types of DBMSs, each of them speaking a different language. To access a database, the user interface must know how to translate its request into the appropriate language and format for that database. The software uses a component called a “wrapper” to access and query the data source. Once requests are translated, the middleware must reformat, or optimize, them to use the least amount of computer time on each database. Individual requests are then sent to their respective databases, where they are executed. The data are returned by the federated databases to the middleware on the originating computer, where the data may be used immediately or placed in a local database for subsequent analysis. The degree to which a common data structure and organization is used can be different in each federated database, which impacts both ease of implementation and system performance. If each database is organized in the same way and uses the same DBMS, less programming is required to integrate data sources and the system’s performance may be as high as that of a data warehouse. However, federations usually include databases with different schema and DBMSs; more elaborate software is required to connect the systems and, consequently, performance may deteriorate. Advantages of the federation strategy include lower cost and shorter development time due to the reduced need for custom software. In addition, a federation can include a wide range of databases; this feature facilitates expansion or reconÞguration of the federation as new databases are built or data requirements change. A signiÞcant disadvantage of the federated approach is that no effort is made to clean the data; hence, individual data sets may contain errors and there may be inconsistencies between database structures such as data classiÞcation terminologies (see the sidebar entitled “Data Terminology”). Another disadvantage of this strategy is that it depends heavily on different network systems and is therefore likely to encounter numerous communications and software incompatibilities. Although federation middleware products are not currently designed to communicate with front-end applications—such as DNA sequence analysis programs—that perform actual data analysis, these capabilities may eventually be available. Recent developments in programming techniques that use modulebased software approaches, such as object-oriented programming and message passing, presage the creation of universal access software that will communicate

A TYPICAL COMPUTER SYSTEM ARCHITECTURE

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TABLE 1. Profiled Companies and Organizations Organization Cellomics geneticXchange, Inc. Groxis IBM

Inxight Software

Technologies CellSpace Knowledge Miner for information retrieval and extraction DiscoveryHub federated database integration middleware Grokker for information retrieval, extraction, and knowledge map construction DiscoveryLink federated database integration middleware; Intelligent Miner for Text tools for searching, categorizing, and extracting unstructured information SmartDiscovery enterprise system for integrated search, categorization, summarization, and entity extraction; Star Tree visual organization tools; tools for natural language processing, file navigation, documents classification, and data visualization Electronic laboratory notebook

Kodak Research and Development Lion Bioscience SRS integrated data warehouse platform; DiscoveryCenter enterprise data integration and analysis environment National Center for ISYS federated database integration, data Genome exchange service software; data Research exchanges services Pacific Northwest Electronic laboratory notebook National Laboratories Reel Two Gene Ontology Knowledge Discovery System database; document classification and information extraction tools Scimagix Scientific Image Management System for analysis, storage, and retrieval of image data SPSS Lexiquest suite of tools for analysis of unstructured data TheBrain BrainEKP (Enterprise Knowledge Platform) Technologies for identification and organization of Corporation unstructured information

Web Site www.cellomics.com www.geneticxchange.com www.groxis.com www.ibm.com

www.inxight.com

www.kodak.com www.lionbioscience.com

www.ncgr.com

www.pnl.gov

www.reeltwo.com

www.scimagix.com

www.spss.com www.thebrain.com

with front-end applications and allow users to focus on analytical tasks while data integration is performed automatically. In the following sections, we describe some of the leading products and development efforts in the area of federated databases. See Table 1 for a list of companies proÞled in this article. DiscoveryLink. DiscoveryLink, IBM’s universal data access middleware package, uses the federated database strategy. DiscoveryLink includes query translators

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and database interfaces (wrappers) for many commonly used DBMSs. DiscoveryLink is built on IBM’s DB2 database software. Users (or applications) generate Structured Query Language (SQL) queries through the DB2 DBMS. The system passes these queries to the federation query optimizer, which calculates a set of queries based on information sent by the system’s wrapper modules. The federation server then distributes the query to the appropriate DBMS for each database speciÞed by the user. Through this process, the user can obtain and view results in a single format—as though a single data source, rather than several heterogeneous sources, had been used—and the original data is maintained in its original form and location. DiscoveryLink integrates with other components of IBM’s Life Sciences Framework product line, such as Lotus K-Station and IBM’s text analysis tools. Integrating DiscoveryLink with other R&D applications requires additional application development. discoveryHub. The discoveryhub middleware from geneticXchange (Menlo Park, California) also uses the federated database strategy. DiscoveryHub allows the user to see data from multiple databases (and other data sources) at the same time. Like DiscoveryLink, discoveryHub does not transform data into the same data structure, even though they appear to be from a single source. One version of discoveryHub works with Oracle’s 9i DBMS and allows the user to store a local copy of the data retrieved from external sources. GeneticXchange states that a strength of discoveryHub is its ability to retrieve data of complex structure and transform it to a less complex form without losing information contained in the original data structure. ISYS. ISYS (Integrated SYStem), developed at the National Center for Genome Resources (Santa Fe, New Mexico), uses module-based software programming to enable “plug and play” integration of databases and analytical programs. Like DiscoveryLink and discoveryHub, ISYS performs query translation through wrappers and data reformatting without transforming the original data. In addition, ISYS binds together software applications as well as data sources. The system functions as a data trafÞc bus: it links databases and analysis programs and passes retrieved data objects between them. An attractive feature of the ISYS bus approach is the high degree of integration of analysis applications into the overall scheme. ISYS creates a common language that allows analytical applications to communicate directly with the available databases. This system facilitates the user’s data exploration by updating all applications with the results of the analyses being performed: for example, a search for DNA sequences that are similar to one the user is interested in will produce a list of sequences; ISYS can then display other information about the genes in that list, such as protein sequence or gene expression data, in other ISYS-compatible applications that are also running; further analysis of the gene expression data, for example, may create a new list of potentially interesting genes. Finally, all the applications that are running are updated automatically with this new list or other information from the analysis.

INFORMATION IDENTIFICATION AND INTEGRATION

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A disadvantage of the current version of ISYS is that the package must be installed and run locally on each user’s computer. This requirement sacriÞces some scalability and computational power. Collaborative Approaches. IBM and Lion Bioscience are also integrating applications with database systems. These companies are collaborating to use IBM’s DiscoveryLink in Lion’s DiscoveryCenter enterprise R&D software system. Lion’s DiscoveryCenter is a suite of analytical programs designed to access information from a single database. Integration of IBM’s DiscoveryLink with DiscoveryCenter will enable these applications to access data from multiple databases by using the federated database approach described above. In October 2002, the companies announced an agreement in which IBM will sell and comarket Lion Hosted Services, a subscription-based hosted computing service based on DiscoveryLink and DiscoveryCenter technologies, for drug discovery researchers. Clients will be able to use these services to obtain and integrate data from a vast array of online biological databases. Other recent collaborations also suggest a trend toward automatic integration of applications with data sources through federation middleware. In January 2002, IBM announced that it has established a collaboration with Accelerys (San Diego, California; a wholly owned subsidiary of Pharmacopeia) to incorporate IBM’s server technology into the Accelerys Discovery Studio platform, which is a suite of life sciences software applications. In one of several programs, the development alliance will also combine IBM’s DiscoveryLink product with the Accelerys Discovery Studio technologies. In July 2002, geneticXchange and Applied Biosystems (Menlo Park, California) struck a global reseller agreement in which Applied Biosystems will sell discoveryHub as part of its Rapid Integration Solutions program. DiscoveryHub will be installed in laboratory information management software developed by Applied Biosystems to facilitate data exchange for high-throughput screening systems. INFORMATION IDENTIFICATION AND INTEGRATION After researchers have accessed and analyzed experimental data, they usually store the resulting information in text-based formats such as written reports and research articles. Although these formats are not easily computable and are therefore referred to as “unstructured,” they are critical to new knowledge generation, a process that mainly results from human mental integration of many types of information (see Figure 4). Text-based information is particularly important in decision making because these documents often provide important contextual information. They record the thinking of previous researchers and provide information on conditions and nuances that cannot easily be attached to traditional data sets. Because of the recent exponential increase in the number of information sources, identiÞcation of the most relevant text-based documents has become

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FIGURE 4. Creating knowledge from data and information.

an increasingly difÞcult task for scientists. Also, certain types of data that may be particularly useful in knowledge generation—such as physicians’ notes in clinical studies—are stored in unstructured formats that require time-consuming analysis. Nevertheless, unstructured information must be identiÞed and organized before relevant information can be extracted and integrated. Researchers need better computational tools to streamline and accelerate these tasks. For many years, various organizations have manually compiled computersearchable compendia of biological knowledge—Ingenuity Systems (Alviso, California), for example, has compiled an extensive knowledge base of scientiÞc Þndings. Development of more effective text-based information retrieval and extraction technologies will provide an important catalyst for knowledge generation. Several laboratories and companies are developing new computational tools to enable automated access to relevant information in near-real time. Results obtained using these tools can be applied to development of organization-speciÞc knowledge bases. In the following sections, we describe some of the tasks required to identify and integrate information as well as various new and emerging tools and technologies designed to accomplish these tasks.

INFORMATION IDENTIFICATION AND INTEGRATION

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Identification of Relevant Documents The majority of text-based documents are in a computer-readable format. The challenge is to identify, within a very large repository, the few documents or other information sources that are relevant to the current task. Three technologies for identifying relevant subsets of documents are important: information retrieval, information extraction, and document categorization. Information Retrieval. Information retrieval technologies use one or more criteria to identify relevant documents. These programs then either retrieve copies of the documents or create a list of documents. The simplest form of information retrieval is the keyword search. Familiar examples of systems that use this approach include the National Center for Biotechnology Information’s PubMed service for searching the Medline database of professional journals, and Internet portal search engines that identify Web pages. Keyword searches may identify large numbers of documents that contain the keyword. However, these technologies do not take advantage of syntactic or grammatical information in the text to ensure the relevance of documents retrieved. ReÞnements to text searching methods help users to Þnd more uniformly relevant documents. If the keywords selected by the user are not the “best” terms to deÞne the subject matter, the search may not return the most useful documents. Query reÞnement is a technique that helps the user to select search terms; an example is the use of the “related articles” link in the PubMed search engine to request a search for “more documents like this one.” Executing this request prompts the search engine to automatically formulate a new query based on terms from the selected document. Users can also specify secondary search elements, such as taxonomies or document types, to further reÞne the output. Information Extraction. Information extraction technologies attempt to distill the deeper, Þne-grained information in a document. Using semantic and grammatical information at the level of sentence and paragraphs, these tools analyze relationships between terms in the document. However, natural language is rife with subtle variations in meaning that depend on the surrounding words, and computational technologies lack the understanding of context that human beings possess. A simple example of the challenges facing information extraction tools is the experience most people have had in using spell checkers in word-processing programs: the phrase, “they cannot determine weather the word is the one wanted,” graphically demonstrates both the impact of context on word choice and an error that will be missed by spell- and grammar-checking software. Natural language processing and other information extraction technologies are active areas of research aimed at developing methods for accurately analyzing text and identifying concepts within documents. Document Categorization. Document categorization software organizes a collection of documents according to user-deÞned taxonomies and ontologies. Taxonomies are knowledge structures that describe the relationships between

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terms and concepts to provide a framework for data and information organization; ontologies are taxonomies that are structured to facilitate computational processing. Examples of ontologies include the Medical Subject Headings (MeSH) used in Medline and the Gene Ontology used to annotate genome databases. Document categorization software packages are built around algorithms that analyze the linguistic content of a document, extract some level of meaning, then assign the document to one of the categories in the taxonomy. The key difference between the available categorization packages is the algorithm used for document analysis. Subtle shades of meaning in natural language that are difÞcult to analyze computationally may pose a signiÞcant challenge for document categorization engines, as they do for information extraction technologies. Changes in the underlying knowledge structure (common in a research environment) further compound the challenge of accurate document classiÞcation. Tools for Identifying and Organizing Unstructured Information Several companies offer products that incorporate information retrieval, information extraction, and categorization functions. These products help researchers to identify and organize documents that contain relevant information. SmartDiscovery. Inxight Software (Sunnyvale, California) offers the SmartDiscovery enterprise system for integrated search, categorization, summarization, and entity extraction (i.e., identiÞcation and extraction of key entities, such as names of people or places) of large bodies of documents. The software can perform these tasks for documents written in any of 23 languages. The SmartDiscovery search engine uses metadata (such as user-speciÞed taxonomies) to improve the accuracy and comprehensiveness of search results. Researchers can reÞne these results further by using the SmartDiscovery summarizer module, a program that extracts information to create abstracts. The summarizer module can also facilitate content-based document analysis by identifying and transferring terms, names, and relationships from documents to secondary databases. Alternatively, the SmartDiscovery categorizer module can be used to create a subset of documents. This module takes a large collection of documents, analyzes them using Inxight’s proprietary natural language processing algorithms and statistical methods, and groups them according to type and content. Intelligent Miner for Text. IBM provides the Intelligent Miner for Text suite of tools for searching, categorizing, and extracting knowledge from unstructured information. The software suite combines search components, a summarizer, a categorization engine, and a document-clustering tool. This product line competes with Inxight’s SmartDiscovery system. The Intelligent Miner for Text suite also provides a Web-based enterprise system for integrated analysis of unstructured data. Like SmartDiscovery, the Intelligent Miner for Text is intended to enable analysis of unstructured content across a wide range of diverse organizational activities.

INFORMATION IDENTIFICATION AND INTEGRATION

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CellSpace Knowledge Miner. Cellomics (Pittsburgh, Pennsylvania) offers the CellSpace Knowledge Miner service for molecular biology literature analysis. CellSpace combines a curated literature database (compiled by Cellomics) with statistical text mining capabilities and a graphical interface to explore relationships between molecules and biological processes. Using keyword searches and a drag-and-drop interface, the user can Þnd molecules or processes that are related to a starting molecule or process, Þnd associations of processes among a group of molecules or genes (such as a gene expression cluster), or Þnd molecules associated with a process. The strength of the statistical association, based on literature analysis, is given for each relationship identiÞed. Access to the underlying literature is given either through links in the graphic display of the relationship or through a conventional ranked list of results. Lexiquest. SPSS (Chicago, Illinois) offers the Lexiquest integrated suite of tools for analysis of unstructured data. The Lexiquest Categorize, Lexiquest Mine, and Lexiquest Guide modules perform document categorization, information extraction, and information retrieval functions, respectively. Gene Ontology Knowledge Discovery System. Reel Two (San Francisco, California) has developed document classiÞcation and information extraction tools and information integration software. In collaboration with GeneEd (San Francisco, California), Reel Two is developing the Gene Ontology Knowledge Discovery System (GO KDS). GO KDS is a searchable database of 12 million Medline abstracts that have been classiÞed using Reel Two’s classiÞcation engine. Users search the database by keyword or by Gene Ontology terms; the software returns document lists ranked by relevance to Gene Ontology concepts. Based on that result set, users can initiate new searches, using the listed concepts as links to explore additional abstracts related to the Gene Ontology concepts. Visual Tools for Organizing and Integrating Information Visual tools for integrating information (as well as data) serve two useful purposes on the path to knowledge generation. First, graphic presentations can improve the researcher’s ability to identify information. Graphic displays of large bodies of documents or data can enable the user to quickly identify items that share properties and are likely to be related. Furthermore, visualization tools can enable an entire organization to view information, a feature that facilitates collaboration between groups and individuals. Graphic information displays can be automatically updated so that all users can see and access new information. Second, graphic displays take advantage of the user’s spatial cognition (or spatial memory) to ease the task of integrating information within a large collection. In other words, providing visual associations among documents makes it easier for a user to scan a large amount of information and helps to highlight concepts. Furthermore, conceptual patterns that are clearly visible in graphic presentations may not be readily apparent in text-based annotations. Links that connect distinct groups of documents help researchers to see connections between concepts, Þnd

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patterns of concepts within a body of information, and subsequently develop a new understanding of the available information. Document clustering technology is a tool widely used for grouping documents and presenting them graphically. This technology uses statistical analyses of term frequencies across a document set to group the documents by similarity in term content. (This approach is conceptually similar to clustering algorithms used to analyze DNA microarray data.) The clustering analysis can be displayed in a graphic format, allowing the user to quickly identify a group of documents with related content. In addition, the terms that form the kernel of a cluster can form a “label” for the cluster that provides the user with a “summary” of the type of information in each cluster. Several companies offer visual tools for organizing and integrating information. We describe some of these tools in the following sections. Intelligent Miner for Text. IBM offers a document-clustering tool as a component of its Intelligent Miner for Text product line (see “Tools for Identifying and Organizing Unstructured Information”). The IBM tool performs hierarchical clustering, in which related clusters fall onto the same branch of a tree that is presented graphically. The tool also performs binary relational clustering, in which documents are grouped in a ßat structure by topic and links are established between groups with related topics. No classiÞcation schemes or taxonomies are required. Star Tree. Inxight Software has developed the Star Tree technology, which organizes unstructured content such as documents, Web pages, or data objects in a dynamic hierarchical structure. Star Tree offers the functionality of a traditional tree Þle structure, with “active reorganization” of the content as the user navigates through the information collection. The underlying organization can be derived either manually or computationally, using programs such as Inxight’s categorizer or other information organization software. To use a Star Tree, the user selects a node of information on the computer screen. When the node is dragged around the screen, all the surrounding connections are reorganized around the node. As new nodes are selected and moved, the visible area of the information collection changes. Inxight also offers the Table Lens product that imposes a graphical organization on tabular data. Table Lens can visualize 100 columns and 65,000 rows of data and dynamically rearrange them based on user selections. This capability enables the user to explore the data set visually for patterns and trends. Brain Enterprise Knowledge Platform. TheBrain Technologies has developed the Brain Enterprise Knowledge Platform (BrainEKP) to identify and organize unstructured information. The user interface is a graphic display of individual elements, called “thoughts,” that represent documents or other information elements. The thoughts have a hierarchical organization but can also be linked to any other thought node in the system. Selecting a thought rearranges the display

INFORMATION IDENTIFICATION AND INTEGRATION

19

to put the active thought at the center, surrounded by linked thoughts. Information contained in the selected thought is displayed in a second frame of the screen. The BrainEKP system incorporates modules that enable universal data access and facilitate internal collaboration. Documents or data sources from a wide variety of applications can be accessed within the BrainEKP environment. New information elements that are entered into the system are immediately available to all users. The taxonomy underlying the organization of data and information in the BrainEKP is a knowledge model tailored to present information in a structure that mirrors the organization’s business processes. Key activities, such as initiation of the patent application process, can be embedded within the knowledge model: for example, when a user reviews information relating to a new drug target, the system can trigger the patent application process and create associated documents, such as messages to attorneys and inventors. Grokker. Grokker, developed by Groxis (Sausalito, California), is a program that provides automated search and organization features and sophisticated visualization capabilities. The program uses search results to construct graphic “knowledge maps” that help users to identify connections between pieces of information—such as documents, raw data, or e-mail—that might otherwise be overlooked. The company plans to release an enterprise version of Grokker in addition to the desktop prerelease version now available. Image Data Mining Tools Images are an important category of unstructured data that researchers have found difÞcult to mine. Although digitizing image data to facilitate computational access and data mining is now common, the task of integrating this type of information remains difÞcult. Currently, no simple methods are available for instructing a computer to logically search an image. Major barriers to the computational integration of image information include the lack of standards for most scientiÞc and medical images and the wide range of content that these images contain. To circumvent these barriers, researchers can use text-based annotation systems to classify and retrieve image data. These annotations deÞne the properties, subject matter, and content of images. The image data are combined with the corresponding annotations and stored as image objects that can be retrieved with object database technology, available from vendors such as IBM and Oracle. Scientific Image Management System. Scimagix (San Mateo, California) has developed the ScientiÞc Image Management System for analysis, storage, and retrieval of image data. The software incorporates image analysis functions to help highlight important elements of certain types of images—for example, the location and intensity of spots on two-dimensional protein gels. In addition, the software bundles annotation with images, assists in providing links between related images, and provides an organizational structure to stored image objects.

20

DATA AND INFORMATION INTEGRATION IN PHARMACEUTICAL R&D

Electronic Research Documentation Tools A research laboratory notebook is an example of a document that is traditionally recorded, usually by hand, in a written format. This practice continues in most laboratories, mostly because electronic records are not acceptable to the legal system (e.g., to prove inventorship on patents), and presents a formidable barrier to implementation of computer-based laboratory record keeping. Although electronic laboratory notebook technology exists, researchers have been slow to adopt it. Developments in security technologies, such as digital signatures and digital time stamps, may eventually offer solutions that are accepted by the legal system and enable use of an entirely computerized laboratory notebook system. Electronic systems will dramatically improve efÞciency in locating relevant information that resides in laboratory research documents. In this section we describe a few examples of electronic laboratory notebook development projects. Pacific Northwest National Laboratories Project. Through the Electronic Laboratory Notebook Project, researchers at PaciÞc Northwest National Laboratories (PNNL; Richland, Washington) are collaborating with other national laboratories to develop a dedicated electronic laboratory notebook. PNNL’s opensource Electronic Laboratory Notebook (ELN), which is based on Java and other Web technologies, allows users to create notebook pages and attach to them a combination of text, images, drawings, equations, and other Þles. When the pages are submitted, notes appear in the notebook’s “table of contents” Þle tree and can be viewed by other authorized researchers using a Web browser. Pages can be digitally signed, time-stamped, and witnessed (i.e., read and signed by a second researcher). Kodak Project. General text-based information input, tracking, and sharing applications—such as Lotus Notes—provide alternatives to stand-alone software packages (such as PNNL’s package) that are designed to function exclusively as laboratory notebooks. For example, Kodak Research and Development (Harrow, United Kingdom) has developed a laboratory notebook system based on Lotus Notes software that appears to work well for research information input and sharing across the organization. As in the PNNL approach, researchers enter textual information directly into a workstation and store it in a database system. This approach enables different groups of researchers, including those at different geographical locations, to readily share data and retrieve experimental notes. Other Projects. Several other electronic laboratory notebook programs are available or in development. The Collaborative Electronic Notebook Systems Association (CENSA; Woburn, Massachusetts) is a group of public and private sector organizations dedicated to developing standards, procedures, and technologies that will enable electronic recordkeeping systems. Other notekeeping and document content software is available from Hallogram Publishing (Aurora, Colorado), Documentum (Pleasanton, California), Doxis (Norwood, Massachusetts), and Adobe (San Jose, California).

OUTLOOK

21

OUTLOOK The quest to discover new, safer, and more effective therapies will rely increasingly on the ability of drug developers to understand biological systems and disease processes. This understanding will require the elucidation of speciÞc molecular mechanisms and their roles within complex biological systems. To develop this knowledge, researchers will need access to tremendous amounts of data and information. The accelerating output of industrialized, high-throughput research programs—such as genome sequencing projects, large-scale proteomics efforts, and single nucleotide polymorphism (SNP) mapping projects—promises to contribute vast amounts of data useful for drug discovery. However, the creation of enormous data and information repositories in the absence of adequate analytical tools may have the net effect of actually reducing R&D productivity. Many of the computational tools described in this article promise to enhance researchers’ ability to Þnd and access relevant data and information and thus increase their productivity. The universal data access technologies described in this article require considerable effort from information technology (IT) departments, which must Þt the receiving organization’s computer systems to those of the data source. However, the trend in new data access systems is toward common communication protocols in which the recipient and provider systems automatically inform each other of their requirements and tailor their communications to suit. Elements of this approach are found in the ISYS system (discussed in “Federated Database Systems”). The World Wide Web is likely to become the conduit for this type of approach on a large scale through the use of Web services. The Web services approach employs platform-independent technologies—such as Extensible Markup Language (XML) and Simple Object Access Protocol (SOAP)—to allow access to data sources through a Web service broker. In this scenario, the service broker enables the service requester (data users and applications) and the service provider (databases) to negotiate a communication protocol for the required data transaction. The automated common-format properties of Web services should promote just-in-time data integration without the need to completely revamp either the provider or requester systems. Once a connection has been established between the user’s computer systems and the provider’s databases, researchers face the greater challenge of locating relevant data and information. The profusion of terminology and data structures used in different databases to describe biological molecules and processes can severely limit the ability of computational tools to retrieve the appropriate information. The development and widespread adoption of standards for terminology and concepts that underlie database organization will be critical to accomplishment of this task. Standardization of communications and content will further impact productivity by driving down the cost of information identiÞcation and access. Because standardization of terminology and concepts involves the codiÞcation of human knowledge, progress will depend on the consensus of domain experts (experts in a particular Þeld who can deÞne terms and codify concepts) and developers of database collections. Several efforts are ongoing to

22

DATA AND INFORMATION INTEGRATION IN PHARMACEUTICAL R&D

develop uniÞed taxonomies of biological concepts (i.e., ontologies). One group leading these efforts is the Gene Ontology Consortium (www.geneontology.org), which is a group of biological database developers and curators. This consortium is producing a controlled vocabulary that can be applied to all organisms to describe molecular function, biological processes, and cellular components. The group intends to develop a mechanism to facilitate updates of the vocabulary as new information becomes available. Existing public biological knowledge taxonomies/ontologies include the Medical Subject Heading (MeSH) and the Uniform Medical Subject Language (UMSL) developed by the National Library of Medicine. A Þnal and important question is whether the new data and information technologies will truly increase R&D productivity and reduce the cost and length of the drug development process. The pharmaceutical industry’s lack of experience in the use of these tools precludes an accurate return-on-investment analysis of their implementation. However, the experience of other industries may provide insight into what may be expected. For example, Center Partners, a provider of outsourced customer care services, implemented the BrainEKP system to enable its agents to access product information more efÞciently. Like researchers in the pharmaceutical industry, Center Partners’ agents need to access a wide variety of information quickly. The company reports that the use of the BrainEKP system improved the quality of speciÞc services and, on average, reduced the time required to deliver them. Metrics for tracking increases in scientists’ productivity in pharmaceutical R&D are not readily available. However, drug companies that are considering further investment in these tools will need to develop ways to measure whether they actually do increase R&D efÞciencies. The number of compounds a company advances into clinical trials or introduces as NCEs each year can serve as estimates of R&D productivity; however, meaningful trends in these developments can only be observed over a relatively long time period. Alternatively, companies can track reductions in R&D expenditures at various development stages. This approach may provide the most useful near-term feedback on the impact of data and information integration technologies. If these tools dramatically reduce the time researchers spend on accessing and integrating data and information—perhaps by tenfold—then companies will be able to justify the costs of their implementation. REFERENCE IBM Business Consulting Services. Executive Strategy Reports. Pharma 2005—silicon rally: the race to e-R&D. 1999. Available at ibm.com/services/strategy/e strategy/pharmapubsilicon.html.

Impact of Technological Advances on Pharmaceutical Productivity

SUMMARY Productivity is a major concern of most pharmaceutical companies. Despite a wealth of new technologies that were expected to boost productivity by accelerating the R&D process, the pharmaceutical industry has experienced an overall decrease in productivity. The rate of new product launches for the top 20 Þrms has plunged drastically to less than one new product per year since 2000. Today, it is commonly accepted that no single technology can revolutionize drug discovery and development, although many new technologies have indeed revolutionized individual processes in discovery and development. In this article, we review the state of pharmaceutical productivity, discuss the ways in which companies are coming to grips with the problem—focusing on the particular strategies of three major companies—and assess the prospects for ending the productivity crisis. BUSINESS IMPLICATIONS •

•

•

The steady and dramatic downturn in pharmaceutical productivity is one of the major concerns facing today’s top companies. This longstanding trend has shown no clear sign of abating; nevertheless, we anticipate that drug candidate attrition rates will begin to improve within the next three to Þve years. Rising R&D costs, high rates of drug failures, increasing pressure on drug prices, and stricter regulatory controls make it increasingly difÞcult for companies to achieve the productivity levels necessary to maintain growth. Many technological advances—including, most recently, genomics and bioinformatics—have been introduced as potential means of accelerating pharmaceutical productivity. Although these technologies have been extremely

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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IMPACT OF TECHNOLOGICAL ADVANCES ON PHARMACEUTICAL PRODUCTIVITY

•

useful in some important ways, none has produced anything close to the results originally expected. To date, some technological advances have greatly accelerated many stages of drug discovery and development, particularly in the areas of target discovery and validation. However, because the result is a much larger ßow of “targets” through the system, and certain key bottlenecks remain, the overall process has not been substantially accelerated.

INTRODUCTION Productivity has become the major concern for the majority of pharmaceutical companies. The rate of new product launches has plunged drastically—from an average of 1.5 new products per year from each of the top 20 Þrms during the 1990s, to less than one new product per year since 2000 (Ansell J, 2003). It is particularly troubling that this reduction has happened despite the introduction of many new technologies—including high-throughput screening, combinatorial chemistry, and genomics—intended to increase productivity. In each case, however, the full promise of the technology has yet to be fulÞlled, and productivity has continued to decline. Today, it is commonly accepted that no single technology can revolutionize drug discovery and development. R&D alone has at least two critical components: identiÞcation of valid and tractable targets and design and generation of appropriate molecules to address those targets in humans. Addressing only a single aspect of either component has been insufÞcient to accelerate the overall process of Þnding a marketable drug. This article reviews the state of pharmaceutical productivity, ways in which companies are coming to grips with this issue, and prospects for solving this problem. THE PRODUCTIVITY PROBLEM Despite the introduction of a wealth of new technologies for R&D, the pharmaceutical industry has experienced an overall decrease in productivity. A slight increase in productivity did occur from about 1970 until the late 1990s, but since then, the number of new compounds has decreased steadily. IMS’s annual review of new active substances (NASs) shows that, in 2002, the industry reached a new twenty-year low of 36 NASs. The FDA approved only 21 new drugs in 2003, compared with 53 during 1996, the peak year for approvals. Some large companies did not receive a single approval in 2003. This dip in productivity is causing great concern because companies are unable to Þll their pipelines fast enough and to garner enough approvals to keep analysts and investors satisÞed. Meanwhile, the cost of drug discovery and development continues to rise. A recent study by Bain & Company (2003) estimated that the cost of developing a new drug to commercialization has reached $1.7 billion, while the cumulative success rate of compounds entering preclinical development

25

THE PRODUCTIVITY PROBLEM

TABLE 1. Major Pharmaceutical Mergers, 1999–2003 Company AstraZeneca (merger of Astra and Zeneca) ˆ Aventis (merger of Hoechst and Rhone-Poulenc Rorer) ´ ´ Sanofi-Synthelabo (merger of Sanofi and Synthelabo) Pfizer (acquisition of Warner-Lambert) Pharmacia (merger of Pharmacia and Upjohn and Monsanto) GlaxoSmithKline (merger of Glaxo Wellcome and SmithKline Beecham) Abbott (acquisition of Knoll) Bristol-Myers Squibb (acquisition of DuPont) Johnson & Johnson (acquisition of Alza) Amgen (acquisition of Immunex) Roche (acquisition of 50.1% in Chugai) Pfizer (acquisition of Pharmacia) Biogen Idec (merger of Biogen and Idec)

Date of Completion

Value ($B)

1999 1999 1999 2000 2000 2000

35.0 22.0 11.0 90.0 27.0 76.0

2001 2001 2001 2002 2002 2003 2003

6.9 7.8 12.3 17.0 1.3 56.0 6.8

has declined by nearly 50%. The consulting Þrm estimated that only one in six new drug candidates will deliver more than its capital costs. Mergers have been a popular short-term solution (Table 1). Companies can get a brief reprieve by combining two large pipelines under one roof, thereby reducing the overall stafÞng and infrastructure that support them. Often, however, merged companies soon Þnd themselves Þghting the same battle for productivity on an even larger scale. In addition, concern is growing within the industry that larger organizations are less innovative than their smaller counterparts; if this is true, merged companies could face an even tougher battle overall. A major question within the industry is whether this vicious circle—wherein companies are spending more and getting fewer novel products in return—is here to stay. Some experts believe that the “easy fruit” has been picked and that pharmaceutical R&D will be much more challenging in the future. Others believe that, although new technologies for drug discovery and development have disappointed so far, the industry is reaching a point where it has Þnally obtained a critical mass of late-stage products. These experts believe that the industry is just about to turn a corner and begin producing novel drugs at an accelerated rate. We agree. Another increasingly popular response to the productivity dilemma is the acquisition of rights to late-stage drug candidates (Table 2). The prices of such products have reached record levels. Aventis’s deal with Genta for cancer drug oblimersen, for example, is worth $480 million. PÞzer recently bid $1.3 billion for cardiovascular drug maker Esperion. However, there is now widespread agreement within the industry that most of the best late-stage products have been acquired. Certainly, new products will emerge, but if competition for them continues to escalate, their prices will make acquisition a less and less satisfying option.

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TABLE 2. Major Pharma/Biotech Product Deals and Alliances, 2002–2004 Companies

Date

Value ($M)

Forest Labs/Cypress Bioscience

1/2004

250

Wyeth/Curis Pharmaceuticals

1/2004

170

Johnson & Johnson/Scios

4/2003

2400

Pfizer/Esperion Therapeutics

12/2003

1300

Novartis/Idenix Pharmaceuticals

5/2003

747

GlaxoSmithKline/Theravance

1/2003

545

Millennium/Ortho Biotech

6/2003

535

Aventis/Regeneron

9/2003

510

Bristol-Myers Squibb/Lexicon Genetics

12/2003

450

Pfizer/Organon

10/2003

370

Novartis/Regeneron

3/2003

350

Merck/Actelion

12/2003

272

Comments Codevelopment and marketing of Cypress’s milnacipran (licensed from Pierre Fabre), in Phase III for fibromyalgia syndrome (FMS). Alliance for licensing and development of Curis Hedgehog proteins and novel small-molecule Hedgehog pathway agonists (activators) to Wyeth Pharmaceuticals for the treatment of neurological and other disorders. Acquisition of Scios, which focuses on cardiovascular and inflammatory diseases, includes nesiritide (Natrecor), marketed for congestive heart failure (CHF). Acquisition of Esperion, which focuses on development of high-density lipoprotein (HDL) targeted therapies for treatment of cardiovascular disease and has several candidates in development. Acquisition of 51% stake in Idenix, including licensing rights to Idenix’s hepatitis B candidate telbividine (LdT), in Phase III, and valtorcitabine (val-LdC), in Phase II, and development of other compounds. Alliance to develop and commercialize novel beta-2 agonists for treatment of respiratory diseases, utilizing GlaxoSmithKline’s drug delivery technology. Collaboration on commercialization and continued clinical development of Millennium’s bortezomib (Velcade), which was launched in May 2003. Codevelopment and commercialization agreement for Regeneron’s VEGF-Trap, currently in Phase I for cancer. Neuroscience alliance for discovery, development, and commercialization; Lexicon will contribute 13 drug discovery programs, and both parties will have exclusive access to Lexicon’s Genome 5000 program for future drug development. Codevelopment and commercialization of Organon’s asenapine, in Phase III trials for schizophrenia and bipolar disorder. Development and commercialization agreement for Regeneron’s Il-1 Trap, in Phase II for rheumatoid arthritis. Alliance to discover, develop, and market new classes of renin inhibitors for cardiorenal diseases.

TABLE 2. (continued) Date

Value ($M)

GlaxoSmithKline/Neurosearch

12/2003

270

Roche/Maxygen

5/2003

230

Pfizer/Eyetech Pharmaceuticals

12/2002

750

´ Sanofi-Synthelabo/Immuno-Designed Molecules

1/2002

545

Roche/Antisoma

11/2002

500

GlaxoSmithKline/Exelixis

10/2002

484

Aventis/Genta

4/2002

480

Pfizer/Neurocrine

12/2002

400

Schering/Immunex (Amgen) Eli Lilly/Amylin Pharmaceuticals

5/2002 9/2002

380 325

Johnson & Johnson/Tibotec-Virco

3/2002

320

GlaxoSmithKline/Nobex

5/2002

283

GlaxoSmithKline/Adolor

4/2002

270

Roche/Kosan Biosciences

9/2002

220

Companies

Comments

27

Five-year research and development alliance that comprises a number of programs within ion channels for treatment of central nervous system (CNS) disorders, including depression, anxiety, and schizophrenia. Collaboration on development and commercialization of Maxygen’s portfolio of interferon-alpha and beta candidates for hepatitis B. Codevelopment and commercialization of Eyetech’s pegaptanib (Macugen) for age-related macular degeneration (Phase III) and macular edema (Phase II). Collaboration agreement in cellular immunotherapy to develop and market immunological treatments for cancer. Roche acquired the rights to develop Antisoma’s entire oncology pipeline, including pemtumomab (Theragyn), in Phase III development. Discovery, development, and commercialization of novel therapeutics in the areas of vascular biology, inflammatory disease, and oncology. Codevelopment and commercialization of Genta’s Phase III oncology product, oblimersen (Genasense). Codevelopment and commercialization of Neurocrine’s indiplon (Phase III) for insomnia. Schering acquired Immunex’s sagromostim (Leukine). Codevelopment and commercialization of Amylin’s exenatidec (AC-2993), currently in Phase III for type II diabetes. Expansion of Johnson & Johnson’s portfolio of treatments for infectious diseases through acquisition of Tibotec-Virco, which has several compounds in development. Codevelopment, manufacturing, and marketing of orally administered insulin products for diabetes. Codevelopment and commercialization of alvimopan, in Phase III for management of bowel paralysis after surgery (postoperative ileus, POI) and constipation caused by opioid use, and development of other indications in outpatient settings. Clinical development of Kosan’s epithilone D (KOS-862), anticancer agent in Phase I development.

28

IMPACT OF TECHNOLOGICAL ADVANCES ON PHARMACEUTICAL PRODUCTIVITY

USING TECHNOLOGY TO IMPROVE PRODUCTIVITY A large number of the technologies that have been introduced during the last ten years were expected to accelerate drug discovery and development. However, the impact of many of these technologies has been much smaller than anticipated. Combinatorial chemistry and high-throughput screening provide examples: combinatorial chemistry enables rapid-Þre, automated synthesis of millions of chemical compounds; high-throughput screening can screen those compounds robotically. Both technologies opened doors and fundamentally changed the way drug discovery is done, adding an industrial aspect to lead identiÞcation. But neither technology has generated a rapid, sustained increase in the rate of new products approved. In fact, some experts argue that they actually slowed the process and eliminated a historically important factor in drug discovery—serendipity. When robots screen compounds in test tubes, they cannot detect potentially promising effects, which human researchers could sometimes observe in animal models. Genomics, which was accompanied by some of the most inßated promises, has provoked a serious backlash. As a widely quoted Lehman Brothers’ (2001) report anticipated, the result of the genomics technology boom has been a huge ßood of poorly validated targets. Instead of following the ßow of scientiÞc literature and investigating drug targets that were widely studied, researchers could pluck a bit of sequence from the genome, put it on a chip, and declare it a target because it was upregulated in diseased tissue. Unfortunately, genomics tools were relatively immature and required expertise for good results. Many spurious or useless data were generated, but because the targets were so novel, a great deal of research had to be done before they could be dismissed. However, the illusion that genomics was going to make drug discovery and development easier was powerful and widely accepted. After all, the Þeld was generating unprecedented amounts of data. For the time being, those data have simply created an even bigger and costlier bottleneck slightly farther down the pipeline in target validation. In retrospect, it is easy to say that the hopes that accompanied all these technologies were irrationally inßated. Drug discovery and development is an extremely complicated task, with so many variables and potential causes for failure that it seems na¨õve to expect a single technology to substantially increase the pace of discovery. But it is also true that new technologies have dramatically accelerated many individual processes in drug discovery. For example, now that multiple animal model genomes have been completed, researchers can almost instantaneously search the genomes of their favorite laboratory species for genes that are similar to a gene of interest. A wide range of new gene suppression techniques, including RNAi and antisense, can be used in concert with automated methods to do highthroughput gene function screens in multiple species. Gene sequences can be mined exclusively for targets that are amenable to high-throughput screens. And, increasingly, cellular assays can be designed to help break down drug mechanisms

USING TECHNOLOGY TO IMPROVE PRODUCTIVITY

29

of action, thereby enabling better understanding of many drugs’ effects long before the drugs enter the clinic. Researchers have also learned how best to apply the new technologies. Genomics, for example, is now being used much more aggressively for target validation than for target discovery. This approach is helping companies to move later-stage compounds forward more quickly, rather than accumulate more compounds in preclinical investigation and Phase I. More importantly, companies are now integrating these technologies in new and more thoughtful ways. The vision of a single industrial process for drug discovery, in which every step is high-throughput, automated, and guided by computer-generated analysis, has given way to a new paradigm: each project is guided by basic, hypothesisdriven biology and enhanced by appropriate use of the right set of machine and “brain-driven” tools. Skeptics must wonder if this hypothesis-driven approach is a real development or just another excuse for R&D labs to hide behind for the next few years—and if it is truly a development, when can we expect to see the rewards? An examination of pharmaceutical products’ rate of progress today reveals that while companies are pumping many more products into development, many more products continue to fail. Why should this situation change in the near future? Several trends suggest that technology will make an increasing contribution to pharmaceutical productivity. First is the widespread adoption of knowledge management tools—or, at least, a greater emphasis on interdepartmental communication and transparency. Advances in information technology and telecommunications are making it much easier to establish efÞcient, secure, and easy-to-use communication channels between teams. As a result, researchers can Þnd useful information from internal sources much more easily, and projects can be more quickly evaluated for their potential. Related to this greater sharing of information is a change in the way projects are selected and triaged, which is effecting a change in the actual scientiÞc research process: where individual researchers might formerly have labored for years on pet projects, now resources are allocated based on regular evaluations by integrated teams. The widely derided “silos” are breaking down as a result of new organizational structures. As scientists at PÞzer admitted in a recent review, even though researchers may resist the idea, “the world of drug discovery has become a managed process” (Schmid EF and Smith DA, 2002). Finally, and most importantly, instead of technology driving drug discovery, drug discovery is driving technology. During the genomics technology explosion of the late 1990s, companies felt pressured to adopt many new tools, sometimes without much consideration of what returns could be expected from them. Today, pharmaceutical companies are extremely clear about their research priorities, and those priorities are driving the development of tools for toxicity screening and biomarker development, two areas that are central to the industry’s current major problem: late-stage failures related to toxicity. As a result of these trends, scientists in research and development are getting much more of exactly what they need—quality information, assistance from

30

IMPACT OF TECHNOLOGICAL ADVANCES ON PHARMACEUTICAL PRODUCTIVITY

colleagues, and the technological tools they need most. This development should help the industry to deal better with the major problem at hand: to identify optimal targets and Þnd suitable compounds to address them. SELECT COMPANY CASE STUDIES While some companies are doing much better than others in terms of productivity, all consider productivity to be one of their chief challenges. More products are failing in clinical trials, and some experts speculate that Þnding new drugs will become increasingly difÞcult. However, most of the industry believes that the key problem is to learn how to exploit the next wave of targets and how to navigate the new, stricter U.S. regulatory environment. In this section, we proÞle four leading companies and some of their strategies for improving productivity. GlaxoSmithKline GlaxoSmithKline (GSK) is probably the best representative of the productivity problem. Having faced this challenge since its inception, the company has taken an innovative approach by creating a new R&D model that we discuss later in this section. One of the top Þve pharmaceutical companies in terms of sales, GSK employs approximately 100,000 people worldwide. Approximately 40,000 of the company’s staff are in sales and marketing; another 42,000 work at nearly 100 manufacturing sites in 38 countries. The company has an R&D budget of approximately $4 billion and employs about 16,000 R&D personnel at 24 facilities in seven countries. GSK had 2003 sales of $35.2 billion; pharmaceutical sales accounted for $29 billion. The company has a particular focus in four therapeutic areas: anti-infectives, central nervous system (CNS), respiratory, and gastrointestinal/metabolic. It is also a leading vaccine developer and has a growing oncology franchise. One of the most exciting compounds it has in development is a dual kinase inhibitor, currently in cancer trials. The company is an aggressive adopter of novel drug-discovery technologies and a standout in the implementation of genomics. GSK was ofÞcially formed in 2001 by the merger of Glaxo Wellcome and SmithKline Beecham. Its creation prompted many experts to consider whether pharmaceutical companies were starting to get too big. The company’s stock dropped signiÞcantly soon after the merger was completed and has inched up slowly, rebounding from more serious dips along the way. GSK’s most serious concern is its ability to produce new products over the next few years. The company is under considerable pressure from generics competition for some of its major drugs, and most of its most promising new compounds are several years away from approval. The fact that so few of these drugs are in later development stages poses greater risk to their prospects.

SELECT COMPANY CASE STUDIES

31

As part of its postmerger reorganization, GSK created a new R&D model by restructuring the company’s preclinical R&D into seven Centers of Excellence for Drug Discovery (CEDDs). Through this approach, the company hoped to reinject innovation into its huge research arm. Each of these centers evaluates and reÞnes drug leads in one of the following areas: • • • • • • •

Cardiovascular and urogenital diseases. Metabolic and viral diseases. Microbial, musculoskeletal, and proliferative diseases (cancer). Neurological and gastrointestinal diseases. Psychiatric diseases. Respiratory and inßammatory diseases. Biopharmaceuticals.

The seventh CEDD—biopharmaceuticals—was created in November 2003 and is the newest of the centers. It was launched because GSK expects to develop a growing number of such agents. Products will include monoclonal antibodies, therapeutic vaccines, and recombinant therapeutic proteins. Unlike the other six centers, which are devoted to midstage pipeline entities, the biopharmaceuticals center will focus on early-stage compounds. The most notable aspect of GSK’s R&D reorganization is that the CEDDs are essentially competitive. Resources are allocated according to success; Tachi Yamada, GSK’s R&D chairman, has publicly stated that unsuccessful CEDDs might be spun off. The fact that GSK sold its former Cell Map Unit to biotech Cellzome in 2001 adds weight to that threat. But the formation of the CEDDs is also a way to specialize preclinical drug development and ensure that optimal compounds go into clinical trials. According to a presentation that GSK made to investors in December 2003, the new R&D model is working. Compared with the numbers from the Þrst year of GSK’s existence, the company’s 2004 Þgures look encouraging. The company currently has 147 projects in clinical trials, including 82 new chemical entities (NCEs), 45 product line extensions (PLEs), and 20 vaccines. Since 2001, GSK has almost doubled the number of drugs in development, and its pipeline is maturing rapidly: NCEs in Phase II/III have increased from 23 to 44. (Table 3 compares the number of GSK products in clinical trials in October 2001 and December 2003.) In addition, GSK expects to Þle a record number of new drug applications (NDAs) in 2004–2008, along with 16 product Þlings in 2004–2005. The company believes that many of these products have the potential to achieve blockbuster status. However, as noted, many of these Þlings will be for PLEs, and GSK will rely heavily on new approvals for these established drugs. Because GSK is certain to lose signiÞcant revenues from generics competition in the near term, approvals for a large number of PLEs and NCEs will be essential. Given the industry’s recent track record in gaining approvals, achieving that goal may be a challenge.

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IMPACT OF TECHNOLOGICAL ADVANCES ON PHARMACEUTICAL PRODUCTIVITY

TABLE 3. Number of GlaxoSmithKline Products in Clinical Trials, 2001–2003 Phase

October 2001

I II III Total

27 15 8 50

December 2003 38 32 12 82

The company’s pipeline of new drugs needs to move forward relatively quickly, and with a high success rate, to restore full investor conÞdence. GSK anticipates a strong inßux of new compounds into its pipeline by 2006. Some of these compounds are indeed novel and represent signiÞcant potential markets; however, at their current, very early stages of development, they cannot yet be considered strong prospects. Nonetheless, the sheer number of drugs (147) in development is impressive. The company has clearly made a signiÞcant investment in new technologies. It has a massive genome-screening project searching for new targets. This endeavor is the largest of its kind in the industry, spanning 18 disease areas, and has been active since 1997. In addition, the company recently established a very sophisticated, ultra-high-throughput screening facility in Tres Cantos, Spain. GSK has integrated these two efforts and is concentrating its genome screens only on targets that are amenable to high-throughput screening. Novartis Novartis is the product of one of the earlier big mergers, that of Ciba-Geigy and Sandoz in 1996. The fact that both were Swiss companies and that the resulting company was large (but not immense) may have helped to make analysts more accepting of this union. Novartis is the industry leader in approvals, having received approval from the FDA for 11 novel compounds in the last four years. Novartis reported sales of $20.9 billion in 2002 for its consumer health and pharmaceuticals divisions. In 2003, the company’s pharmaceuticals business had $16 billion in sales. The company reports that R&D spending increased substantially in 2003, reaching approximately $3 billion. The company expanded its pipeline to include 79 projects in clinical development or registration. It is particularly encouraging that 63 of these projects are in Phase II or III. The company estimates that ten of these later-stage compounds could achieve blockbuster status. The Þve business units of Novartis Pharmaceuticals include oncology, transplantation, ophthalmics, mature products (branded, mostly older drugs), and primary care. Within primary care, the company specializes in cardiovascular, CNS, gastroenterology, dermatology, respiratory, and rheumatology/bone/hormone replacement therapy. In 2002, Novartis announced it was moving its pharmaceutical R&D headquarters from Basel, Switzerland, to the new Novartis Institutes for BioMedical Research in Cambridge, Massachusetts. This move was a result of the higher

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level of activity and innovation in the United States and Cambridge’s increasing popularity as a site for pharmaceutical R&D. Novartis has approximately 3000 R&D scientists located at its research institutes in the United States (Cambridge and East Hanover, New Jersey), Switzerland (Basel), United Kingdom (Horsham and London), Austria (Vienna), and Japan (Tsukuba). Novartis has been a pioneer of both genomics and electronic data capture (EDC) in clinical trials. In genomics, the company took an unusual path. During the genomics boom, many other companies, such as Merck and SmithKline Beecham, were rushing to discover genes, but Novartis decided to emphasize functional genomics. Besides establishing a strong functional genomics department within pharmaceutical R&D, the company also established the Genomics Institute of the Novartis Research Foundation (GNF) in La Jolla, California, partly to exploit California’s active biotechnology environment. Little obvious beneÞt has come from that move, however, and it is likely that instead of looking for synergies with biotechnology companies, Novartis will seek greater synergies between the GNF and the pharmaceutical R&D genomics department. In its focus on functional genomics, Novartis has been prescient. The race to patent genes has turned out to be worth little to the winners, and most companies are shifting their efforts to identifying gene function and determining which genes are the best drug development targets. Novartis has not yet reaped the beneÞts of its foresight, but the company is well positioned to make optimal use of genomics both for target discovery and for drug development. Novartis was also an early implementer of EDC and uses it for most of its clinical trials. EDC has obviated the need for manual data entry and facilitated the automated collection and transfer of data. The company reports that it has saved $50 million in database management fees by implementing its own EDC system and is on track to go from last patient visit to database lock in just one day—an extraordinary feat because using traditional methods, this process can take months, even a year or longer. Pfizer Like GSK, PÞzer has reached a size that raises questions about its ability to manage productivity. The company’s acquisition of rival Pharmacia in 2003 has buoyed sales; according to IMS, PÞzer earned close to $30 billion in 2003 in U.S. prescription sales alone—9.7% growth, compared with 2002 earnings. The company reported that its worldwide sales totaled more than $45 billion last year, a 40% increase of its 2002 revenue. Approximately $39 billion of this revenue was from pharmaceutical sales. But PÞzer now has to exceed those levels—its worldwide revenue goal for 2004 was $54 billion, unprecedented in the industry. Most of the restructuring related to the Pharmacia merger has been completed; PÞzer now has 120,000 employees worldwide. The company claims to have the world’s largest R&D organization: it spent $7.1 billion on R&D in 2003 and $7.9 billion for 2004. PÞzer has approximately 13,000 people in its R&D units, which are located at six international research sites. These scientists are working in 18 therapeutic areas on approximately 300 discovery projects.

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PÞzer is well positioned in the market. In 2003, according to IMS, 14 of PÞzer’s products were at the top of their respective therapeutic categories—more top-ranked products than any other company. The company sells 8 of the world’s 25 top-selling medicines, including atorvastatin (Lipitor). Still, by most accounts, PÞzer needs about twice as many new products per year as most of its rivals to maintain its success. The company’s stated goal is to submit 20 NDAs over a Þve-year period ending in 2006. Six of those NDAs are already Þled. A large number of those drugs will likely be licensed in or codeveloped. Because of its size, PÞzer can afford a great many deals and can offer the best terms when necessary. For example, when amortized over twenty years, the $1.3 billion recently paid for Esperion lowers PÞzer’s earnings per share by less than one cent. Esperion is crucial to PÞzer because this small company is developing drugs against arterial disease, including one popularly referred to as the “arterial draino” because it appears not merely to inhibit further plaque buildup, but to actually reduce the amount of plaque already present. PÞzer is aggressive in the deal arena, leaving little to chance. For example, PÞzer has deals with all four of the world’s major antibody companies: Abgenix, Cambridge Antibody Technology, Mederax, and Morphoses. In light of its new size, PÞzer has adopted a new strategy and now aims to develop drugs to improve health and reduce health care costs, which means developing a complete range of drugs against some of the world’s most common diseases. In cardiovascular disease, for example, the company aims to develop treatments for managing blood pressure, cholesterol levels, smoking, and diabetes. This strategy is intended in part to address the increasing pressure to reduce drug costs. By demonstrating the cost savings that disease-management drugs can effect, companies like PÞzer hope to lessen that pressure. Because of atorvastatin’s unprecedented success, PÞzer already has a huge share of the cardiovascular market. PÞzer would like to augment its cardiovascular franchise with several new entrants, particularly torcetrapib, which raises the level of high-density lipoprotein (HDL). This new compound raises HDL levels by inhibiting cholesteryl ester transfer protein (CTEP). PÞzer is comparing combination doses of torcetrapib and atorvastatin with monotherapy in clinical trials, hoping to eventually market a blockbuster combination product. Encouraging results from a Phase II trial were recently published in the New England Journal of Medicine (Brousseau ME, 2004). A record $800 million has been budgeted for the Þnal torcetrapib trials. PÞzer reports that it is the largest Phase III program any company has ever undertaken. Wyeth Formerly American Home Products, Wyeth employs more than 52,000 people worldwide, including nearly 6000 employees devoted to R&D. It has eight major R&D sites, including sites in Pennsylvania, New York, and Massachusetts (Cambridge). The company reported sales of approximately $9.5 billion in 2003; all but $1.5 billion of that amount was generated by pharmaceuticals. Wyeth spent approximately $2.2 billion overall on research and development in 2003.

OUTLOOK

35

Wyeth emphasizes pharmaceutical, vaccine, and biotechnology products and Þve major therapeutic areas: oncology, cardiovascular and metabolic diseases, inßammatory diseases, neuroscience, and women’s health and bone diseases. Over the past several years, Wyeth has undergone major changes to help integrate new technologies into the R&D process and to improve overall productivity. The company has taken three major steps: adding two new R&D functional teams, using an integrated approach to advance development projects, and outsourcing clinical trial data management as a cost-saving measure. One component of the R&D reorganization was the establishment of two groups, discovery medicine and experimental medicine. Both groups aim to help bridge the gap between discovery and development. The physicians and scientists in discovery medicine use information about human disease pathophysiology to help guide target selection and validation; they also aim to use new technologies to accelerate clinical trials. Meanwhile, the experimental medicine department facilitates appropriate incorporation of new technologies (e.g., gene expression analysis) into preclinical and clinical development. Although part of their goal is to help introduce new technologies successfully into the drug development process, the two groups focus on alignment of discovery and development. Lack of feedback between these two units has been widely cited as one of the major obstacles to efÞciency. Wyeth is also making a special effort to integrate information from a variety of groups when making decisions about which projects to advance. For example, high-throughput screening (HTS) projects are initiated by the genomics division and one of the Þve therapeutic area (TA) departments. The senior vice president of discovery approves these projects, but the decision process is supported by the chemistry and core technology divisions. Wyeth has six core technology divisions: cell culture, protein expression, protein biochemistry, the compound library, assay development, and HTS. In an unusual move, Wyeth cut 80 jobs in March 2003 after hiring Accenture to manage its clinical trial data. At the same time, the company transferred 135 employees to Accenture. The company reported that this action was a cost-cutting move. Wyeth appears to be on target to meet its new, more aggressive productivity goals. The company has more than tripled the number of drugs in development. Most importantly, it has exceeded its goal of Þling at least eight investigational new drug applications (INDs) every year. Over the next few years, Wyeth expects to Þle mainly for new indications or dosing regimens for existing products, but it hopes to keep on growing the number of drugs in its pipeline rapidly, with a total of 51 clinical development projects involving 36 separate compounds under way. OUTLOOK Pharmaceutical companies have responded to the productivity crisis in a variety of ways: • •

Mergers. Aggressive acquisition and licensing of late-stage products.

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IMPACT OF TECHNOLOGICAL ADVANCES ON PHARMACEUTICAL PRODUCTIVITY

• • •

Establishing new methods for evaluating projects and clinical trial candidates. Enhancing communication and transparency between teams and units. Becoming more selective and demanding of technology.

At this point, the impact of the productivity crisis is not reßected in overall sales. The worldwide pharmaceutical industry is still growing. But the pressure to increase productivity is likely to continue; in fact, it will quickly intensify because of increasing generics competition and price sensitivity. The positive aspect of this problem is that it has already generated dramatic change in the management and organization of pharmaceutical R&D—something many experts have long called for but which seemed difÞcult to achieve before the current pressure to increase productivity took effect. Pharmaceutical R&D has been criticized for lack of coordination and communication between divisions. Many experts also believed that there was too much independence within research and not enough objectivity in deciding which projects best satisÞed overall business objectives. In addition, because R&D departments are spending so much and producing less, management is forcing them to become more accountable. Again, while these corporate requirements may have negative effects (e.g., they have had a somewhat stißing effect on innovation overall), they should have a positive effect on the quality of products that do make it to market. Finally, critical technological advances have made huge differences in pharmaceutical timelines. They have not solved the productivity crisis, but recombinant biology, high-throughput screening, combinatorial chemistry, genomics, and bioinformatics have transformed drug discovery and have dramatically accelerated various stages of the process. Now, new technologies are emerging to Þll in the major gaps. For example, the introduction of reagents for RNAi and for doing gene expression analysis on formalin-Þxed, parafÞn-embedded (FPE) samples has made certain genomics tools much more powerful approaches for target validation and biomarker discovery and validation. RNAi facilitates the study of gene function by enabling much faster, genome-wide gene knockdown. Before the development of RNAi reagents, determining the function of a particular gene product was a long, costly, and very uncertain process. In another recent development, the ability to analyze gene expression analysis in FPE samples has enabled easier performance of powerful biomarker-generating experiments using many tumor samples. Tumor samples are typically Þxed in formalin and embedded in parafÞn. Although many years worth of such samples exist, these samples could not be used for gene expression analysis because their RNA is chemically altered. Consequently, researchers had to collect fresh frozen samples for each study. Because studies require hundreds of samples in order to provide useful information, this chore was a major impediment to research. It is possible that the road ahead will be even tougher than it has been in the past. Human biology is complex, and indeed, many of the simplest problems may have been solved already. But if the problems that lie ahead are more difÞcult, researchers are now equipped with a much more sophisticated and complete set

REFERENCES

37

of tools. These tools allow scientists to better understand the targets they address and the complete mechanism of action of any drug; they can also measure the impact of compounds with greater sensitivity in many more model systems. In addition, Þelds like pharmacogenomics, medical devices, and medical imaging are introducing novel technologies that can be used together with pharmaceuticals in innovative ways. Drug-covered stents are one excellent example of such innovation. We anticipate that drug/prognostic combinations and novel applications of imaging will also create whole new categories of products and greatly prolong the lives of many current and new drugs. The recent ßood of new preclinical and early-clinical-stage products into leading pharmaceutical companies’ pipelines could be a frantic attempt to appease investors, a sign of a true breakthrough in drug discovery and development processes, or a mixture of both. The last explanation is the easiest to believe, but the answer will be clearer over the next few years as those early-stage projects progress. We believe that lasting signs of the reversal of the productivity crises will start emerging soon. SpeciÞcally, we anticipate that within the next three to Þve years, pharmaceutical drug candidate attrition rates will begin to slow dramatically and several novel types of products (e.g., drug/prognostic combinations) will earn approval. Given the already burgeoning supply of compounds entering the clinic, this increase should greatly shore up company pipelines. Certainly, some ugly surprises lie ahead: some of these compounds will have to address novel targets, and because great redundancy exists in almost all biological pathways, most of these targets will be tough to hit cleanly and with sufÞcient efÞcacy. Nevertheless, the huge explosion of immature, poorly tested tools is over. The best of those tools are maturing, and key new enabling technologies are emerging. Therefore, we believe that the industry is well positioned to weather this particular storm. REFERENCES Ansell, J. Do mergers beneÞt R&D? Scrip Magazine. September 2003:28–29. Bain & Company, 2003. Has the pharmaceutical blockbuster model gone bust? Brousseau ME, et al. Effects of an inhibitor of cholesteryl ester transfer protein on HDL cholesterol. New England Journal of Medicine. 2004;350:1505–1515. Lehman Brothers. The fruits of genomics. January 2001. Schmid EF, Smith, DA. Should scientiÞc innovation be managed? Drug Discovery Today. 2002;7(18):941–945.

Advances in Medicinal Chemistry and Lead Optimization

SUMMARY Medicinal chemistry applies both wet chemistry and computational methods to design, synthesize, and optimize the biological properties of compounds. An important phase of medicinal chemistry is lead optimization, a process whereby researchers design, synthesize, and retest analogues of primary lead compounds. During this process, medicinal chemists seek to identify compounds that are bioavailable and potent and that have high afÞnity and selectivity for their biological target. The goal of lead optimization is to Þnd compounds that have acceptable pharmaceutical properties and can enter the next phases of drug development and clinical testing. In this article, we discuss advances in medicinal chemistry and its integration throughout drug discovery and early-stage development, and we proÞle several companies specializing in this Þeld. BUSINESS IMPLICATIONS •

•

Medicinal chemistry encompasses the discovery, identiÞcation, design, and optimization of biologically active compounds. Researchers have increasingly integrated medicinal chemistry into the upstream discovery stage, in which various screening techniques and design processes are applied to generate more drug-like compounds. Now, medicinal chemistry is also being integrated into the downstream, early-stage drug development process, in which chemists make highly targeted attempts to create drug candidates that have a good prospect of reaching the market and staying there. Medicinal chemistry is transitioning to an industrialized discipline that bears the hallmarks of modern, high-throughput drug discovery. Integral components

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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INTRODUCTION

•

•

39

include designed compound libraries, high-throughput parallelization technology, sophisticated databases, and molecular modeling. Parallel methods that accelerate processes and increase throughput are of increasing importance for lead generation and optimization. Development teams increasingly emphasize early lead prioritization by attempting to predict a compound’s potential to be developed into a pharmaceutically useful drug (developability). Because absorption, distribution, metabolism, excretion, and toxicity (ADMET) properties are so critical for a drug’s success, researchers are applying computational tools and in vitro models to predict a compound’s ADMET properties and subsequently prioritize the compound during development. By the end of this decade, advances that enable drug developers to bring only the most promising candidates into development could ultimately help shorten preclinical development times and reduce the number of late-stage failures. Advanced clinical trials could be much more tightly focused and far more predictable in their outcome. These changes will help reduce the overall length, cost, and risk of drug development for pharmaceutical and biotechnology companies.

INTRODUCTION Medicinal chemistry encompasses the discovery, identiÞcation, design, and optimization of biologically active compounds intended for pharmaceutical purposes. It applies both wet chemistry and computational methods to design, synthesize, and optimize the biological properties of compounds. Medicinal chemistry also analyzes drug metabolism and mode-of-action and determines a drug’s quantitative structure–activity relationship (QSAR). An important phase of medicinal chemistry is lead optimization, a process in which researchers design, synthesize, and retest analogues of primary lead compounds—which themselves have been developed and prioritized from validated hits (compounds that are active in drug screens). During this process, medicinal chemists seek to identify compounds that are bioavailable and potent, and that have high afÞnity and selectivity for their biological target. The goal of lead optimization is to obtain compounds that have acceptable pharmaceutical properties and can enter the next phases of drug development and clinical testing. In recent years, researchers have increasingly integrated medicinal chemistry into the upstream discovery stage, in which various screening techniques and design processes are applied to generate more drug-like compounds. Now, medicinal chemistry is also being integrated into the downstream, early-stage drug development process (see Figure 1). A discussion of advances in medicinal chemistry must therefore also include in vivo lead optimization testing and the database technology used to manage and evaluate resulting data. This article focuses on advances in medicinal chemistry and its integration throughout drug discovery and early-stage development. We emphasize parallel strategies in medicinal chemistry and trends in early-stage lead prioritization, and

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ADVANCES IN MEDICINAL CHEMISTRY AND LEAD OPTIMIZATION

FIGURE 1. The central role of medicinal chemistry in drug discovery and development.

we discuss various other technological advances. In addition, we proÞle several companies that are specializing in medicinal chemistry and lead optimization. PARALLEL STRATEGIES IN MEDICINAL CHEMISTRY Parallel methods that accelerate processes and increase throughput are of increasing importance for lead generation and optimization. The concept of “parallelization” in chemistry is similar to parallelization in computing: the term refers to accelerating a process by running a multitude of tightly coordinated subprocesses (which are variations of a common theme) in batches during the same period. After the process is Þnished, analysis of the processes can also proceed in a batch manner. Researchers have developed many parallel methods in chemistry and applied them to accelerate and streamline the drug discovery process. An example of this approach is parallel synthesis, a method whereby a series of synthetic chemical reactions are conducted simultaneously (see Figure 2). In parallel synthesis, one key parameter—such as the concentration of a reactant—is varied at a time. Parallel synthesis is now the preferred method for creating focused compound libraries for lead optimization. To fully capitalize on the advantage of parallel methods, researchers must integrate them with a project database that, ideally, is tied into a corporate knowledge management system. Linking parallel chemical processes with knowledge management systems can reduce redundancy in tasks, such as compound library building and testing, and helps avoid bottlenecks during the data evaluation phase. Parallel Approaches in Solid-Phase Synthesis Solid-phase synthesis Þrst became popular in the early 1960s when Dr. Bruce R. MerriÞeld and co-workers introduced it as a method of peptide chain synthesis (Merrifeld won the 1984 Nobel Prize in chemistry for this work). In this approach, the Þrst reactant (a molecular building block) is covalently bound via a linker to a chemically inert, solid support (resin) such as a plastic bead. Further reagents can be added sequentially, and excess, unbound reagents can be washed away at the end of each synthetic step. At the end of the reaction sequence, the reaction product is cleaved from the solid support. In the 1990s, new forms of this general method were applied to the synthesis of organic, small-molecule compounds.

PARALLEL STRATEGIES IN MEDICINAL CHEMISTRY

41

FIGURE 2. Schematic of parallel synthesis.

Solid-phase synthesis is now a commonly applied strategy for the preparation of large combinatorial chemical libraries. Advantages and Limitations. Solid-phase synthetic methods enable multistep chemical reactions and can simplify and accelerate puriÞcations between reaction steps; nevertheless, they have some disadvantages. The support resin can withstand only relatively benign reaction conditions, its loading capacity may be low and therefore limit the scale of reactions, and reactions may be slower and more difÞcult to monitor than solution-phase reactions. Researchers have developed various techniques to address these limitations, including using solid-supported reagents and scavengers that can easily be removed from the reaction mixture. When solid-phase methods are used to produce mixtures of compounds (e.g., via the split-and-pool method; see Figure 3), chemists also face the difÞcult task of determining the structure of compounds that are found to be active in biological screens. Thus, researchers and companies have developed

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ADVANCES IN MEDICINAL CHEMISTRY AND LEAD OPTIMIZATION

FIGURE 3. Schematic of split-and-pool synthesis.

several innovative encoding and tagging strategies for tracking and identifying compounds throughout the synthetic process. These methods generally involve adding a chemical or physical “tag” to each bead that essentially records its chemical reaction history. Positionally Encoded Arrays. Combining positionally encoded solid-phase supports with the split-and-pool technique offers medicinal chemists a powerful tool for efÞciently creating and using compound libraries. In this approach, solid supports are arranged in an array format, and compounds are synthesized on the solid supports. This strategy greatly facilitates the deconvolution effort because the position of the compounds in the array is preserved throughout the sequential synthetic steps and thereby introduces an ordering (encoding) principle for their expected chemical structure. Identifying groups of desirable reaction products is much faster, and screening them can be much more focused. Interest in classical solid-phase synthesis surged in the second half of the 1990s and then waned during the Þrst years of the new millennium. Now, the

PARALLEL STRATEGIES IN MEDICINAL CHEMISTRY

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increased availability of automated instrumentation is making solid-phase array technologies that use two- or three-dimensional positional encoding popular as a method for building compound libraries. A good example of two-dimensional positional encoding in parallel synthesis is the Spot method. This method was initially developed for peptide synthesis but can also be applied to medicinal chemistry (in cases that use certain types of relatively mild chemistry and require only small amounts of reaction product). Small droplets, each containing a single compound, are spotted onto an inert polymer membrane in an array pattern and allowed to dry. The membrane array can then be subjected to chemical reactions during which the coordinate of each spot is preserved and thereby serves as a code for the identity of each compound. Researchers are also applying photolithographic technologies, which were originally developed for oligonucleotide arrays, to solid-phase chemical synthesis. Three-dimensional compound arrays add another dimension to the positional coding approach. This approach uses sturdier carrier structures and allows much more compound to be synthesized than with the Spot method. These carriers can also be individually encoded using tags that are compatible with automation, such as bar codes and radiofrequency chips. The tags serve as unique identiÞers for each position in the 3-D array and allow samples to be removed from the array for complete processing. Tag-Encoded Methods. Discovery Partners International’s Irori division provides products for solid-phase chemical synthesis and tracking. Irori’s Directed Sorting technology can be used to synthesize compounds within separate microreactors (Kans) that contain a solid support resin and either a glass-encased miniature radio-frequency tag or a two-dimensional bar code. The company’s sorting stations handle the microreactors according to their radiofrequency tags. Novel solid-phase tagging technologies include SynPhase Lanterns and so-called Lucky Charms. Mimotopes created SynPhase Lanterns, which are ßow-through cylinders for solid-phase synthesis of organic small molecules or peptides and which use radiofrequency tagging. Chemical reactions proceed on a reactive mobile polymer phase grafted onto a nonreactive rigid polymeric support. SynPhase Lanterns can also be used as scavengers in solution-phase chemistry. Another original technique is called Euclidean-shape-encoded combinatorial libraries, or Lucky Charms. Developed at the Scripps Research Institute in La Jolla, California, Lucky Charms are swellable polymer carriers that can be distinguished by their different geometric shapes (Vaino AR and Janda KD, 2000). Parallel Solution Synthesis in Arrays Many array-based solution synthesis systems are available for conducting parallel chemistry on a scale compatible with the demands of lead optimization. One example is Scigene’s FlexChem II Synthesis System, in which parallel synthesis is carried out in glass reactor vials. The vials are used in arrayed sets of 96 or 24 in a format that is compatible with automated liquid-handling equipment.

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Compound Purification and Management To keep up with the output of parallel synthesis in lead optimization, compound puriÞcation technologies also need to accommodate higher throughput. Purity is important because the use of puriÞed libraries can increase the efÞciency of highthroughput compound screening and because it avoids misleading results introduced by contaminating pharmacologically active by-products of synthesis. The drug discovery industry recently began to reemphasize puriÞcation and quality control of lead optimization libraries (processes that had waned in importance during the late 1990s) prior to their use in compound-screening operations. Three trends in library puriÞcation are now apparent: an increased focus on phase trafÞcking techniques aimed at integrating synthesis with puriÞcation; the application of polymeric and/or solid-phase extraction; and the use of puriÞcation methods, such as supercritical ßuid chromatography, a technique that is typically three to Þve times faster than conventional high-performance liquid chromatography (HPLC). Several companies offer compound puriÞcation systems and/or services. One of the current leaders is Argonaut Technologies, which offers the Trident Sample Processing Station, a liquid-handling system for extraction and Þltration, and the Flash Master II system, which has binary gradient capability and can purify up to ten samples in parallel. Berger SFC, a member of the Mettler-Toledo Group, specializes in supercritical ßuid chromatography. Combipure provides library screening and puriÞcation services and specializes in quality control and puriÞcation of lead optimization libraries. Discovery Partners International’s Accelerated Retention Window (ARW) HPLC technology accelerates compound puriÞcation and reduces the number of steps required for postpuriÞcation processing and the use of materials. Although compound management (the storage, handling, and retrieval of compounds) may be somewhat remote from medicinal chemistry, it nevertheless is a critical and nontrivial component of pharmaceutical laboratory operations. Ideally, compound management should be fully integrated with the design, synthesis, and puriÞcation of the lead optimization library. Although the loss of any particular library compound has little signiÞcance in the discovery stage, such a loss can have a severe impact during lead optimization. Today’s medicinal chemists rely on computerized handling systems for compound management, such as systems from the Automation Partnership, that are integrated with compound management databases. High-Throughput Crystallization The inclusion of systematic screening for pharmaceutically optimal solid forms (i.e., salts, solvates, and polymorphic crystals) at the early lead-optimization stage is a new development in medicinal chemistry. Surprisingly, drug developers had, until recently, considered this step a matter of the later stages of the drug development process. However, the choice of the salt form can signiÞcantly affect the bioavailability (the ability of a drug to be absorbed from the gastrointestinal tract, enter general circulation, and become available to the target tissue) of solid drug formulations, and the spontaneous conversion of polymorphic salt forms can greatly affect stability. Regulatory agencies worldwide now require companies to

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make a reasonable effort to identify and characterize critical polymorphs of the compound and to control for variations that could affect the product’s bioavailability. Going into in vivo testing with the right solid form not only avoids the additional studies that would be required if the form had to be changed but also discloses the full potential of a lead in the early stages of development. The explanation for the earlier neglect of the so-called salt and crystal factor during lead optimization is that X-ray crystallography, the technique used to determine a compound’s three-dimensional crystal structure, was historically a manual, low-throughput, labor-intensive process. Strategies to miniaturize and perform crystallization studies in parallel have only recently become available. Companies that pioneered these approaches for protein targets (and protein–ligand interactions) include Syrrx and Structural Genomix. Several companies have commercial systems that greatly facilitate screening for crystal forms by providing standardized reagents and/or conditions in plates or removable vials. They include Hampton Research (Crystal Screen HT, Index HT), Molecular Dimensions (Structure Screen), and deCode Biostructures (Emerald Biostructures Products). TransForm Pharmaceuticals offers clients high-throughput crystallization studies and formulation design (see “Corporate Highlights”). Chemical Genetics in Library Design In the broadest sense, chemical genetics refers to the large-scale use of smallmolecule organic compounds to determine protein function and to identify and validate drug targets. By introducing chemistry into the earliest stages of discovery, chemical genetics permits researchers to simultaneously select drug targets and generate small-molecule drug leads. In the forward chemical genetics approach, researchers would select compounds that bind to and alter the activity of a speciÞc cellular protein and cause an observable phenotypic change in the organism. They could then use the compounds to identify the protein targets of the selected compounds. More interesting to the medicinal chemist is the reverse chemical genetics approach, which can also allow lead optimization to proceed almost in parallel with the validation of a target. In reverse chemical genetics, potential target proteins can be used to probe compound libraries. By focusing further optimization attempts on those compounds that exhibit binding (above a predeÞned threshold) to the target, the lead optimization library can be improved and winnowed to a smaller number of leads that can then be used to observe their in vivo effects in cell cultures, tissues, or animal models. These observations can then be used to infer plausible biological roles of the target proteins. Cellular Genomics, Vertex Pharmaceuticals, and InÞnity Pharmaceuticals (a company founded by Harvard University’s Dr. Stuart Schreiber) are prominent examples of biotechnology companies using the chemical genetics approach. MICROWAVE-ASSISTED SYNTHESIS AND RANDOM IRRADIATION METHODS Many reactions employed by medicinal chemists must overcome the “activation energy” threshold in order to proceed. Chemists usually overcome this threshold

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by providing energy in the form of heat, which imparts kinetic energy to the reactant molecules and vibrational energy to some of its chemical bonds. Imparting this activation energy by irradiating the reactant mixture can offer possibilities far beyond conventional heating and allow access to chemistries that are not easily reached in more traditional ways. The use of microwaves to aid chemical synthesis can cut reaction times from tens of hours to a few minutes or seconds. Because microwaves excite chemical bonds directly instead of kinetically, microwave-assisted synthesis can also be applied to reactions that are conducted at very low temperatures (as low as −100◦ C) to control undesirable side reactions. Microwave-assisted synthesis allows rapid reaction sequences to proceed, an approach that is ideal for the iterative process of alternating synthesis, library validation, and testing used to build focused lead optimization libraries (Blackwell HE, 2003). This cutting-edge Þeld is even expanding to include carbohydrate chemistry. Drug developers used household microwave ovens for microwave-assisted synthesis until 2000, when Personal Chemistry (now part of the Discovery Chemistry division of Biotage) introduced the Þrst single-mode microwave reactor speciÞcally designed for organic synthesis. The company’s state-of-the-art product is Coherent Synthesis, a computerized system consisting of a workstation, software reaction kits, and other accessories. The commercial leader in microwave synthesis is now CEM. The company’s Focused Microwave technology is an open-architecture, user-conÞgurable platform for microwave-assisted synthesis. Of CEM’s line of equipment, its Discover System, Navigator Automated Microwave Compound Factory, and Explorer Automated Optimization Systems are of most interest to the medicinal chemist. Although the utility of microwave-driven reactions lies in accelerating and facilitating chemical reactions that are essentially controlled and predictable, gamma irradiation introduces a strong random element that can be useful when more signiÞcant changes to a molecule are desired. Irradiation with a high-energy 60 Co source breaks most reactant molecules down into hundreds of fragments and forms radicals that can recombine to produce distinct reaction products. If the conditions are chosen properly, a signiÞcant fraction of these products will have molecular weights and solubilities that satisfy the Lipinski Rule of Five (see “Early Lead Prioritization” for a description of the Lipinski Rule). Because the solvent is usually aqueous, deconvolution of such semirandom minilibraries (i.e., their fractionation into hierarchical classes of compounds according to structural relation) is relatively straightforward. COMPOUND- AND REACTION-CENTERED DATABASES Pharmaceutical R&D departments generate increasing amounts of chemical and pharmacological data to help them better decide which compounds (or targets) to pursue. Theoretically, these data should provide the basis for a data-driven decision support architecture in which every piece of data is captured, archived with proper indexing, and made readily available for interpretation from a wide variety of perspectives. Although the pharmaceutical industry has not yet achieved

VIRTUAL SCREENING AND STRUCTURE-BASED DESIGN FOR AFFINITY OPTIMIZATION

47

such an enterprise wide support structure, medicinal chemistry departments have made great strides toward this goal. Efforts to abstract huge amounts of data in order to facilitate their handling while allowing better access to the underlying information are progressing on several avenues. Researchers have designed mathematical descriptors that store essential information on compound structure and link it to biological effects; this computational Þngerprinting can greatly facilitate the elucidation of QSAR that is so essential to lead optimization. Chemical reactions that create modiÞed leads can also be stored in an algorithmic form (reaction phenotyping). If such databases are properly annotated, they can make medicinal chemistry knowledge much more directly available than conventional, purely relational databases can. APPLICATIONS OF VIRTUAL SCREENING AND STRUCTURE-BASED DESIGN TO LEAD AFFINITY OPTIMIZATION Chemists use computational structure-based design and virtual screening tools to screen virtual compound libraries and select and design drug-like molecules that will most likely be pharmacologically active. At Þrst sight, computational modeling techniques such as receptor–ligand docking should provide researchers with an almost magical solution to the problem of optimizing a lead’s afÞnity for its target. In concept, researchers could modify a lead compound on the computer screen, dock the modiÞed structures to a model of the lead’s target, and select the highest-afÞnity molecules for synthesis. This approach should enable such huge strides toward afÞnity-optimized compounds that only a little wet medicinal chemistry would subsequently be needed to complete lead optimization. Although this scenario is feasible in principle, its application is still unrealized. The failure of pure computational lead optimization lies not in a lack of raw computing power or clever programming of docking software but rather in the lack of sufÞciently detailed structural information about targets. Only 10–15% of all proteins can be adequately crystallized for three-dimensional structure determination via X-ray crystallography. This percentage is even lower for the active sites of enzymes; it is also very low for G-protein-coupled receptors (GPCRs), an important class of current drug targets. Moreover, to perform meaningful lead optimization, high-resolution X-ray structures are required not only for the active site but also for the ligand–target complex in a conformation that resembles the in vivo state. Also, subtle interactions between a ligand and its binding site can alter the conformation of both partners in fashions that are frequently unpredictable. Because of these factors, computational screening and design tools have been more successful when applied to lead discovery (in which the criteria for quantitative prediction are less stringent) rather than to lead afÞnity optimization. Computational techniques have also had an impact on predictive ADME (absorption, distribution, metabolism, and excretion) and toxicology approaches (see “ProÞling ADMET Properties”). The utility of computational tools for lead afÞnity optimization may improve over the next few years as knowledge-based computational paradigms are increasingly applied. For example, Predix Pharmaceuticals (see “Corporate Highlights”) has developed a unique, proprietary

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computational technology for predicting the three-dimensional structure of any GPCR, even in the absence of high-resolution physical data. Tools that combine numerical algorithms with databases containing metadata on structurally related targets and ligands will also aid in the development of better computational tools for medicinal chemistry. EARLY LEAD PRIORITIZATION A pervasive concept underlying recent developments in medicinal chemistry is that one priority lead compound should be selected as quickly as possible. Additional, lower-priority leads are typically kept in reserve, and often programs to characterize these compounds—which might be needed as backups if the priority lead fails—proceed simultaneously, albeit more slowly. A lead discovery and selection process might be compared with a military campaign in which the commander can never be sure that he has all relevant knowledge about the enemy force but nevertheless must at some point proceed to attack. Likewise, the head of a lead optimization team has to drive the process to the point at which an acceptable compromise among time, material resources, and data collection is reached. Although development teams cannot make lead prioritization decisions with complete certainty, today’s technologies enable chemists to approximate certainty better than ever before. The Shifting Definition of Developability The most basic requirements for lead selection are the compound’s afÞnity for the intended target (the avidity with which it binds to its target) and the compound’s selectivity (the ability of a compound to discriminate between target and nontarget molecules—which are often similar). Beyond these requirements are other criteria important for predicting a compound’s potential to be developed into a pharmaceutically useful drug—its developability. Although the concept of a compound’s developability has always been a central objective for medicinal chemistry, the criteria for determining developability are changing dynamically in number, type, and content. To determine whether a drug is developable, three pertinent questions must be answered: •

• •

Will it be possible to get enough of the compound to the target tissue, and will it remain there long enough to have a clinically useful therapeutic effect? Will the compound have unintended and/or toxic effects when administered in the doses required? Does the compound have the pharmaceutical properties necessary to make a good drug?

Ideally, researchers should begin to address these questions at the earliest stages of drug discovery by employing approaches that will generate drug-like compounds that can be screened for biological activity. However, current drug

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49

discovery technologies, which aim to accelerate discovery and frequently rely on combinatorial chemistry, tend to generate compound structures that are larger and less soluble (and therefore more difÞcult to optimize and therefore less developable) than those pursued in earlier times. This tendency cannot be entirely avoided, even if library construction is purposefully biased toward compounds with more acceptable physicochemical properties. Advances in drug delivery technology can ameliorate some of the disadvantages of using larger and less soluble compounds. Still, the need to work with leads that are more difÞcult to optimize is a key reason why medicinal chemistry is now a serious bottleneck in drug development. Probably no single individual has had a greater inßuence on medicinal chemistry than Dr. Christopher A. Lipinski of PÞzer’s Global Research and Development. Lipinski (now retired) established the famous Rule of Five in the 1990s as a guideline for quickly proÞling compounds with respect to their drug-likeness. The Rule of Five considers molecular weight, the number of hydrogen bond donors, aqueous solubility/hydrophobicity, and the number of hydrogen bond acceptors—to determine the likelihood of a compound being poorly absorbed (see Table 1). (The value limit for each of these parameters is a multiple of Þve: hence the term Rule of Five.) Most drug discoverers have embraced Lipinski’s rule (or some of its variants) as a general guideline for eliminating compounds and for designing drug-like libraries. Despite its utility, many successful pharmaceutical compounds violate the rule, and it has limited applicability in lead optimization in which medicinal chemists work with relatively small, structurally restricted optimization libraries. Chemists also use other criteria tailored to the particular compound family to assess its potential during lead optimization. For example, the polar surface area (PSA), a parameter that can be directly calculated from a compound’s chemical formula, can be combined with in vivo-determined brain penetration data on related compounds to generate a good computational predictor of absorption and blood–brain barrier penetration.

TABLE 1. Lipinski’s Rule of Five and Its Limitations Parameter Molecular weight

Lipinski Value Limitsa Molecular weight is greater than 500 daltons

Remarks

Current drug delivery technology can increase the bioavailability of certain compounds that exceed this limit. Hydrogen bond Number of hydrogen bond Frequently invalid for larger interaction donors is greater than five molecules. Aqueous solubilOctanol/water partition Solubility might be influenced by ity/hydrophobicity coefficient (c Log P) is greater particle size and than 5 pharmaceutical form. Heteroatoms for orbital Sum of hydrogen bond acceptors Generally valid. interaction is greater than 10 a According to the Rule of Five, poor absorption is likely under these conditions.

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Profiling ADMET Properties Drug-like molecules possess pharmaceutically desirable ADME properties: they can be orally absorbed; they are distributed to the desired therapeutic target; they are metabolized to nontoxic metabolites in a reasonable length of time; and they are excreted (or eliminated), also within a reasonable amount of time. In addition, ideal drug-like compounds are not toxic to humans. Because these properties are so critical for successful drug development, researchers have developed computational tools and in vitro models for predicting a compound’s ADME/toxicity (ADMET) properties and subsequently prioritizing the compound during development. Computational Profiling. Computationally predicting a compound’s absorption and distribution properties (both of which underlie its bioavailability) has proved easier and less complex than predicting its metabolism and excretion. Thus, most commercial activity in computational ADME prediction has been concentrated in the areas of absorption and distribution. Aqueous solubility (the mass of dissolved compound per volume unit of saturated solution) and hydrophobicity (also lipophilicity; lack of afÞnity for water as measured by the octanol/water partitioning coefÞcient) are key properties for all molecules handled by medicinal chemists. These properties are linked to each other and tie into the Lipinski rule. They are also used to predict a compound’s oral absorption and bioavailability: a compound with insufÞcient aqueous solubility will be unsuitable for oral administration unless its activity is very high; a compound with insufÞcient hydrophobicity might be easily soluble in water but would not be able to passively diffuse through cellular membranes. Methods to measure solubility and hydrophobicity experimentally are straightforward and well established, but they are difÞcult to adapt to higher-throughput formats and they require the compounds to be synthesized in relatively large amounts (at least tens of milligrams) and in relatively high purity. As a result, much attention is now devoted to developing computational methods of predicting solubility and hydrophobicity. Such methods would allow researchers to focus their efforts on synthesizing only the most promising compounds. However, the parameters of solubility and hydrophobicity are notoriously difÞcult to predict across highly diverse chemical structures. Most commercial software packages make ab initio predictions (i.e., based on the chemical formula alone, without experimental data from related compounds) with only limited accuracy. Fortunately, medicinal chemists rarely need to work completely ab initio; measuring these parameters on relatively few key derivative molecules synthesized during lead optimization provides experimental data. This information can be used to calibrate and Þne-tune the prediction process and allow computational methods to replace some of the iterative steps of synthesis and measurement. The earliest computational ADMET approaches developed in the 1990s were essentially based on reductionist paradigms (i.e., they operated on the belief that the broad spectrum of ADMET properties seen among drug candidates should be governed by a limited set of abstract rules). However, because different pharmacological mechanisms can lead to the same property, algorithms for their

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TABLE 2. Select ADMETa Prediction Software Suitable for Lead Optimization Software Package

Vendor

KnowItAll ADME/Tox Edition Discovery Studio MedChem Explorer ADME

Bio-Rad Laboratories

GastroPlus

Simulations Plus

Idea pkExpress

Lion Biosciences

ADME Boxes 2.2

Pharma Algorithms

”Cloe PK

Cyprotex Discovery

BioPrint

Cerep

Accelrys (formerly a subsidiary of Pharmacopeia)

Remarks Uses consensus modeling environment technology. MedChem Explorer module enables design and optimization of compound libraries in an Excel environment. Predicts oral absorption of compounds using neural network technology; several additional modules available. Predicts oral absorption from in vivo data. Software modules for various ADME predictors. Provides virtual human and rat models to predict pharmacokinetic properties. Subscription-based pharmacoinformatics platform with ADME modules.

a ADMET, absorption, distribution, metabolism, excretion, and toxicity.

prediction must be capable of handling interactions of compounds with more than one pharmacologically relevant key mechanism. Because such multiple modes of action are always present, prediction methods based on chemical structure alone would have to operate on an impractically large set of rules—provided that all these rules were known (which is not currently the case). In modern ADMET prediction software, a combination of QSAR and knowledge-based approaches are applied. Table 2 lists select commercial ADMET software offerings. Researchers have developed other tools as well, including those narrower in scope as well as highly specialized toxicology packages, many of which are not yet commercially available. In vitro Methods. Most drug metabolism occurs in the liver, so hepatic metabolism determines the bioavailability of a drug as well as its metabolites, some of which might be pharmacologically active in ways that may be either desirable (e.g., prodrug metabolites that induce the therapeutic effect) or highly undesirable (e.g., metabolites that induce toxic side effects). Determining a compound’s metabolism is therefore a critical component of lead prioritization and optimization. The standard approach to elucidate hepatic metabolism is to incubate a compound with liver microsome preparations from several species typically used in drug development. Rat, dog, rabbit, monkey, and human preparations are available from several commercial sources. Liver tissue slices are also frequently employed to corroborate the results obtained with the respective microsome preparations.

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An emerging approach that recently became commercially available utilizes cultured liver cells. MultiCell Technologies has established a porcine hepatocyte cell line (HepLiu) that proliferates in culture and retains typical hepatic functions. The company has also created the second-generation cell line e-HepLiu by engineering HepLiu to increase expression of CYP3A4 (an important drugmetabolizing enzyme) nearly tenfold. According to MultiCell, the cell lines closely emulate the metabolic state of the human liver; this feature suggests they could serve as useful tools in catabolic and liver toxicity optimization. Excretion has so far resisted most efforts at theoretical modeling or in vitro testing. Researchers are, therefore, very interested in the development of assays that can identify the uridine diphosphate (UDP)-glucuronosyl transferase (UGT) enzymes involved in hepatic glucuronidation, a metabolic pathway in compound detoxiÞcation and, subsequently, excretion. Once brought into standardized formats, such assays could serve to predict excretion in vivo. Metabonomics Metabonomics is the systematic, comprehensive analysis of the full complement of nonpeptidic, organic, low-molecular-weight molecules (metabolites) in a particular cell, tissue, biological sample, or organism. In this emerging technology, researchers perform nuclear magnetic resonance (NMR) or mass spectroscopy on sequentially obtained unprocessed body ßuid samples from a laboratory animal and subject the resulting data to a statistical analysis. This process can reveal correlations between particular metabolic Þngerprints (or proÞles) and speciÞc physiological states. During lead optimization, differences between closely related compounds can lead to different physiological effects on an organism. Compounds that elicit metabolic Þngerprints that have been previously determined to be relatively innocuous can be prioritized over others that show more complicated patterns. Therefore, metabonomics provides a new tool for the prioritization of leads. Toxicogenomics As its name indicates, toxicogenomics is a marriage of toxicology and genomics; it combines the conventional tools of toxicology with microarray-based analysis. Brießy, toxicogenomics addresses the question of how toxicological markers shift in response to drug exposure. Although lead optimization is only one of its many applications, toxicogenomics is bound to become increasingly helpful to medicinal chemists as more knowledge about drug-induced changes at the molecular level becomes publicly available in structured formats. A step toward this goal is the creation of Minimum Information About a Microarray Experiment in Toxicology (MIAME/Tox), which is a set of guidelines deÞning the minimum information required to clearly interpret and potentially reproduce and verify array-based toxicogenomic experiments. MIAME/Tox’s major objective is to guide the development of toxicogenomics databases and data management software. Researchers are now developing such databases at the National Institutes of Environmental Health Sciences (NIEHS) National Center

CORPORATE HIGHLIGHTS

53

for Toxicogenomics and at the European Molecular Biology Laboratory (EMBL) European Bioinformatics Institute (EBI). Predictive Biosimulation The ambitious approach of predictive biosimulation attempts to integrate genomic, proteomic, and physiological data about an organism (including humans) into a computational model that can then reproduce the organism’s key processes and responses in both healthy and disease states. Although much of what happens in an organism on the molecular level is still not completely understood, most initial efforts in predictive biosimulation have used a “bottom–up” modeling approach. In this methodology, scientists begin with relevant sets of genes, proteins, and other molecules and attempt to model their interactions within a cell. By running simulations that can be compared against experimental data, developers have demonstrated that some of these systems can, in some circumstances, serve as operational models of biological processes. Entelos uses a “top–down” strategy. In this approach, researchers deÞne a process of interest—such as a particular disease state or drug response—and then add in models of component subsystems until sufÞcient detail is present to allow the emulation of actual biological behavior. Entelos has used this approach to develop its proprietary PhysioLab technology, which allows researchers to model dynamic, large-scale human disease processes. PhysioLab model systems are available (through licenses and R&D partnerships) for asthma, obesity, diabetes, and adipocytes, and they provide integrated expertise in the areas of metabolism, immunology, and respiratory diseases. Predictive biosimulation is still in its infancy, and its applicability to drug development remains very narrow; however, this technology may eventually become useful in lead optimization. In this application, predictive models could be used as decision support tools to focus the direction of compound synthesis and the design of animal experiments. Moreover, it could be employed for the evaluation of lead optimization libraries in a so-called virtual human—a computational model of a human patient—potentially adding information that would otherwise never be obtained in this early stage of drug development. CORPORATE HIGHLIGHTS The companies highlighted in this section comprise a cross-sectional overview of the current business segment focused on medicinal chemistry and lead optimization. Each company represents a particular business focus in this area. Table 3 lists these and other companies with medicinal chemistry or lead optimization technologies, products, and/or services. ArQule: From Strategic Collaboration to Integrated Developer ArQule’s business model has been to provide chemistry and lead optimization services through strategic partnerships. ArQule has integrated extensive lead optimization capabilities into its technology platform and offers clients a wide array

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TABLE 3. Select Companies with Medicinal Chemistry and/or Lead Optimization Technologies, Products, and/or Services Company Argonaut Technologies ArQule

The Automation Partnership

Berger SFC Biotage (Discovery Chemistry division) CEM Chemical Diversity

Combipure deCode Biostructures/deCode Chemistry Discovery Partners International

Web Site

Technology/Products/Services

www.argotech.com

Instruments and reagents for chemical synthesis and purification. www.arqule.com High-throughput parallel synthesis and lead optimization; ADMETa prediction tools; oncology drug discovery. www.automationpartnership.com Automated instrumentation for compound management, high-throughput screening, genomics, cell culture, and sample management. www.bergersfc.com Supercritical fluid chromatography. www.biotage.com Instrumentation, kits, and accessories for microwave chemical synthesis. www.cem.com Microwave laboratory instrumentation. www.chemdiv.com Chemical libraries, medicinal chemistry services, bioscreening, lead optimization, custom synthesis, software. www.combipure.com Chemical purification and library synthesis. www.decode.com Structural biology and chemistry services. www.discoverypartners.com

Entelos Hampton Research

www.entelos.com www.hamptonresearch.com

Mimotopes

www.mimotopes.com

Molecular Dimensions

www.moleculardimensions.com

Pharmacopeia

www.pcop.com

Combinatorial libraries and products; chemical synthesis; software; ADMET profiling; high-throughput purification, high-throughput screening, crystallization and gene-expression-profiling products. Predictive biosimulation software. Tools and products for crystallization screens. Solid-phase synthesis products for high-throughput chemistry and medicinal chemistry; libraries; lead optimization services; peptides. Kits, reagents, and other technologies for biomolecular crystallization studies. Compound libraries; high-throughput screening; combinatorial chemistry, medicinal chemistry; parallel synthesis; ADMET analysis.

CORPORATE HIGHLIGHTS

55

TABLE 3. (continued) Company

Web Site

Pharmidex

www.pharmidex.com

Predix Pharmaceuticals

www.predixpharm.com

Scigene

www.scigene.com

TransForm Pharmaceuticals

www.transformpharma.com

Technology/Products/Services Computational and tissue-based screening methods to predict and assess blood–brain barrier penetration. Computational tools for GPCRb and ion-channel three-dimensional structure prediction; synthetic chemistry; GPCR-based drug discovery and optimization. High-throughput chemical synthesis; ovens; incubators. High-throughput crystallization; formulation design, discovery of alternative salts, solvates, and polymorphic forms and/or formulations.

a ADMET, absorption, distribution, metabolism, excretion, and toxicity. b GPCR, G-protein-coupled receptor.

of drug discovery services. Its Automated Molecular Assembly Plant (AMAP) is the foundation of its high-throughput parallel synthesis and lead optimization program. Through its Parallel Integrated Lead Optimization Technologies (PILOT) system, ArQule uses computational tools and ADMET prediction methods to optimize leads. The company employs its Directed Array program to synthesize analogue compounds based on “hits” from partner companies’ internally developed compounds. In January 2001, the chemistry company ArQule (Woburn, Massachusetts) acquired the computational ADMET modeling company Camitro (Menlo Park, California). This acquisition enabled ArQule to add predictive ADMET capabilities. More recently, ArQule repositioned itself as a biotechnology company developing small-molecule drugs for cancer and inßammation. This strategy followed the 2003 acquisition of cell-cycle and cancer specialist Cyclis Pharmaceuticals. This acquisition gave ArQule Cyclis’s Activated Checkpoint Therapy platform technology for developing oncology drugs as well as its lead oncology compound, CO-501. Chemical Diversity: Focus on Synthesis and Libraries Chemical Diversity’s (San Diego, California) core expertise is the synthesis of drug-like molecules during lead optimization. The design and synthesis of small optimization libraries (50–3000 compounds) and hit-to-lead optimization libraries for QSAR analysis and identiÞcation are central to the company’s custom chemistry business. Chemical Diversity also offers structure-based design, bioassays and ADMET assays, and computational and medicinal chemistry services. It can

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also provide larger-scale compound synthesis for preclinical and early clinical studies. Pharmidex: Lead Optimization for the Brain Pharmidex (London, United Kingdom) is a highly specialized contract research organization that helps drug discovery and development companies select and optimize lead compounds for central nervous system drug development with a particular emphasis on blood–brain barrier penetration. Clarity is Pharmidex’s computational platform technology that rates a compound’s ability to cross into the brain. Integrity, the company’s integrated package of screening methods to assess brain penetration in vitro (Penetrability) and in vivo (NeuroPK), therapeutic efÞcacy, and side-effect liability, is suited for assessing compounds during lead optimization. Predix Pharmaceuticals: Computational Medicinal Chemistry Predix Pharmaceuticals (formerly BioInformation Technologies, Woburn, Massachusetts, and Ramat Gan, Israel) has algorithms for predicting the threedimensional structure of GPCRs and ion channels and uses its virtual screening technology to discover, develop, and optimize compounds that target these molecules. Predix combines laboratory-based medicinal chemistry with structurebased computational approaches to synthesize and optimize lead compounds. In February 2004, the company announced that it had initiated Phase I clinical trials for its drug candidate PRX-00023, a compound discovered and optimized using the company’s computational GPCR modeling. PRX-00023 is a dual-action serotonin 1A (5HT1A ) receptor agonist/sigma-1 antagonist intended to treat anxiety, attention-deÞcit/hyperactivity disorder (ADHD), and other neuropsychiatric disorders. In August 2003, Predix merged with Physiome Sciences, a predictive biosimulation company. Predix uses Physiome technology to model a compound’s effect on the cardiac action potential and to prioritize compounds for further development. TransForm Pharmaceuticals: Formulation Design and Improvement TransForm Pharmaceuticals (Lexington, Massachusetts) offers services designed to enhance the bioavailability and improve the performance of its client’s compounds. The company is able to discover alternative salts, solvates, and polymorphic forms and/or formulations. TransForm specializes in high-throughput crystallization studies and formulation design. It uses its automated CrystalMax technology to perform up to 10,000 crystallization experiments, in parallel, within two weeks. This capability enables researchers to determine optimal or alternate solid forms of the active pharmaceutical ingredient. TransForm also has the capacity to rapidly proÞle the solubility of optimization compounds in parallel and in multiple solvents or vehicles. These capabilities help medicinal chemists to more systematically integrate considerations of developability into the lead discovery process.

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OUTLOOK Medicinal chemistry has entered a state of transition that has already extended the discipline’s reach far beyond the conÞnes of the traditional chemical laboratory. The time-honored practice of handcrafted, “one-at-a-time” lead optimization is being replaced with an industrial approach that bears the hallmarks of modern, high-throughput drug discovery. Integral components include designed compound libraries, high-throughput parallelization technology, sophisticated databases, and molecular modeling. The initial steps of this transition were largely defensive in nature (i.e., they were taken mainly in an attempt to cope with the deluge of primary leads that were being delivered by combinatorial chemistry and highthroughput screening technologies). Recently, an upstream integration has taken place that considers the requirements of medicinal chemistry at the early-stage discovery process. This approach has already led to the adoption of focused compound library design, which is replacing purely technology-driven, brute-force screening. As high-throughput crystallization studies generate more structural information about more targets and receptor–ligand interactions, we anticipate an increase in the adoption and use of computational modeling techniques—such as receptor–ligand docking—for both library design and lead optimization purposes. We are now witnessing a downstream integration of medicinal chemistry in the discovery and development process, in which chemists make highly targeted attempts to create drug candidates that have a signiÞcantly improved prospect of reaching the market and staying there. Accomplishing this objective will require signiÞcant improvements in the predictive power of ADMET technologies. By recruiting high-power computational methods, employing massive parallelization in its laboratory processes, and linking both with proteomics technologies, medicinal chemistry could soon assume the role of a truly pivotal hub in the entire drug development process. By the end of this decade, advances in medicinal chemistry that enable drug developers to bring only the most promising candidates into development could ultimately help shorten preclinical development times and reduce the number of late-stage failures. Advanced clinical trials could be much more tightly focused and far more predictable in their outcome. These changes would help reduce the overall length, cost, and risk of drug development for pharmaceutical and biotechnology companies. Regulatory processes, which are intrinsically slow in accommodating change, might prove to be more signiÞcant impediments in this respect than technological issues.

REFERENCES Blackwell HE. Out of the oil bath and into the oven–microwave-assisted combinatorial chemistry heats up. Organic and Biomolecular Chemistry. 2003;1:1251–1255. Vaino AR, Janda KD. Euclidean-shape-encoded combinatorial chemical libraries. Proceedings of the National Academy of Sciences. 2000;97(14):7692–7696.

Bio-Defense Opportunities in the Biotechnology and Pharmaceutical Industries

SUMMARY Biodefense is an emerging industry that offers new opportunities for both existing and new drug companies, as well as for biotechnology companies with technologies for drug discovery and pathogen detection. Incentives to enter the Þeld include government funding of R&D, accelerated regulatory approval, and reduced time-to-market. Still, companies need to weigh those advantages against several risk factors, such as liability, a limited market, and the potential for overregulation and patent challenge. In this article, we discuss the threat of biowarfare, the various technologies used in biodefense, and the commercial opportunities and challenges inherent in the Þeld, and we brießy proÞle vaccine and therapeutic drug companies that have already entered the biodefense business. BUSINESS IMPLICATIONS •

Soon after the dissemination of anthrax throughout the U.S. Postal Service in 2001, biodefense funding in the United States increased dramatically. The National Institute of Allergy and Infectious Diseases (NIAID) refocused its efforts to emphasize biodefense-based research programs and signiÞcantly increased its biodefense budget. In May 2004, the U.S. Senate passed Project Bioshield, which will provide $5.6 billion in biodefense funding to the commercial sector. This new focus on biodefense creates many opportunities for companies developing vaccines and therapeutic drugs.

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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INTRODUCTION

•

•

•

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The threat posed by biological warfare agents to large population centers is signiÞcant. Pathogens that cause diseases such as anthrax, smallpox, plague, tularemia, botulism, and hemorrhagic fevers are potential bioterror and biowarfare agents. New vaccines and drugs will be needed to protect against and effectively treat these diseases. Facing the need for new stockpiles of smallpox and anthrax vaccines, various governmental agencies are offering large development and supply contracts. Many companies, including some that are specializing in biodefense, are taking advantage of these contracts as well as other types of government funding. Among the leading biotechnology companies developing new biodefense vaccines are Acambis, Bavarian Nordic, BioPort, DynPort Vaccine, and VaxGen. Incentives such as government funding of R&D, accelerated regulatory approval, and reduced time-to-market will encourage many biotechnology companies to explore the biodefense arena. Still, these companies will need to weigh those advantages against various risk factors, including complicated routes to obtaining government contracts, unpredictable markets, liability, price controls, and the loss of patent protection for products that become important to national security.

INTRODUCTION One month after the terrorist attacks of September 11, 2001, the deliberate dissemination of anthrax spores through the U.S. Postal Service resulted in 22 documented or suspected cases of anthrax, 5 of which proved fatal. This attack highlighted the vulnerability of the United States to bioterrorism and biowarfare. Soon after the anthrax incident, biodefense funding in the United States increased dramatically, and the National Institute of Allergy and Infectious Diseases (NIAID) refocused its efforts to emphasize biodefense-based research programs. In September 2003, NIAID granted funds totaling $350 million (over Þve years) to universities and institutions to establish eight Regional Centers of Excellence for Biodefense and Emerging Infectious Disease Research. That same year, the agency’s biodefense budget increased to $1.16 billion, from $20.7 million in 2000. In his 2003 State of the Union Address, President George W. Bush announced Project Bioshield, a national initiative to enable the development of drugs and vaccines to combat biological and chemical weapons. Project Bioshield legislation, which was passed by the House of Representatives in July 2003 and by the Senate in May 2004, will provide $5.6 billion over ten years for the commercial development of vaccines and therapies for biowarfare agents. Under the project, the government will guarantee drug purchases from companies developing therapies or vaccines against speciÞc biowarfare pathogens. This new focus on biodefense creates new opportunities for vaccine and therapeutic drug companies, as well as for biotechnology companies with technologies for drug discovery and pathogen detection. Proposed legislation to encourage the commercial development of biodefense products and the U.S. government’s

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commitment to provide major funding for biodefense programs are stimulating signiÞcant research on hitherto little-studied pathogens. In this article, we explore the commercial opportunities and challenges in the emerging biodefense industry for biotechnology and pharmaceutical companies. THE THREAT OF BIOLOGICAL WARFARE AGENTS The threat posed by biological warfare agents to large population centers is signiÞcant. As early as 1970, a World Health Organization (WHO) committee predicted that 50 kilograms of virulent tularemia germs dispersed over a metropolitan area of 5 million people would result in 250,000 incapacitating casualties and 19,000 deaths and that 50 kilograms of plague organisms released would result in 150,000 incapacitating casualties and 36,000 deaths. In 1993, the U.S. OfÞce of Technology Assessment (OTA) estimated that 100 kilograms of anthrax spores could result in 130,000 to 3 million deaths. Particular threats are posed by illnesses such as tularemia, pneumonic plague, botulism, and viral hemorrhagic fevers, for which no vaccines and few therapies are available. Smallpox, which has a 30% case fatality, is another major concern. Because the United States discontinued smallpox vaccines in 1972, the majority of the U.S. population may be susceptible to this virus. NIAID and the Centers for Disease Control and Prevention (CDC) have classiÞed potential bioterror and biowarfare pathogens into three categories—A, B, and C—based on the ease of dissemination and person-to-person transmission; mortality and morbidity rates; the severity of consequence (including public panic); and the level of consequence management (efforts to manage a wide variety of short- and long-term effects of the attack) required to address an attack. Category A pathogens, which are considered the most medically severe and have the highest destructive potential, include those that cause anthrax, botulism, plague, smallpox, tularemia, and viral hemorrhagic fevers. Category B pathogens, which are relatively easy to disseminate and cause moderate to low mortality, include those that cause Q fever, brucellosis, and various toxicities (e.g., from ricin toxin, Clostridium perfringens epsilon toxin, staphylococcal enterotoxin B). Category C pathogens include those that cause emerging infectious diseases, including Nipah viruses, hantaviruses, tickborne hemorrhagic fever viruses, and yellow fever virus, and those that cause multidrug-resistant tuberculosis. These pathogens are at risk for being engineered for mass dissemination because they are readily available, relatively easy to produce and disseminate, and have a potential for high morbidity and mortality. An attack with Category A pathogens would most likely occur via aerosol dispersal (dispersal through water supplies or insect or animal reservoirs is possible but less efÞcient). Aerosol dispersal results in inhalational versions of disease, which are typically more severe and present with less familiar symptoms than versions caused by natural routes of infection by the same organism. For example, pneumonic plague caused by inhaling Yersinia pestis-containing aerosol is more

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severe than the bubonic plague that is acquired from infected ßeas. Inhalational anthrax is signiÞcantly worse than cutaneous or gastrointestinal anthrax, which is caused by touching or eating contaminated animal products. Two recent cases predict the destructive power of biowarfare agents that cause inhalational anthrax. In 2001, of the 11 cases of inhalational anthrax in the United States from contaminated letters, almost half resulted in death. Earlier, in 1979, in the Russian town of Sverdlovsk, 79 local inhabitants acquired inhalational anthrax after a nearby Soviet bioweapons plant accidentally released weaponized anthrax spores into the air. Sixty-eight of the victims reportedly died (Inglesby TV, 2002). Factors that distinguish biowarfare and bioterror agents from other weapons of mass destruction include the pathogens’ potency and their ability to escape early detection, the requirements of Þrst responders to bioterror attacks, and the type of consequence management needed to deal with an attack. Only very small quantities of biological agents are required for mass destruction: 100 kilograms of anthrax spores that could kill 3 million people could Þt inside a briefcase. Biological agents are also invisible, odorless, colorless, tasteless, and otherwise difÞcult to detect. Unlike attacks with chemical or explosive weapons, for which the source of an attack is immediately apparent and which result in instantaneous mortality around an epicenter (which can then be managed by rescue workers, ÞreÞghters, police, and other Þrst responders), bioterrorist acts may not be detected for days and may be initially confused with other diseases (such as ßu). First responders to bioterror include emergency units and health care personnel in hospitals who have to deal with the effects of speciÞc diseases. Consequence management includes input from agencies including the Federal Bureau of Investigation (FBI) and the CDC. Part of this management need includes the control of public panic, which can result in people ßeeing quarantined areas and spreading the disease further. WEAPONIZATION OF BIOLOGICAL AGENTS Simply having access to dangerous pathogens does not enable the creation of biological weapons. Pathogens (or the toxins they generate) must Þrst be “weaponized” to be effective as biowarfare agents. Weaponized agents are different from naturally occurring pathogens. These differences are important for drug developers to understand so that they can develop effective therapeutics to defend against pathogens used in biowarfare. Production requires technical know-how for the selection, handling, engineering, containment, and large-scale production of biological pathogens. Delivery involves not just technologies for delivery via air or water supply but also technologies for the concentration, puriÞcation, and stabilization of biological agents for packaging and storage. Dispersal involves apparatus for scattering pathogens and toxins as well as technologies that improve the survivability of biological agents in media (e.g., aerosol, water), explosive devices, and the environment (e.g., sunlight, air, meteorological conditions). The sophistication required for the production, delivery, and dispersal of biological agents limits the number of countries and organizations that can participate in biological warfare or bioterrorism.

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BIODEFENSE TECHNOLOGIES Companies are developing vaccines, small-molecule drugs, protein therapeutics, and detection methodologies to aid in combating biological weapons. Virtually every drug discovery technology at the disposal of the biotechnology and pharmaceutical industry (e.g., high-throughput target identiÞcation technologies, high-throughput screening, informatics) can be applied to biodefense. In this section, we discuss current technologies for and approaches to biodefense-related drug discovery. Target Identification Target identiÞcation refers to the discovery of molecules against which drugs and vaccines can exert a therapeutic effect. This discovery process requires signiÞcant basic research directed toward understanding pathogen life cycles, host immune responses to pathogens (including innate immunity), and the molecular mechanisms by which pathogens create disease. Despite tremendous advances in molecular and cell biology, surprisingly little is known about many biological warfare pathogens. This dearth of knowledge is due mainly to the previous perception that the threat of biological warfare was relatively low and to the lack of funding available for studying potential biowarfare agents. Moreover, the difÞculty of working with many of these biological agents has limited the number of people who could study the most dangerous and contagious pathogens. Only four U.S. facilities have operational biosafety level-4 (BSL-4) containment laboratories necessary to work with Category A organisms: the CDC in Atlanta, the U.S. Army in Fort Detrick, Maryland, the Southwest Foundation for Biomedical Research in San Antonio, and the University of Texas at Galveston. Recently, Boston University and the University of Texas won $128 million and $110 million NIAID awards, respectively, to construct National Biocontainment Laboratories, which are new buildings for BSL-4 facilities for the study of lethal pathogens and biological toxins. Tremendous opportunities are available in biodefense for companies with functional genomics and proteomics technologies. Because astonishingly little is known about many potential biowarfare pathogens, companies that specialize in microorganisms and anti-infectives have an advantage in this area. For example, the complete nucleotide sequence of Francisella tularensis, the organism that causes tularemia, has yet to be determined. Further DNA and RNA sequencing studies will be required to enable the comparison of strains and mutations, and to understand the contribution of bacterial plasmids and their encoded proteins in disease etiology. Structural analyses of pathogen proteins are also needed to perform structure–activity relationship analysis of drug–target interaction. Animal Models A general lack of postmortem studies in humans who died from bioweapons contributes to the unsatisfactory understanding of the clinical manifestations and pathology of inhalational diseases caused by biowarfare agents. Testing

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biodefense therapeutics is also complicated by the fact that researchers cannot, for ethical reasons, determine the therapeutic efÞcacy of compounds directly on humans in Phase III clinical trials. To address this problem, the FDA announced in May 2002 that it would rely on proof of efÞcacy in animals—instead of humans—to approve biodefense therapeutics in cases where Phase III testing in humans is not feasible or ethical. These circumstances place greater importance on reliable animal models to predict the efÞcacy and safety of potential drug candidates for biodefense. Liability issues for drugs that have not been tested in large-scale Phase III clinical trials also become signiÞcantly more consequential. New and better animal models that replicate the physiological sequence from inhalation and natural routes of infection from biological weapons are therefore required. Companies that specialize in creating new animal models, including transgenic animals, may Þnd opportunities in the biodefense sector. An example of a new animal model is the bubonic plague ßea-to-mouse transmission model that NIAID recently developed to test new vaccines (Jarrett CO, 2004). This model, which transmits plague organisms (Yersinia pestis) to mice through ßeabites, mimics routes of bubonic plague infection more naturally than do models that rely on syringe-injection of the pathogen. This model is superior for testing bubonic plague vaccines against naturally acquired plague because plague organisms express a different set of antigenic proteins when grown in ßeas instead of cell culture (and therefore exhibit different immune responses). Hence, by replicating natural routes of infection, researchers may be able to Þnd vaccines that are more likely to be effective against ßea-transmitted bubonic plague (a potential bioterrorism method) in humans. Vaccines The sudden urgency to provide vaccines against biological weapons has encouraged the resurrection of discontinued vaccines that were prepared by outdated and potentially unsafe methodologies. Moreover, vaccines that were developed for natural infections often do not work for inhalational versions of disease caused by weaponized pathogens and toxins. For example, discontinued plague vaccines that were effective against bubonic plague do not work on the pneumonic plague caused by plague organisms that have been weaponized into aerosols (Inglesby TV, 2000). Current versions of vaccines for organisms such as smallpox are also still based on methodologies developed in the early 1900s. During the smallpox eradication campaign of the last century, four basic vaccinia virus strains were used as smallpox vaccines: the Lister strain (in Japan and other countries), the New York City Board of Health (NYCBH) strain (in the Americas), the EM-63 strain (in the Soviet Union), and the Patwadanger strain (in India). The current frozen stocks of smallpox vaccine (Dryvax), which is based on the NYCBH strain and was manufactured by Wyeth Laboratories in the early 1980s, are made from crude extracts harvested from the pustules on the skin of infected calves. Although

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15 million aging doses (which in principle could be expanded into 75 million doses) of frozen Dryvax exist, the CDC is reserving vaccinations for voluntary inoculation in health care providers and Þrst responders. The agency fears that mass inoculation of the U.S. population with the Dryvax strain could result in hundreds of deaths and thousands of severe side effects, including permanent brain damage. The Dryvax strain is especially dangerous and contraindicated for the millions of people with immunodeÞciencies, heart conditions, eczema, and atopic dermatitis. Clearly, more modern vaccines based on recombinant DNA, cell culture, or safer strains of pathogens are needed. PuriÞed immunizing virus grown in cell culture can be used to create cleaner vaccines free of contaminating animal products. Still, cell-culture-derived viral or bacterial vaccines created using live, attenuated organisms or inactivated viruses pose signiÞcant side effects and safety issues, including death. These safety concerns underlie the CDC’s decision not to mass-inoculate the U.S. population against smallpox. Hence, some companies are basing their new biodefense vaccines on milder strains in the hope they will induce the same immune response without undesirable side effects. With the support of the U.S. Army and the National Institutes of Health (NIH), several companies are attempting to develop even safer recombinant subunit vaccines based on bacterial or viral proteins made from cloned genes. The challenge is that many bacterial and viral proteins are not immunogenic when used alone. For example, recombinant hepatitis B vaccines based on the viral surface antigen worked only after researchers discovered that the viral proteins must Þrst be isolated as aggregates of 22 nm particles and that the proteins were ineffective as single monomers. Determining which protein or combinations of proteins will evoke an immune response can be a difÞcult task. Therapeutic Drugs New drugs are needed to Þght the diseases that could be caused by biowarfare. Despite the availability and effectiveness of standard antibiotics to Þght many pathogens used as bioweapons, antibiotics are often ineffective by the time symptoms present or have been diagnosed. For example, by the time inhalational anthrax presents, anthrax bacilli are already rapidly germinating and releasing toxins (against which antibiotics are not effective). Moreover, the rise in antibiotic-resistant strains and the ease by which bacteria can be made antibioticresistant through genetic engineering (simply by routine transfection of plasmids encoding antibiotic-resistance genes into bacteria) call for new therapies against novel drug targets. The goal in antibiotic drug development is to Þnd a broad-spectrum orally bioavailable antibacterial or antiviral drug that works through a mechanism that is difÞcult to evade through mutation (which leads to drug resistance). Pharmaceutical companies favor small-molecule drugs over protein therapeutics because the former can be stable at room temperature, orally bioavailable (protein therapeutics must be injected), and chemically synthesized relatively easily. The

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military also favors broad-spectrum, small-molecule drugs because soldiers in the Þeld (who are potentially vulnerable to bioweapons) cannot carry a portfolio of difÞcult-to-administer drugs dedicated to different diseases. Still, protein therapeutics such as monoclonal antibodies and cytokines have potential; they can combat biological weapons by inactivating toxins, by blocking the action of host or pathogen molecules that mediate pathogen growth or toxicity, or by modulating the immune system. Other Technologies Other technologies that contribute to biodefense include RNA interference, aptamers, drug delivery technologies, and adjuvant technologies. Many companies are also pursuing detection methodologies for biowarfare pathogens (see Table 1). FACTORS THAT INFLUENCE THE BUSINESS OF BIODEFENSE Companies attempting to transform biodefense technologies into a business must overcome several signiÞcant challenges. Drug and vaccine development is an expensive and lengthy process that requires 10–15 years and hundreds of millions of dollars. Because biotechnology companies will not see sales revenue from the products they develop for many years, these Þrms are dependent on outside sources of Þnancing, including government grants, venture capital, corporate partnership revenue, and public equity (see Figure 1). The U.S. government has taken steps to provide more funding and to ease the regulatory path to promote commercial biodefense efforts. In this section, we discuss some of the catalysts and hurdles that are inßuencing the business of biodefense. Government Biodefense Financing Many biotechnology companies are exploring biodefense because of the tremendous amount of funding that has suddenly become available. U.S. government appropriations for biodefense funding from NIAID have increased 80-fold in only four years, from $20.7 million in 2000 (1% of NIAID’s total budget) to $1.6 billion in 2004 (more than one-third of NIAID’s total budget) (see Figure 2). The government is now the largest source of funding for academic institutions and biotechnology companies that want to conduct biodefense basic research or develop novel vaccines and therapeutics. Prior to 2001, the Defense Advanced Research Project Agency (DARPA) was the largest principal source of biodefense funding. In 2003, the agency devoted approximately $160 million of its total $2.65 billion budget to biodefense, and in 2004, its biodefense funding allocation totaled approximately $150 million. DARPA, which favors funding of “blue sky,” long-term efforts, has funded projects such as the discovery of modiÞed stem cells that can detect and produce therapeutic responses against pathogens, pathogen communication disruption, toxin-binding decoys, immunomodulation, broad-spectrum small-molecule and

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TABLE 1. Select Companies Active in Biodefense Biodefense-Related Disease/Pathogen/Technology

Company

Web Site

Vaccines Acambis

www.acambis.com

Avanir Bavarian Nordic

www.avanir.com www.bavariannordic.com

Berna Biotech BioPort BioSante Chiron (PowderJect) Crucell Dor Biopharma Dynavax DynPort Vaccine

www.bernabiotech.com www.bioport.com www.biosantepharma.com www.chiron.com

GeneMax Microscience Nanotherapeutics Panacea Biotec VaxGen Vical

www.genemax.com www.microscience.com www.nanotherapeutics.com www.panacea-biotec.com www.vaxgen.com www.vical.com

Ebola virus, West Nile virus Ricin toxin, botulinum toxin Anthrax, plague Smallpox, anthrax, plague, tularemia, botulinum toxin Smallpox Anthrax Anthrax, ricin Anthrax Smallpox, anthrax Ebola virus, West Nile virus

www.anacor.com www.biophage.com www.biopolymer.com

Anthrax Anthrax Anthrax

www.biota.com.au www.cengent.com

Smallpox Anthrax

www.chimerix-inc.com www.genelabs.com

Smallpox Broad-spectrum antivirals

www.gilead.com www.hemispherx.net

Anthrax Broad-spectrum antivirals, smallpox, Ebola virus Antibacterials Smallpox (USAMRIIDa using Kemin libraries) Anthrax Smallpox Smallpox Smallpox

Small-Molecule Drugs Anacor Biophage Biopolymer Engineering Biota Holdings Cengent Therapeutics Chimerix GeneLabs Technologies Gilead Sciences Hemispherx

www.crucell.com www.dorbiopharma.com www.dynavax.com www.dynport.com

Ibis Kemin Pharma

www.ibisrna.com www.kemin.com

PolyMedix Siga Technologies TransTech ViroPharma

www.polymedix.com www.siga.com www.ttpharma.com www.viropharma.com

Protein Therapeutics Abgenix

www.abgenix.com

Smallpox, West Nile virus, dengue fever, yellow fever, Japanese encephalitis virus, Clostridium difficile Anthrax Smallpox, dengue fever, Japanese encephalitis virus Smallpox Anthrax Anthrax, plague, ricin toxin Yellow fever virus

Ebola virus (USAMRIID using Abgenix technology)

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TABLE 1. (continued) Company Alexion Antibody Technologies Cangene EluSys Therapeutics Human Genome Sciences Medarex Nuvelo PharmAthene Serologicals Detection Systems/ Devices Aclara Biosciences Bioveris (through Igen acquisition) Cepheid EraGen

Web Site

Biodefense-Related Disease/Pathogen/Technology

www.alxnsd.com

Anthrax

www.cangene.com www.elusys.com

Smallpox, anthrax, botulism Anthrax

www.hgsi.com

Anthrax

www.medarex.com www.nuvelo.com www.pharmathene.com www.serologicals.com

Anthrax Ebola virus Anthrax Smallpox

www.aclara.com www.bioveris.com

e-Tag assay system Test kits for botulinum and ricin toxins, anthrax, and Staphylococcal enterotoxin B www.cepheid.com GeneXpert system for anthrax www.eragen.com GeneCode detection system for smallpox and anthrax Isis www.isispharm.com Triangulation Identification Genetic Evaluation Pharmaceuticals of Risks (TIGER) biosensor technology MedMira www.medmira.com Smallpox Nanogen www.nanogen.com NanoChip electronic microarray Nanosphere www.nanosphere-inc.com Nanoparticle probes, assays, instruments Northrup Grumman www.northgrum.com Automated Bio-Agent Detection System Orchid BioSciences www.orchid.com SNP-based detection technology Response www.responsebio.com Detection systems for smallpox, anthrax, and Biomedical botulinum and ricin toxins Tetracore www.tetracore.com Test kits for anthrax, plague, ricin and botulinum toxins, Staphylococcal enterotoxin B, tularemia, and Brucella a USAMRIID, U.S. Army Medical Research Institute for Infectious Diseases.

peptide therapy, gene shufßing to develop superantigens, gene therapy, and immune stimulation with edible plants. Other sources of biodefense government funding include the CDC, which is responsible for contracting national stockpiles of vaccines, antibiotics, and other therapeutics for the civilian population; the U.S. Army Medical Research Institute of Infectious Diseases (USAMRIID); the National Science Foundation (NSF); and the U.S. Department of Energy (DoE). As mentioned previously, the U.S. Senate passed Project Bioshield legislation in 2004. The legislation will provide $5.6 billion over ten years for the commercial development of vaccines and therapies for biowarfare agents. It will aid the stockpiling of vaccines and therapeutics against biowarfare pathogens and provide other incentives for companies to develop biodefense products. Guaranteed

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FIGURE 1. Drug discovery value chain and sources of revenue

government contracts to purchase vaccines from commercial suppliers and other funding will provide a signiÞcant boost to the emerging biodefense industry. Accelerated Regulatory Approval for Biodefense Products In 2002, the FDA eased regulatory requirements for the approval of new drugs that treat the effects of exposure to biological, chemical, radiological, and nuclear

FIGURE 2. National institute of allergy and infectious disease biodefense funding

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agents. Under the FDA’s Animal EfÞcacy Rule, the agency can approve biodefense drugs based on their efÞcacy in animal models alone in cases in which human trials for efÞcacy are either not feasible or ethical. By allowing companies to avoid conducting human Phase III clinical trials for certain biodefense-related products, the Animal EfÞcacy Rule potentially saves the companies tens of millions of dollars. Also in 2002, the Public Health Security and Bioterrorism Preparedness and Response Act authorized the FDA to confer fast-track status on and permit priority review of biodefense products. These changes provide tremendous incentives for companies to pursue biodefense therapeutics because they reduce both the cost and time required to develop a therapeutic drug or vaccine. Market Challenge The market for biowarfare and bioterrorism-related diseases is extremely small. Both the incidence and prevalence of diseases such as Ebola virus hemorrhagic fever and anthrax are very low. Unless a company can develop an anti-infective that can be applied to many diseases beyond biodefense-related pathogens, concentrating on biodefense—a market characterized by high demand only immediately after a bioterrorism incident—is an extremely risky business proposition. The small biodefense market affects the ability of biodefense companies to raise working capital through venture capital and corporate partnerships. Venture capitalists and pharmaceutical companies are not interested in investing in or partnering with a biotechnology company unless they can realize signiÞcant returns on investment. This disincentive creates Þnancial bottlenecks within the biodefense development chain (Figure 3). One remedy to the Þnancial challenge is a government guarantee to prepurchase stockpiles of vaccines or therapeutics from biotechnology companies, even if they are still in development. In November 2001, the CDC awarded Acambis a contract to purchase 155 million doses of smallpox vaccine for $428 million. This contract built on the company’s previous contract with the CDC, originally awarded in September 2000 and updated in October 2001, for the production of

FIGURE 3. Challenges and catalysts in the biodefense drug discovery chain

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54 million doses for $343 million. Acambis, which had been a struggling biotechnology company, saw its stock skyrocket after the 2001 contract was announced. Guarantees by the government to prepurchase vaccines and therapeutics from companies developing new and hitherto untested products would reduce risk and could provide an incentive for venture capital investments in this arena. Limited Pool of Funding Sources and Customers Companies focusing on biodefense face the prospect of depending on a small number of government funding sources for Þnancing and supply contracts. The biodefense industry does not enjoy the large markets, consisting of thousands of hospitals and potentially millions of patients, that comprise the traditional pharmaceutical segments. Competing with multiple biotechnology and pharmaceutical companies to win business with a handful of government contractors and agencies is highly risky. Also, although the amount of biodefense funding has signiÞcantly increased, there is no guarantee these levels will be maintained or increased. Liability Challenge Liability is a critical issue that may deter many biotechnology and pharmaceutical companies from entering biodefense. New biodefense vaccines and therapeutics will be based on efÞcacy in animal models with no assurance of efÞcacy or acceptable side effects in humans. Unless governments clearly address the issue of liability, investors and large pharmaceutical companies will not pursue the development of new biodefense products, and only a few biotechnology companies will focus on biodefense. A step in the right direction is new legislation that enables the Department of Homeland Security to designate biodefense products as antiterrorism technologies and shifts liability to the federal government. In addition, legislation, such as Senate bill S. 666 (the Biological, Chemical, and Radiological Weapons Countermeasures Research Act), has been introduced that includes liability protection for companies developing biodefense therapeutics. Overregulation and Patent Challenge The threat of government control of biodefense products, simply because they are perceived by politicians to be in the domain of national defense, is discouraging the active participation of large pharmaceutical companies and investors—parties whose involvement is vital to drive the creation of new biodefense products. The health care industry is already one of the most regulated industries, characterized by complex rules, codes, and procedures designed to ensure drug safety and efÞcacy. The perception of further or undue government regulation could inhibit the innovation required to create new vaccines and therapeutics. Another threat posed by the government is in the area of patent protection. Without patents, the pharmaceutical industry could not exist. The protection afforded by patents enables pharmaceutical companies to pay for the overwhelming cost of R&D necessary to create high-risk new products. However, in the month following the anthrax scare in October 2001, the guarantee

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of patent protection came under serious threat. During negotiations related to increasing the supply of the antibiotic ciproßoxacin (Bayer’s Cipro), an approved drug to treat inhalational anthrax, members of both the U.S. Senate and Canadian government proposed bypassing Bayer’s patent. Although these proposals were not carried out, they raised legitimate fears among pharmaceutical and biotech companies. Bayer agreed to provide large volumes of Cipro to the U.S. government at a signiÞcantly reduced price. The company’s revenue from Cipro fell 37% between 2001 and 2002. Scientific Freedom and Innovation Basic scientiÞc research and the free exchange of information through scientiÞc publications and other communications form the foundation from which all new drugs are created. The U.S. Patriot Act and the Public Health Security and Bioterrorism Preparedness and Response Act, which were hurried into law in 2001 and 2002, respectively, have created unanticipated challenges in basic research that threaten to slow the development of biodefense products (see Figure 3). Standard scientiÞc practices necessary to conduct basic research and drug discovery, including the publishing and sharing of scientiÞc information—such as DNA sequences of pathogens—have become regulated, monitored, and, in a few cases, prosecuted. In one extreme and alarming case, a U.S. attorney charged, under the Patriot Act, a 26-year-old graduate student from the University of Connecticut with possessing a potential bioterrorism pathogen. While cleaning out an old freezer, the student had moved and refrozen forgotten vials from the 1960s that contained anthrax-contaminated cow tissue. The threat of such criminal prosecution under the Patriot Act could lead universities to avoid conducting needed research on diseases such as anthrax. Clearly, progress in developing new drugs and vaccines against bioterror agents will require a more rational balance between security and scientiÞc freedom. CORPORATE HIGHLIGHTS Vaccine Companies Although very few biotechnology companies specialize in biodefense alone, vaccine companies are an apparent exception. The current need for new stockpiles of smallpox and anthrax vaccines has created large development and supply contracts by the governments of many countries. This need has enabled several vaccine companies to speciÞcally focus on biodefense. In many cases, biodefense has been the salvation for vaccine-based biotechnology companies. Acambis. Acambis (Cambridge, United Kingdom) develops and manufactures vaccines for smallpox, West Nile virus, travel and endemic diseases (e.g., yellow fever, dengue fever, Japanese encephalitis), and Clostridium difÞcile (a toxinproducing intestinal bacteria). Under a $771 million, twenty-year contract (which consolidates two previous contracts) with the CDC, Acambis will manufacture

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and maintain 209 million doses of ACAM2000 smallpox vaccine for civilian use. As mentioned previously, the company’s Þrst contract (originally awarded in September 2000 and updated in October 2001) provided $343 million for the production of 54 million doses of ACAM1000 vaccine. The company reports that it will be replacing this vaccine with its newer ACAM2000 vaccine. Under the second contract (November 2001), Acambis and its partner, Baxter, will receive $428 million for 155 million doses of ACAM2000. Acambis reports that it has completed delivery of 155 million doses to the CDC. Acambis’s new smallpox vaccine is an improvement over previous vaccines. The original smallpox vaccines were based on a live poxvirus (vaccinia virus) that cross-protects humans against smallpox. PuriÞed from crude extracts from the lymph of calves infected with vaccinia, the original Dryvax vaccine (which was manufactured by Wyeth Laboratories) is today considered unacceptable for mass inoculation because of the threat of contamination by infectious agents from animal tissue. Acambis’s smallpox vaccine, ACAM2000, which uses the same vaccinia virus strain as the Dryvax vaccine, is safer because it is derived from cell culture instead of calf tissue. ACAM2000 is a cloned viral isolate derived from Dryvax that had been adapted to grow on Vero cells (cultured mammalian cells) and screened for side effects in rabbits and mice. Acambis is conducting Phase II clinical trials for ACAM1000. In the meantime, the company is manufacturing ACAM2000 as an investigational new drug (IND). In April 2004, however, it announced that it had suspended recruitment for its Phase III clinical trials for ACAM2000 after three suspected cases of myopericarditis were discovered in both ACAM2000- and Dryvax-inoculated volunteers (Dryvax was the comparator product). Acambis is reviewing its data to discover whether a relationship exists between the myopericarditis and the smallpox vaccines and plans to request guidance from the FDA and the CDC on how to proceed. With its partner Baxter, Acambis is also developing modiÞed vaccinia ankara (MVA), an attenuated form of current vaccinia strains. A vaccine based on this strain would be most appropriate for immunocompromised patients. In March 2003, Acambis won a $9.2 million contract with NIAID to develop the MVA vaccine. Net proÞts of approximately $61 million in 2003 enabled Acambis to acquire Berna Products for $8.4 million. This acquisition gives Acambis the necessary salesforce to market its yellow fever vaccine, Arilvax. The company Þled a biologics license application (BLA) with the FDA for Arilvax in December 2003. Bavarian Nordic. Bavarian Nordic (Copenhagen, Denmark) is developing Imvamune, a third-generation smallpox vaccine based on the MVA strain of vaccinia virus. The company’s core technology, MVA-BN, exploits the MVA virus as a viral vector to deliver vaccine antigens. The MVA strain is a highly attenuated, replication-impaired virus that is thought to be safer, particularly for immunocompromised patients, than the Dryvax strains. In February 2003, NIAID awarded Bavarian Nordic a three-year contract to develop the MVA vaccine. Under the contract, the company must develop a plan to manufacture and deliver

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up to 30 million doses of the vaccine to the U.S. government. NIAID will also support clinical development of the vaccine. Bavarian Nordic has completed a Phase I clinical trial of Imvamune in healthy volunteers. In May 2003, the company announced that it had completed a Phase II dose-Þnding trial and that the vaccine had demonstrated a good safety proÞle at all doses tested and incurred only mild-to-moderate injection-site reactions. Subjects receiving the highest dose demonstrated a rapid immune response, and subjects receiving the lower doses seroconverted after the full vaccination course. The company is planning followup studies. Bavarian Nordic is proÞtable, reporting 2003 revenues of DKK 504.6 million ($76.9 million) and DKK earnings before interest and taxes of DKK 214 million ($32.6 million). In 2003, it acquired a cGMP (current good manufacturing process)-compliant manufacturing facility in Berlin-Buch, Germany, from Schering and a Denmark-based production facility from Orion (Finland). Bavarian Nordic reports that the latter plant can be expanded to produce 120 million doses of smallpox vaccine per year. In 2002, the company secured several contracts with various countries to produce smallpox vaccines, including a $26.4 million contract to produce its Elstree-BN smallpox vaccine and other smallpox vaccine contracts worth $47.5 million. Bavarian Nordic is focusing on using its proprietary vector technology to develop vaccines against smallpox and human immunodeÞciency virus (HIV). The company intends to partner its original programs, including its cytochrome P450 cell therapy program for pancreatic cancer vaccines. In October 2003, Bavarian Nordic entered into a partnership with Pharmexa to develop HER-2 vaccines for breast cancer. It also has partnerships with Chiron and Epimmune. BioPort. BioPort (Lansing, Michigan) is the only licensed U.S. manufacturer of anthrax vaccines. The company makes its vaccine, BioThrax, by collecting Þltered media from cell cultures of avirulent Bacillus anthracis, which contains anthrax protective antigen and other proteins. BioPort was created in 1998 when the State of Michigan privatized the Michigan Biologics Products Institute (MBPI) and transferred ownership of assets to BioPort. The institute had produced anthrax vaccine for the U.S. Department of Defense (DoD); after privatization, BioPort entered into a new DoD contract to supply the vaccine. In 1997, MBPI faced closure of its anthrax vaccine plant by the FDA. The agency cited quality control violations and, in 1998, found additional regulatory deviations that resulted in the quarantine of 11 lots of anthrax vaccine destined for the U.S. Army. BioPort renovated the facilities and obtained FDA approval of its vaccine production plant in late 2001. In December 2003, BioPort signed a $245 million contract with the Pentagon to supply BioThrax anthrax vaccine for up to three years. In June 2003, BioPort completed its acquisition of Antex Biologics (Gaithersburg, Maryland) for $3.4 million. The Antex acquisition expands BioPort’s product portfolio into vaccines to prevent respiratory and enteric infections as well as sexually transmitted diseases. In addition to the vaccine portfolio, the acquisition

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provides BioPort with an opportunity to develop several classes of small molecules as novel antibiotics. DynPort Vaccine. DynPort Vaccine (Frederick, Maryland) is a joint venture between Computer Sciences (formerly DynCorp) and Porton International, a wholly owned subsidiary of the French pharmaceutical company Ipsen. In 1997, DynPort was awarded a $322 million, ten-year contract with the DoD’s Joint Vaccine Acquisition Program (JVAP). Under the contract, DynPort is JVAP’s prime systems contractor to manage the U.S. stockpiles of biodefense vaccines (except for anthrax vaccine, which is managed by BioPort). In addition, DynPort develops or subcontracts the development of new biodefense vaccines for JVAP, including smallpox, next-generation anthrax, plague, Venezuelan equine encephalitis, tularemia, and botulinum toxin vaccines. DynPort is developing a cell-culture-derived smallpox vaccine (CCSV) based on the NYCBH strain used in the Dryvax vaccines. A Phase I clinical trial involving 350 healthy volunteers compared CCSV with Dryvax. The investigators found that CCSV elicited a 100% immune response with 8% fewer side effects than those of Dryvax. DynPort subcontracts with ID Biomedical (Vancouver, British Columbia) to develop subunit vaccines against pneumonic plague. The existing vaccine for plague, which has been discontinued, is not effective against the pneumonic plague that would be expected to result from aerosolized plague organisms used in biowarfare or bioterrorism. As part of an alliance with the U.S. Army Medical Research and Material Command (USAMRMC), ID Biomedical demonstrated that intranasal delivery of plague antigens using its proteosome technology protected mice from aerosolized plague organisms. In January 2003, ID Biomedical received a $6 million contract to clone, express, and cGMP manufacture plague antigens for use in DynPort’s vaccine; in December 2003, DynPort awarded the company a second contract worth $4.5 million to produce a subunit antigen. In March 2003, DynPort initiated Phase I clinical trials to test the safety of an injectable anthrax vaccine based on recombinant B. anthracis protective antigen (rPA). DynPort is also pursuing the development of next-generation anthrax vaccines for inhalational anthrax by subcontracting with two companies. In January 2003, DynPort awarded Avant Immunotherapeutics (Needham, Massachusetts) an $8 million, two-year contract to develop and conduct preclinical testing on an oral combination anthrax/plague vaccine. In April 2004, DynPort awarded a $3 million subcontract to Avant to develop an oral combination vaccine against anthrax and plague. In January 2004, DynPort awarded BioSante Pharmaceuticals (Lincolnshire, Illinois) a $658,000 subcontract to develop a noninjectable anthrax vaccine using BioSante’s calcium phosphate nanoparticles as an adjuvant. In April 2004, DynPort was awarded a $570,527 contract from NIAID to develop a live attenuated vaccine for tularemia. The company will develop a pilot batch of vaccine in two years for clinical testing, and it will evaluate the vaccine’s safety and immunogenicity, and support NIAID in Phase I testing. VaxGen. VaxGen (Brisbane, California) was spun out of Genentech in 1995 as a vaccine company specializing in the development of the AIDSVax vaccine against

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human immunodeÞciency virus (HIV). After the vaccine elicited disappointing results in Phase III trials, the company switched its focus to biodefense. In September 2002, NIAID awarded VaxGen a contract worth $13.6 million (later increased to $20.9 million) to conduct preclinical testing and to develop a feasibility plan to manufacture a 25-million-dose emergency stockpile of nextgeneration anthrax vaccines. The vaccines are based on an rPA102 vaccine that was initially developed by the U.S. Army Medical Research Institute of Infectious Diseases (USAMRIID). Protective antigen by itself is nontoxic and is thought to be one of the major antigenic components in BioPort’s crude cell-free Þltrate vaccine. In June 2003, VaxGen initiated a 100-person Phase I clinical trial to compare the safety and immunogenicity of rPA102 with that of BioPort’s vaccine. Within four months, this milestone led to an exclusive license agreement with USAMRIID whereby VaxGen received the worldwide rights to develop and commercialize rPA102. The company also received an $80.3 million, three-year contract from NIAID to conduct further animal studies, Phase II clinical trials, manufacturing scale-up, and validation of the vaccine, and to produce 3 million doses. In addition, VaxGen and the Health Protection Agency for England and Wales signed a letter of intent for a royalty-bearing contract under which the agency will manufacture and market the vaccine in the United Kingdom. In January 2004, the FDA granted rPA102 fast-track status to accelerate the development of the recombinant anthrax vaccine. In March 2004, VaxGen reported that Phase I of the rPA102 study demonstrated that immune responses at the higher doses tested were comparable to those elicited by BioPort’s vaccine and that the rPA102 vaccine was well-tolerated. In April 2004, the company initiated 13-month, Phase II trials of the anthrax vaccine and announced that it had submitted a proposal to the U.S. Department of Health and Human Services to produce up to 75 million doses. In January 2004, VaxGen partnered with Avanir Pharmaceuticals (San Diego, California) to manufacture and develop Avanir’s monoclonal antibody candidates, generated using its Xenerex technology, against B. anthracis infection and toxins. Avanir has demonstrated that a single dose of two of its most potent antibodies can provide protection against a lethal challenge of anthrax spores in mice. VaxGen is also developing smallpox vaccines based on the LC16m8 Lister strain of vaccinia virus. LC16m8 is an attenuated strain of Lister and has been shown to cause the least side effects among all the conventional smallpox vaccines. VaxGen hopes that its smallpox vaccine will have a better safety proÞle than vaccines based on the NYCBH strain. In December 2002, VaxGen entered into an agreement with the Chemo-Sero-Therapeutic Research Institute in Japan to initiate the development of the LC16m8-based smallpox vaccine in the United States. Under a previous agreement, the institute had already manufactured 1 million doses for VaxGen to test in animals. In April 2004, VaxGen announced that the results of animal efÞcacy studies (conducted in mice and rabbits) of its smallpox vaccine demonstrated that a single dose was as effective as DryVax in protecting against a lethal poxvirus challenge.

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Preparations to launch its HIV vaccine serendipitously positioned VaxGen to competitively bid for large government supply contracts for biodefense vaccines. In 2002, VaxGen established a $120 million joint venture, Celltrion (Incheon, South Korea), with a group of South Korean investors to construct a 48,000-liter mammalian cell culture facility in South Korea (expandable to 144,000 liters). In 2003, VaxGen opened a new ßexible manufacturing facility in South San Francisco. The facility is capable of mammalian or bacterial cell culture (with suites of 1000-L, 200-L, and 100-L bioreactors). VaxGen anticipates that the San Francisco facility will be able to manufacture approximately 100 million doses of the rPA102 anthrax vaccine. Therapeutic Drug Companies Several biopharmaceutical companies that develop protein or small-molecule therapeutic drugs have branched out into the Þeld of biodefense. Most of these companies are leveraging their existing technology and taking advantage of government funding to develop treatments for various biodefense-related pathogens, particularly anthrax and smallpox. Chimerix. Chimerix (La Jolla, California) is a privately held company that is developing drugs for smallpox, drug-resistant HIV infections, and viral hepatitis. Chimerix has a unique proprietary lipid-conjugate drug-delivery technology that can signiÞcantly enhance the oral bioavailability of existing drugs that are currently restricted to administration by injection. By covalently attaching lipids that promote intestinal absorption to known drugs, the company creates new compounds that are orally bioavailable. Once delivered into cells, intracellular cellular enzymes cleave the attached lipid and release the original drug. In September 2003, Chimerix licensed rights from Gilead Sciences to develop orally bioavailable derivatives of cidofovir (Vistide) for the prevention and treatment of smallpox infections. Cidofovir is an injectable nucleoside analogue approved for cytomegalovirus retinitis in AIDS patients. The drug also exhibits antiviral activity against a variety of DNA viruses and retroviruses, including vaccinia virus and smallpox virus. Chimerix has developed lipid derivatives of cidofovir and demonstrated that several derivatives administered orally protect mice against lethal challenges of cowpox and vaccinia virus. Also in September, Chimerix received a $36.1 million grant from NIAID to develop and test CMX-001, the company’s lead cidofovir derivative, an orally available smallpox therapeutic. In April 2004, the company was granted a U.S. patent (#6,716,825) covering CMX-001. Human Genome Sciences. Human Genome Sciences (HGS; Rockville, Maryland) is a genomics-based biopharmaceutical company with a market capitalization of $1.6 billion that develops protein therapeutics and antibody-based drugs. As part of its drug discovery program, HGS has developed a large infrastructure for creating monoclonal antibodies against therapeutically relevant receptors and secreted proteins identiÞed through the company’s target identiÞcation program. HGS has rights to multiple antibody technologies, including a

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collaboration with Cambridge Antibody Technology. In addition, HGS has developed signiÞcant cGMP manufacturing capacity to produce protein therapeutics, including therapeutic monoclonal antibodies, and has constructed a large-scale biologics manufacturing facility. Capitalizing on the antibody discovery and production capability that the company developed for its genomics-based drug discovery efforts, HGS has developed a therapeutic monoclonal antibody for the prevention and treatment of anthrax. In March 2003, HGS reported that its monoclonal antibody, ABthrax, neutralizes the ability of B. anthracis protective antigen (PA) to bind to its receptor. The company found that a single dose in rabbits and nonhuman primates signiÞcantly increased survival. Based on these results, the company initiated, in June 2003, Phase I placebo-controlled, dose-escalation clinical trials to test the safety, tolerability, and pharmacokinetics of intramuscularly injected ABthrax. In March 2004, the company announced that the studies showed ABthrax was well tolerated in healthy volunteers and achieved the required blood levels predicted by animal models for protection against inhalation anthrax. In August 2003, the FDA granted fast-track status to HGS’s ABthrax. Considering the new laws that enable marketing of a biodefense product on the basis of efÞcacy in animal models alone, HGS could have an anthrax therapeutic in one or two years. However, HGS has stated that further development of ABthrax is dependent on the government’s willingness to commit to the purchase of the product. OUTLOOK Biodefense is an exciting area that creates new opportunities for both existing and new companies. Incentives such as government funding of R&D, accelerated regulatory approval, and reduced time-to-market will encourage many biotechnology companies to explore the biodefense arena. Still, these companies will need to weigh those advantages against various risk factors. Navigating the complicated routes to government contracts can be challenging; know-how in this area can provide a company with a signiÞcant advantage. For example, Acambis, which secured government contracts worth $771 million, beneÞted from the expertise of its chief scientiÞc ofÞcer, Dr. Thomas Monath, who received his M.D. from Harvard Medical School, did Þeld research in Africa for the CDC on yellow fever and Lassa fever, and became a U.S. Army colonel and chief of virology at the U.S. Army Medical Research Institute of Infectious Diseases at Fort Detrick. Risks for companies pursuing biodefense include unpredictable markets, liability, price controls, and the loss of patent protection for products that become important to national security. Biotechnology companies entering the biodefense market will follow a variety of business models. There is an important question for companies to consider. Can a biotechnology company develop a sustainable business on biodefense alone? A few companies will probably try to focus solely on biodefense and procure large government supply contracts. Because one of the criteria for obtaining such

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contracts is the possession of large and expensive biotherapeutic manufacturing capacity, corporate development for small Þrms pursuing this avenue will be tricky. Most Þrms seeking biodefense funding will leverage it for the development of existing product pipelines in other therapeutic areas. Platform technology companies with drug discovery technologies (such as genomics and proteomics) that suffer from commoditization will Þnd new opportunities in governments looking to characterize the genomes and the biology of microorganisms relevant to biodefense. Incentives to stimulate development and commercialization of biodefense therapeutics await the enactment of new laws. Some biodefense laws were passed just six months after the terrorist attacks of September 11, 2001, and the subsequent anthrax bioterrorist attacks. However, Project Bioshield, proposed by President Bush in 2003 to stimulate biodefense, struggled through an inefÞcient lawmaking process. Project Bioshield provides $5.6 billion over ten years for the commercial development of vaccines and therapies for biological, chemical, nuclear, and radiological warfare agents. Under the legislation, the government guarantees drug purchases from companies developing therapies or vaccines against speciÞc biowarfare pathogens, accelerates the drug approval process, and allows the distribution of unapproved drugs under emergency conditions. In July 2003, the House of Representatives passed its version of the bill; the Senate Þnally passed its version in May 2004. Competition is already strong for companies developing biodefense products to address anthrax, smallpox, and plague, but opportunities are open for companies seeking to pursue certain Category A pathogens (including tularemia and hemorrhagic fever viruses) and most of the Category B pathogens. Although therapies for tularemia and Ebola virus may appear to be less of a funding priority than anthrax and smallpox, governments should not wait for a bioterrorism incident with those pathogens to act. Another avenue of investigation that governments and companies should explore are bioterror pathogens that extend beyond human diseases, such as veterinary and agricultural bioterror pathogens. The disruption of feed and food supplies could be as socially and economically disruptive to nations as the destruction of human life.

REFERENCES Inglesby TV, et al. Plague as a biological weapon. Journal of the American Medical Association. 2000;283:2281–2290. Inglesby TV, et al. Anthrax as a biological weapon. Journal of the American Medical Association. 2002;287:2236–2252. Jarrett CO, et al. Flea-borne transmission model to evaluate vaccine efÞcacy against naturally acquired bubonic plague. Infection and Immunity. 2004;72:2052–2056.

Fear of Theranostics: Why an Industry Has Not Really Emerged SUMMARY Theranostics—on-the-spot, highly accurate, DNA-based diagnostics that identify disease, predict a patient’s therapeutic response, or select patient subtypes for treatment—have arrived. Several such products are now available, and many companies are developing theranostics products or are exploring the tandem marketing of theranostics and therapeutics. But is theranostics an industry of its own or a new decision-making business tool? In this article, we examine this question in detail, reviewing the various roles theranostics might play, including functioning as a provider to Big Pharma under the made-to-order business model, developing as a stand-alone hybrid industry, acting as a revolutionary in the transition to integrated health care, and serving as a productive business tool that will be an integral part of the future health care industry. BUSINESS IMPLICATIONS •

•

Contrary to current thinking, theranostics is not a symbiotic, made-to-order business for the pharmaceutical industry. Despite the existence of a cluster of theranostics companies enabling pharmaceutical R&D, the dearth of comarketing deals and alliances indicates deep-seated issues (e.g., perceived differences in pricing, distribution, and customer service) between these two sister industries. Some companies (e.g., Roche, with its HepCare program) are attempting a hybrid approach to health care by offering both diagnostics and therapeutics in test-and-treat packages. However, size and scale issues will prevent most such companies from becoming major players and establishing a viable theranostics industry.

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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•

•

It is widely accepted that theranostics is a vital missing piece in the shift toward a world of predictive medicine—indeed, some industry pundits forecast that theranostics will revolutionize health care. However, the evolutionary pace of change denies theranostics a rapid path to growth as an industry on its own merit. Is theranostics an industry at all? The best approach may be to regard theranostics instead as a new decision-making business tool. This approach requires a shift away from pure-play theranostics companies that would serve as major product suppliers and toward a more service-based orientation. Theranostics can be a productive business discipline that will be critical to the future of integrated health care.

BUILDING BLOCKS OF AN INDUSTRY Theranostics—on-the-spot, highly accurate, DNA-based diagnostics that identify disease, predict a patient’s therapeutic response, or select patient subtypes for treatment—have arrived. Several such products are in use; Table 1 provides a sampling of theranostics product activity. There is no doubt that the concept of theranostics is entrenched and that theranostics has the potential for widespread use, as the following statement from Eduardo Balbona, M.D., formerly chief medical ofÞcer of HealthScreen America, which offers health education as well as screening tests and services, indicates: “The future belongs to those who move the identiÞcation process, using sophisticated medical technologies, to earlier stages. This proactive shift to a predictive medicine model will greatly improve outcomes, longevity and overall quality-of-life.” Trend analyses already portray theranostics as a major industry in its own right. Consider the following: • • •

One leading diagnostics group (Bayer Diagnostics) forecasts 20% annual growth of the theranostics industry until 2009. Predictive tests for a dozen or more conditions are likely to become available by 2010. By 2020, gene-based designer drugs for diabetes, hypertension, and other complex diseases will be commonplace.

BUT IS THERANOSTICS AN INDUSTRY YET? Theranostics is not yet established as an industry. Given the aforementioned predictions, the reason may seem to be timing and awaiting the correct moment in the evolution of the relationship between therapy and diagnostics. But it is not. In reality, the question requires a fundamental insight into the type of industry theranostics is now—and, more accurately and importantly, what type of industry it will become. It involves an analysis of the complex web of relationships and synergies that theranostics has spun in its relatively brief history. (The

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TABLE 1. Select Companies with Theranostics Tests in Use Company Abbott Diagnostics

Affymetrix

Axis-Shield

Bayer Diagnostics

CytoCell

Cytogen

DakoCytomation

diaDexus

Diagnoswiss

Digene

Comments Extensive portfolio of tests, platforms, and services for cancer, cardiology, endocrinology, infectious diseases, metabolic disorders, and women’s health, as well as numerous assays for therapeutic drug monitoring of patients. GeneChip microarray systems and various chips including kits for HIVa genotyping for drug-resistance determination, patients at high risk for osteoporosis, and various cancers. Developed by Axis-Shield and recently launched by Abbott Laboratories, the IMx Sirolimus assay monitors sirolimus (Rapamune) levels in transplant patients, which have to be properly maintained in each patient to optimize response. Sirolimus, developed by Wyeth, is a potent immunosuppressive drug indicated for the prophylaxis of organ rejection in patients receiving renal transplants. Diverse diagnostics, genotyping systems, and services in the areas of cardiovascular and kidney disease, oncology, women’s health, diabetes, and virology. Various genotype-based tests and screening solutions based on FISHb technology for cytogenetics and cancer. Developed various imaging and targeted cancer products including Quadramet, which is used to treat pain associated with prostate, breast, and other cancers that have metastasized to the bone; ProstaScint for detection of prostate cancer; and NMP22 BladderChek for screening and monitoring bladder cancer. Developed the HercepTest, a diagnostic test for HER-overexpression to fight breast cancer, used in conjunction with trastuzumab (Genentech/Roche’s Herceptin), and EGFR pharmDx, a test for the overexpression of EGFR used with ImClone’s cetuximab (Erbitux). Developing genomics-based tests and has launched the PLAC test, which measures blood levels of lipoprotein-associated phospholipase A2 (Lp-PLA2), a novel marker for heart disease. Developed products based on biochip technologies that integrate new methods for high-throughput protein analysis. Developed Hybrid Capture technology for gene-based tests including the hc2 High-Risk HPVc DNA test for cervical cancer, and DNAwithPap Test for adjunctively screening women to assess the presence or absence of high-risk HPV. Additional tests include detection of chlamydia, gonorrhea, hepatitis B, and cytomegalovirus.

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TABLE 1. (continued) Company

Comments

Gen-Probe

Developed nucleic-acid based tests to detect pathogens including chlamydia, HIV, HCV,d West Nile virus, and others. Developed various products and tests specializing in three areas: infectious diseases (hepatitis C, hepatitis B, and HIV), genetic testing (HLAe tissue typing and cystic fibrosis), and neurodegeneration (Alzheimer’s disease). Developed predictive nucleic-acid-based tests for BRCA1 and BRCA2 cancer genes for breast and ovarian cancers, MLH1 and MSH2 genes for colorectal and endometrial cancer, and the p16 gene for melanoma and pancreatic cancer. In collaboration with Aventis, developed the Enox test, which is a rapid, point-of-care test that detects anticoagulant effects of enoxaparin (Aventis’s Lovenox). Diverse portfolio of tests and services for infectious diseases, cardiology, cancer, diabetes, osteoporosis, and others. Recently launched the AmpliChip CYP450 (based on the Affymetrix GeneChip DNA analysis platform) microarray technology as a broad diagnostic to test two genes that can influence drug metabolism and adverse drug reactions in individual patients. Developed high-density genome-wide scans through its MassArray platform, and DNA analysis technology to identify genetic variation, and has discovered multiple candidate genes in breast, lung, prostate, and skin cancers, central nervous system disorders, metabolic and cardiovascular diseases, and musculoskeletal and inflammatory conditions. Developed Therasol, a drug delivery/therapeutic agent. The technology specifically carries the therapeutic agent directly to the target. Current drug formulations are neutral carrying vehicles. Subsidiary of Abbott Laboratories that has developed various genotyping tests based on FISH and microarray technology to detect cancer and infectious diseases. In August 2004, the FDA approved Ventana Medical Systems’ VentanaDX c-KIT rabbit monoclonal antibody diagnostic test that allows doctors to detect the presence of the c-KIT protein in gastrointestinal stromal tumors (GISTs), which will be pertinent in the selection of patients eligible for imatinib (Gleevec) because the drug inhibits the kinases c-KIT and platelet-derived growth factor (PDGF).

Innogenetics

Myriad Genetics

PharmaNetics

Roche Diagnostics

Sequenom

TheRyte

Vysis

Ventana

a HIV, human immunodeficiency virus. b FISH, fluorescence in situ hybridization. c HPV, human papillomavirus. d HCV, hepatitis C virus. e HLA, human leukocyte antigen.

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FIGURE 1. Role of theranostics in patient/therapy management

concept of proactively designing tests for therapy is less than ten years old.) This analysis will reveal how to sow, nurture, and harvest theranostics so that it can assume its rightful role in integrated health care. (Figure 1 highlights key roles for theranostics in integrated health care.) THERANOSTICS AS A MADE-TO-ORDER MODEL? The vast majority of companies operating under the theranostics banner today are promoting their ability to enable a particular R&D dimension of the pharmaceutical supply chain (Table 2 and Figure 2). In essence, they are applying a tried and tested business model based on made-to-order principles. Under a made-to-order model, a pharmaceutical partner might require a test to screen for a disease or to monitor patients in a clinical trial. It would then commission a diagnostics company, expert in its particular therapeutic Þeld, to develop and supply the test within a given time frame. In return, the pharmaceutical company would pay a contract fee. The diagnostics company might then be asked to continue to support the diagnostic or help with regulatory approval and commercialization. TABLE 2. Select Companies Active in Theranostics by R&D Focus R&D Focus Pharmacogenomics/genetics

Pharmacodynamics Therapy prerequisite tests Integrated health care marketing

Select Companies Cytogen, Diagnoswiss, Genaissance, Nuvelo (formerly Variagenics), Pharmacogenomics, Sequenom PharmaNetics, Roche DakoCytomation, GlaxoSmithKline, Novartis Cholestech, Prometheus, Roche

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FIGURE 2. Primary focus of theranostics players

The made-to-order model has worked well in other industrial sectors, and several companies that have followed that model have emerged as leaders in their Þeld. Examples include the following: • • •

Boeing built its commercial airplane business on the back of development contracts with the U.S. military. Microsoft was a ßedgling software house until it responded to a particular request for a PC operating software system for IBM, called DOS. Pixar grew rapidly on the strength of lucrative development and distribution deals with Disney.

On initial examination, the made-to-order label seems appropriate for theranostics: under this model, the pharmaceutical industry can commission the speciÞc diagnostics it requires to develop and market its products and, in turn, can help diagnostics companies offset the costs of lengthy, high-risk product development. The same type of contractual relationship between, for example, Boeing and the Pentagon—a relationship that allows Boeing to utilize speciÞc parts of the technology developed with the military on its own commercial aircraft—is often present in theranostics deals forged to date. Diagnostics partners have the freedom to seek other partnerships—in addition to the deal with the commissioning pharmaceutical company—for the same product. The pharmaceutical industry has a huge Þnancial incentive to critically assess and commission theranostics that can help improve its ailing productivity at an R&D level. The Tufts Center for the Study of Drug Development suggests that the success rate in pharmaceutical development, from target to market, is approximately 1.5–2%. From initiation of clinical trials to market, it is 10%. Thus, the fully loaded cost to bring a new drug to registration, according to the Tufts Center, is now $897 million. Between the end of 1998 and 2002 alone, costs

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have risen by just over $200 million as a result of a 50% decrease in productivity in the same four-year time frame, according to the Centre for Medicines Research International, based in the United Kingdom. Too many drugs are failing to travel successfully through the development pathway, resulting in ailing productivity and skyrocketing development costs. Theranostics can improve the success ratio and have a profound Þnancial impact: the ability to better identify patients for whom therapy is effective or less toxic will yield huge productivity increases in target selection. Promoters of the made-to-order theranostics model say the beneÞt to the pharmaceutical industry is considerable. Theranostics, they say, will facilitate smarter decisions regarding product development at multiple levels. The Centre for Medicines Research International believes that the application of disease genetics and pharmacogenetics together could save as much as two-thirds of the current cost to develop a drug and reduce development time by nearly two years. Moreover, a PriceWaterhouse Coopers report forecasts—quite boldly—that pharmacogenomics alone could save drug manufacturers approximately $130 million per approved compound. Theranostics can also have a Þnancial impact beyond the walls of R&D. For example, the use of tailored patient screening at the clinical trial stage can help select the 30–50% of patients for whom typically a therapy has no impact. With this capability, drugs previously withdrawn or orphaned as a result of narrow therapeutic effectiveness have the potential to be resurrected. In his recent “win some, lose some” analysis (published in Pharmaceutical Executive in October 2003), Stan Bernard (a management consultant who teaches pharmaceutical management at the Wharton School of Business) uses a mathematical approach to illustrate and quantify the beneÞts in brand share to those marketing a drug in conjunction with a theranostic—the combination, he suggests, can provide a 75% (or greater) increase in revenue, far outweighing the reduction in the target market that results from the diagnostic deselecting patients for whom the drug will be ineffective. If these suggested Þnancial beneÞts are borne out in practice, the pharmaceutical industry will have a clear incentive to commission diagnostics companies to develop theranostics under a made-to-order model. After all, the last time the pharmaceutical industry was presented with such R&D productivity gains—in the late 1980s, via combinatorial chemistry and pharmacokinetics—pharmaceutical companies quickly sought to internalize the new skills and to acquire the ßedgling companies that offered new approaches. Given this historical perspective, we wonder why—if the pharmaceutical industry truly is convinced that theranostics is an enabling industry—pharmaceutical companies have not started rapidly acquiring their theranostics partners. Yet no such industry effort to internalize the skills inherent in a theranostics focus has happened to date. One of the last companies to conduct a mega-merger, GlaxoSmithKline, actually closed its embryonic predictive medicine division (which focused on theranostic links with its therapy pipeline) as part of the merger.

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Closer inspection of the putative marriage between pharma and diagnostics reveals deep-seated fears among many in the major pharmaceutical companies, fears that emanate from the perceived dissimilarities between the therapy market and the diagnostics market (in terms of pricing, distribution, reimbursement, intellectual property, branding, and customer service) and from the loss of control implicit in implementing theranostics universally, outside R&D settings. QUANTIFYING THE BENEFITS OF THERANOSTICS Within pharmaceutical R&D, it is relatively easy to quantify and thus Þnancially measure the synergies of theranostics and pharmacotherapies in therapeutic target selection by identifying the patient subsets for whom the therapy is either ineffective or toxic. Such data are painstakingly gathered in the early stages of drug development. However, as Eddie Blair of Integrated Medicine notes in an interview conducted for this article, “quantiÞcation of beneÞt becomes much more woolly when it comes to marketing.” This statement refers speciÞcally to the lack of precedents and published proof that a diagnostic can improve the market share or competitive position of a therapeutic with which it is partnered. In turn, many pharmaceutical marketing directors and boards are skeptical about the claims made for theranostics at the marketing stage and are extremely cautious—at best—about theranostics. John Funkhouser, CEO of PharmaNetics, which partnered with Aventis to produce a test used with Aventis’s enoxaparin (Lovenox), agrees. According to Funkhouser, the Aventis partnership was constructive at the R&D phase, but problems of shared expectations arose at the marketing phase. Aventis’s R&D department saw the test, Enox, as a way to help underpin the clinical efÞcacy of enoxaparin in restricted patient populations. To Aventis’ marketing department, however, Enox was a potential barrier to empirical prescribing. According to a Morgan Stanley report, “Aventis have long continued to resist any suggestion that there is a need to monitor Lovenox. Indeed, this claim is the principal thrust of their competitive advantage over UFH [unfractionated heparin].” Chris Towler, who was a senior member of the technical marketing team that rolled out GlaxoWellcome’s ßu drug zanamivir (Relenza), tells a similar tale. While rapid ßu tests were useful in clinical development, GlaxoWellcome’s marketing team perceived the tests as a major implementation problem and thus as a threat to the empirical prescribing of zanamivir. Indeed, despite the availability of a range of physician-ofÞce ßu tests from companies including Binax and Quidel, no pharmaceutical companies are marketing any notable test-and-treat programs for ßu. Likewise, no companies are tapping the obvious beneÞts of reducing antibiotic resistance via pretreatment screening. Although pharmaceutical companies are unable to quantify the Þnancial advantages of theranostics, they can measure the Þnancial downside of prescreening and deselecting patients for whom a drug is not appropriate. Particularly for blockbuster drugs administered as part of primary care, the downsides—such as, for example, removing 30–50% of the target patient population because it is

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known that the drug may not have any beneÞt for those individuals—are easily quantiÞed because these epidemiological data will have been gathered during clinical development. It is therefore simple for marketing directors to see how a diagnostic that targeted certain patients and not others for treatment might reduce their market share. Completely unknown and untested, however, are the potential upsides of using theranostics to increase the number of patients receiving a therapy or to increase revenues through better patient compliance. Understanding and realizing these beneÞts requires a fundamental reengineering of the tandem marketing of drug and diagnostic. REENGINEERING THE MARKETING PROCESS TO INCLUDE THERANOSTICS The CEOs of diagnostics companies have a remarkably common complaint about Big Pharma: unlike the pharma company R&D teams with whom they often work well over long time frames, the pharma company brand managers rotate (relatively speaking) like revolving doors. This inconsistency is no minor irritation, given that the brand manager is not incentivized to a speciÞc collaborative theranostics R&D program and (as illustrated previously) more often views the theranostics as a threat than as a boon. Pharmaceutical brand managers are equally and legitimately dismayed at what they perceive as a lack of basic direct sales and marketing principles among the diagnostics companies. This lack of experience in, for example, selling diagnostics directly to primary care physicians is not surprising because so many diagnostics executives have been trained within a culture of volume sales, discounts, and thirdparty distribution arrangements. In the U.S. health care market, Þnding diagnostics partners that can and will promote their own unique diagnostic tests directly to physicians is not an easy task. Typically, small to midsize diagnostics companies expect the pharmaceutical company, as the party with theoretically the most to gain, to Þnance and even handle sales and marketing. Pharmaceutical brand managers are accustomed to a sophisticated marketing environment where the predominant skill is creating a context conducive to drug prescribing. Big Pharma creates this environment by “prelaunching” products (i.e., marketing them before launch through channels such as publications, patient forums, and disease awareness programs) and by investing heavily to develop the market. The pharma brand managers’ world spans everything from the sponsored development of clinical guidelines to patient advocacy programs. The results of their efforts are easily measured by sales—sales that are generated via tailored messages delivered in intricately planned sales campaigns by direct representatives who might have three minutes to capture a physician’s attention. As such, these pharmaceutical managers are currently ill equipped to assume the task of copromoting tests that seldom employ the same level of branding disciplines, have uncertain adoption curves, and require several 20- to 30minute sessions with the physician. Add to these difÞculties the age-old problems (or perceived problems) associated with diagnostics—low, highly variable, or

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nonexistent reimbursement proÞles as well as uncertain intellectual property proÞles—and the copromotion of theranostics with therapy is revealed to require a fundamental reengineering of the selling and marketing process to ensure the successful diffusion of the theranostics prior to or in parallel with the therapy. Some say that the theoretical increases in sales of pharmaceuticals based on packaging of those pharmaceuticals with theranostics are a clear indication that a reengineering of sales and marketing is appropriate. However, those charged with ensuring the success of sales and marketing approaches are unlikely to welcome theoretical sales increases driven by the fundamental shifts in sales and marketing approaches that would be necessary to address untested copromotion techniques. THERANOSTICS FROM THE PAYER/REGULATORY PERSPECTIVE Payers, such as those in managed care, have employed a variety of means to reduce costs: examples include generic substitution, use of formularies, centralization of laboratory costs, and encouragement and development of capitated physician networks. (In fact, in response to increased levels of payer control, pharmaceutical marketing has over the past ten years shifted toward the consumerization of U.S. health care and away from the control of managed care. Direct-to-consumer advertising and patient advocate programs caused a major upsurge in pharmaceutical sales during the early 1990s and have become a mainstay of blockbuster marketing programs ever since.) Pharmaceutical companies’ fear of theranostics in the hands of managed care is not irrational. As the health care world evolves toward integration of health care decisions, in the future, payers will likely embrace the results of studies that help preauthorize expensive treatment. The pharmaceutical industry has major boardroom concerns regarding this development. Resisting universal promotion of theranostics is one way to dam the ßoodtide. Although managed care has as yet promulgated no wholesale policy to use theranostics for pretreatment authorization purposes—which would result in a deselection of patients from expensive treatments (as discussed later in the context of integrated health care)—the studies have begun. Promoters of theranostics cite recent FDA pronouncements as good news for the theranostics industry because they will theoretically force the pharmaceutical industry to embrace theranostics more openly. As then-FDA commissioner Mark McClellan stated in 2003, “Certain new therapies will be developed along with genetic or phenotypic tests that can identify the responding sub-population, detect individuals who need a different dose, or Þnd people who are prone to a certain toxic effect. Development of these test/therapy combinations must be facilitated, because they have the potential to maximize drug beneÞts while minimizing toxicity.” As the rebuttal of Hillary Clinton’s health care reforms in the early 1990s suggests, the pharmaceutical industry can move very slowly on proposed revolutions in the treatment paradigm. After the National Riße Association, the pharmaceutical industry is the largest spender of lobby dollars in Washington. Although

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the FDA has clearly put the debate on the agenda, it has not established any mandatory policies regarding theranostics. EXCEPTIONS TO THE RULE: THERANOSTIC/DRUG COPROMOTION Many cite copromotion deals between theranostics companies and the pharmaceutical industry as evidence that Big Pharma has changed its approach to marketing theranostics. However, many joint theranostics initiatives are made in response to a speciÞc market or technical problem and are thus focused in scope and are seldom geared toward major-market development. Table 3 details select theranostics/therapy projects. As we have discussed, the impact of using theranostics to screen patients who are candidates for treatment with blockbuster drugs remains unknown but is perceived to have downsides to the industry in that the screening will limit TABLE 3. Rationale for Testing: Examples of Theranostics Projects Driving Company Merck

Genentech

GlaxoSmithKline (GSK)

Novartis

Bristol-Myers Squibb

Rationale HDL/LDL were not widely agreed as markers of heart disease. Merck had to prepromote cholesterol testing as a diagnostic to change its cholesterol-lowering drugs from therapies targeted at restricted patient numbers to a new billion-dollar drug category. Trastuzumab (Herceptin) likely would not have received approval from the FDA without parallel approval of a dedicated diagnostic test from DakoCytomation known as the HercepTest. Abacavir (Ziagen), an anti-HIV drug, has a potentially fatal hypersensitivity reaction in approximately 5–10% of patients who take the drug. Together with the FDA, GSK is conducting clinical trials aimed at matching single nucleotide polymorphisms (SNPs) to drug toxicity responses including a program for abacavir to determine whether biomarkers exist that may help to predict a higher risk for developing this condition. Traditional karyotyping is used to find a particular single genetic defect associated with leukemia (the Philadelphia chromosome, caused by the splicing of chromosomes 9 and 22) and led to the development of imatinib (Gleevec). Without this marker to screen patients, imatinib would likely not have received approval. BMS’s drug paclitaxel for breast cancer relies on Myriad’s BRACAnalysis test, which assesses a woman’s risk of developing breast or ovarian cancer based on detection of mutations in the BRCA1 and BRCA2 genes.

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the treated patient population. Interestingly, the opposite is perceived to be true with specialty drugs, where the pendulum of unknowns swings in favor of theranostics programs. In these cases, the unquantiÞable dangers that lurk in toxicity or ineffectiveness drive the need to identify appropriate patient populations to treat. The prospect of copromotion with therapies that require very careful patient selection is good news for theranostics companies whose development programs target orphan drugs or very speciÞc marketing problems—for example, the need to monitor a therapy once administered. This upside, however, should not be mistaken for a shift toward a universal market development approach to theranostics in combination with major blockbuster drugs. PROBLEMS WITH THE MADE-TO-ORDER MODEL For theranostics to become a truly made-to-order industry for Big Pharma, both parties need to be in harmony. This harmony does not yet exist, however. Those who forecast a growing synergy between these two sister industries ignore two major and conßicting points: • •

Today’s pharmaceutical industry is highly conservative when it comes to driving internal change or facilitating external change. Theranostics is all about change at every level of the supply chain.

The recent lackluster share performance of major drug companies—such as GlaxoSmithKline, whose shares have fallen by as much as 20% from their Þveyear peaks, and AstraZeneca, whose share price suffered dramatically after the FDA failed to approve its new anticoagulant, ximelagatran (Exanta)—may force consideration of new approaches to blockbuster marketing. However, the lack of new hybrid skills at the board table that would enable integration of theranostics into all aspects of the pharmaceutical supply chain will deny theranostics the requisite focus. Deals between theranostics and pharmaceutical companies will continue to be forged, but they will be selective to R&D and specialist pharmaceutical products, reactive to speciÞc marketing problems, and limited in scope. If theranostics is to emerge as a viable, separate industry in the near term, it needs to move beyond complete reliance on the made-to-order model. The degree of change required on the part of the pharmaceutical companies is unlikely to occur in the foreseeable future. Thus, theranostics will never be afforded the central role needed to become a prerequisite standard if it relies only on momentum from the pharmaceutical industry. Rather, it needs to Þnd that momentum by concurrently pursuing other strategies. THERANOSTICS AS A STAND-ALONE HYBRID INDUSTRY A small number of theranostics companies have decided to develop and promote integrated diagnostic and therapy products in parallel rather than wait to be

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a made-to-order provider for the pharmaceutical industry. Given the potential problems of relying only on pharmaceutical partnerships to drive the theranostics industry, this independent development and promotion seems outwardly like a strategically smart move, giving birth to a truly novel hybrid business model. Already, integrated diagnostic and pharmaceutical companies such as Roche and Abbott appear to a have a lead in such approaches. In particular, Roche’s recent strategic moves in diseases such as hepatitis (with its HepCare program) and HIV (with its HIV and hepatitis initiative) suggest a focus on integrating the company’s product offerings in these conditions into one-stop-shopping programs. In doing so, Roche hopes to reposition itself as a “health information provider” and offer a full array of services, from laboratory analysis and molecular typing right through therapy. Such a move would take advantage of Roche’s two major business arms and differentiate the company from the pureplay pharmaceutical companies that are so at odds with the diagnostic side of the industry. Although some mega-companies are identiÞed as integrated diagnostic/pharmaceutical companies, in reality the business divisions (diagnostics and pharmaceuticals) of these behemoths remain distinctly and culturally unjoined. Of their current revenue, 90% comes from traditional business silos in therapy or laboratory sales. As such, their focus will, for a considerable time, remain in the traditional model, and their pharmaceutical arms will be subject to the same reticence with regard to diagnostics/theranostics common in the pharmaceutical industry. We are unlikely to see large integrated companies such as Abbott or Roche rebrand themselves as theranostics companies any time soon. Smaller companies are also emerging with a hybrid therapy/diagnostic approach focusing on either a disease or a business segment. Examples include the following: •

•

•

Millennium is focused on using the molecular understanding of disease to more effectively design therapeutic treatments and diagnostic products for cardiovascular, inßammation, and oncology indications. Interleukin Genetics aims to be the world leader in understanding many chronic diseases or disabilities such as coronary artery disease and diabetes. It too is working on therapy and diagnostic combinations. Prometheus Laboratories uses existing tests to protect and revitalize generic therapies in many conditions, including Crohn’s disease.

However, such companies face a series of size and scale hurdles. Companies that build expectations and aim to develop their own proprietary diagnostics and therapies are subject to at least double the R&D investment risks as companies dedicated to developing only diagnostics or pharmaceuticals. In the early 1990s, Cortecs tried to develop both therapies and diagnostics for diabetes and osteoporosis. However, the 1999 failure of the company’s lead therapy, an oral calcitonin for osteoporosis called Macritonin, to make it through Phase III clinical trials dashed the hopes of the company and its investors, leaving Cortecs with

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a handful of point-of-care tests and a decimated share value. In fact, Cortecs now also has a new name (Provalis) and business plan (two divisions focus on diagnostics and health care). Furthermore, the integrated diagnostic/therapeutic strategy may be restricted to specialist areas given that many of the major mainstream diseases are proving to be polygenomic—diabetes, for instance, is perhaps the product of ten or more different genetic lesions. For these polygenomic diseases, small companies will Þnd it difÞcult (if not impossible) to provide the comprehensive test-and-treat solutions they are using to differentiate themselves in the Þrst place. The alternative strategy of avoiding expensive and risky therapy development by selecting established generic therapies—as Prometheus has done with its “inch wide, mile deep” approach (in other words, a dedicated focus to be an integrated health care provider but to only a small number of therapeutic areas)—has signiÞcantly lower development risks but is instead subject to potential competition emanating from novel therapies that could outmode the therapy part of the test-and-treat equation. Lack of marketing dollars increases risk as well. The cost of appropriate marketing for novel markers (either in the laboratory or in the physician’s ofÞce) alongside a drug is considerable, even if the therapy has an established patient pool. The FDA is already worried about moving serious diagnostic decisions out of the hands of trained pathologists. Small niche players, although they may be culturally engineered to market therapy alongside diagnostics, simply do not have access to the marketing budgets of their larger brethren that would allow them to compete effectively outside the highly focused clinical domains. In addition, theranostics that are based on molecular testing platforms will need to overcome several infrastructural and ethical challenges before they make it into the physician’s ofÞce. Note, for example, the issue of providing counseling to patients for whom a genetic test might show a predilection for a particular disease. Questions of who will provide and fund these additional services are already being debated in regulatory and industry circles. In sum, considerable hurdles impede the evolution of a stand-alone theranostics industry based on a test-and-treat paradigm. Large integrated diagnostic/pharmaceutical players with the deep pockets required to fund the marketing budgets essential to be successful do not in the short term want to reposition themselves as stand-alone theranostics companies. As a result, their participation will be selective and conÞned to speciÞc disease sets. A part from marrying existing therapies with existing tests, the cost of developing entirely new integrated tests and therapies appears to be prohibitive for smaller specialist stand-alone players in theranostics. If such companies are well funded or are acquired at the commercialization stage, they will be able to handle R&D risks as well as the marketing costs that arise when promoting novel tests with novel therapies. Without the right level of funding, the smaller companies are likely to remain highly specialized. Although a specialist niche is a respectable and often lucrative home for investment dollars, it does not an industry make.

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THERANOSTICS AS A HEALTH CARE REVOLUTIONARY Another raison d’etre ˆ bestowed upon theranostics is that of the revolutionary oil to the engine of integrated health care. At its heart is a vision where a patient’s physician delivers seamless and tailored care, without the huge cost inefÞciencies inherent in our current disintegrated system. The vision is not without strong logic. For diseases characterized by educated physicians, symptomatic patients, great therapy, and cost-conscious payers, theranostics can facilitate the remaining requirements, which are as follows: • • • •

Find the patients for whom the therapy will be most effective. Test those patients on the spot. Monitor their response to therapy remotely. Adjust their therapy accordingly and make them better.

Two stakeholder segments have the potential to harness the lubricating potential that theranostics brings and will thus contribute to driving the industry forward: managed care and specialist network providers. Managed care companies such as Wellpoint, Anthem, and Healthnet are in the business of triaging patients effectively. Together, these companies cover 30 million lives and have created a variety of preferred provider organizations (PPOs), HMOs, and various hybrid and specialty network health care services to combine effective cost-control techniques. Healthnet boasts a major reduction in its drug bill from an industry average of 9% of costs annually to 5%. Duncan Moore of Morgan Stanley believes that it is only a matter of time before these groups adopt theranostics as a way to impose prior authorization for treatment. In moves that seem to validate the fears within Big Pharma, a number of managed care groups including Kaiser and United Healthcare are beginning to seek independent analysis from theranostics companies of a new drug’s relevance and cost–beneÞt prior to reimbursement. Physician and patient demand can also promote adoption of theranostics. Integrated networks of clinics or providers such as Salick and HealthScreen are already marketing their ability to more efÞciently and more accurately manage routine health checks (for conditions such as heart disease and high cholesterol) and costly diseases (such as cancer). Theranostics are a fundamental part of their future as they seek to offer payers or patients a more cost-effective management program. Interestingly, motivated physician networks that internalize new theranostics as part of their product also have the ability to eliminate the costly physician detailing and marketing issues that arise for developers wishing to market novel diagnostics allied to novel therapies. When a physician community moves to deÞne how, where, and when a test-and-treat program is appropriate, it removes the need for sales representatives to promote to individual physicians (the most expensive step in the marketing chain). As Christopher Fey, HealthScreen America’s founder, puts it: “We have the capability to pioneer the way pharmaceutical companies, academic medical

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centers, employers, insurance companies, governments, and others involved in health care address a population’s future health risks.” We can expect provider networks and managed care to increasingly utilize theranostics in the short to medium term. This move bodes well for the future of theranostics. However, although theranostics will help facilitate better and more integrated clinical decisions, health care is a truly interdependent machine. A theranostics revolution would require a shift of profound proportions in all aspects of clinical care (including pharmaceutical adoption) toward a fundamentally different clinical delivery model. A move toward integrated medicine will not be fast—more evolution than revolution. Thus, the nascent theranostics industry cannot rely solely on a push to integrated medicine to provide its forward momentum. THERANOSTICS AS A DECISION-MAKING BUSINESS TOOL Is theranostics an industry yet? We indicated at the start that because theranostics is an embryonic business, the answer to this question lies in an analysis of the type of industry theranostics will become. It cannot exist solely as a made-toorder provider to Big Pharma or as a series of integrated test-and-treat companies. Nor can it wait around for providers and payers to integrate health care and in so doing embed theranostics as part of the equation. Perhaps the best architecture for theranostics promoters is to step outside the box and proclaim that theranostics will never be an industry in its own right. A better approach is to regard theranostics as a new decision-making business tool, much like accountancy. This approach requires a shift in the thinking of the pure-play theranostics CEOs, away from their vision of becoming major product suppliers and toward a service-based orientation. But it is worth considering some parallel examples of this label. Accountancy as a practice has evolved over generations via policies, rules, and guidelines into a virtually invisible enabler of business today. Like theranostics, accountancy involves tools that can be drawn into speciÞc business problems across the supply chain on an as-needed or opportunistic basis, helping gain insights, select better choices, and deÞne target groups. Like accountancy, theranostics is used by multiple stakeholders across the supply chain but is not always a prerequisite to good business decisions. In support of the idea of theranostics as a multirole business tool, consider the multiple attempts to rebrand theranostics away from diagnostics—for example, as personal medicine, integrated medicine, predictive medicine, preventive medicine, pharmanostics, and test-and-treat solutions. Table 4 shows the value that theranostics information has across the multiple stakeholders of the supply chain. Parallels can also be drawn between theranostics and the Internet. The many bullish predictions for theranostics suggest the birth of a new industry, much like the Internet was hailed ten years ago. Although Wall Street has applied capital to theranostics much more cautiously than it did with the dot-com businesses, it is worth reßecting that (with the exception of a few companies) the Internet turned out to be only a new productivity tool and not an industry after all.

TABLE 4. Uses and Value of Theranostics Across the Supply Chain Perceived Value to Regulators

Perceived Value to Payers

Perceived Value to Physicians

Perceived Value to Blockbuster Pharma

Perceived Value to Specialty Pharma

Pharmacogenomics

High

High

High

High

High

Toxicogenomics

High

High

High

High

High

Clinical trial/approval Pharmacodynamics

High Med

Med High

Low High

Low High

High High

Initial market development

Low

Low

Low

Low-med

High

New indications

Low

Low

Low

Low

High

Generic market management

Low

Low

Low

Low

High

Purpose of the Theranostic

Objective To determine how potential patients will respond to a molecule and thereby determine which molecules to develop To identify patients who might have adverse reactions to specific agents To facilitate approval of drugs To adjust drug dosage to obtain optimal therapy per patient To provide physicians with valuable information enabling them to individualize and optimize the therapeutic regimen of the patient To extend therapy into new sets of patients not previously identified To better identify patients for whom orphan drugs, which are difficult to market, are appropriate

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Theranostics might never be an industry in its own right, but it certainly is a productive business discipline that health care will be unable to live without. A roster of theranostics companies could evolve along the lines of international accountancy or information technology (IT) consultancy practices, getting involved with multiple players (big and small), bringing and codeveloping tools to help enable their clients’ business, and evolving with the future rather than driving it. We should not regard such a role as a diminution of the status of theranostics—it will not cramp the development of new theranostics products, services, or deals. However, it should force us to reconsider how we invest in, incentivize, and embed theranostics in all aspects of the value chain as we journey toward integrated health care. Equally, it should force us to better Þnancially quantify how theranostics will improve each clinical decision, an analysis that is greatly lacking in this Þeld. More progressively, health care companies and clinical networks that do not think of holding board meetings without representation by their chief Þnancial ofÞcer or chief information technology ofÞcer should consider bestowing equal boardroom status to new chief theranostics ofÞcers. These senior managers would be accountable for selling, promoting, enabling, quantifying, and reporting the many beneÞts of theranostics to their own particular supply chain. In turn, theranostics promoters should welcome this approach because it will shortcut the need to continually prove their net worth as external suppliers representing a disruptive industry. Instead, they would be seen as core elements of conducting competitive business in the new health care era.

Beyond Commodity Drugs: Strategic DiversiÞcation in the Generics Industry

SUMMARY The traditional model of generic pharmaceuticals as a purely commodity industry is moving closer to extinction. Intense competition has driven down margins and made sustainable growth in crowded markets nearly impossible. Many companies have responded with aggressive patent challenges in hopes of gaining Þrst-to-Þle status and 180 days of marketing exclusivity, but these legal Þghts are often long and complex and bring ßeeting rewards. As a result, generics manufacturers are turning to other strategies in their search for sustainable growth, increased pricing power, and reduced earnings volatility. In this article, we discuss the trends that support a healthy generic drug industry, the trends that are making it difÞcult for generics companies to compete, and the strategies that companies are pursuing to remain competitive. BUSINESS IMPLICATIONS •

The increasing presence of generics manufacturers from India is keeping downward pressure on prices in many commodity generics markets. Companies such as Dr. Reddy’s, Ranbaxy, and Cipla enjoy a signiÞcant competitive advantage over their counterparts in the United States and Western Europe thanks to their generally well-educated labor force, low-cost environment, and weak intellectual property laws. Although India began conforming to TRIPS in 2005 and thus lost its traditional head start, several companies there have had time to achieve “critical mass” and can effectively compete in the global generics industry.

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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•

•

•

Pharmaceutical companies that focus exclusively on commodity generics will face constant problems maintaining proÞtability and sustaining adequate growth. Successful companies have diversiÞed or will diversify into the manufacture of active pharmaceutical ingredients (APIs), the development of generic biologics, the development of branded products, the acquisition of niche products, or the development of their own new molecular entities (NMEs). The most common response to the intense competition and limited growth prospects in the commodity generics market has been to move into branded products. Companies such as Ivax and Teva started on this path years ago and have aggressively launched a variety of branded products. DiversiÞcation into branded products creates a hybrid pharmaceutical company that combines low-margin commodity generics with higher-margin brands. The lines between branded and generics manufacturers will continue to blur. One by one, surviving generics companies will evolve into more traditional midtier pharmaceutical companies with focused portfolios of drugs targeted at smaller markets and/or enhanced drug delivery. Some will try to achieve this reinvention through organic growth, but many will Þnd mergers and acquisitions to be the fastest route.

INTRODUCTION Each day, the traditional model of generic pharmaceuticals as a purely commodity industry moves closer and closer to extinction. Intense competition, particularly from manufacturers in India, has driven down margins and made sustainable growth in crowded markets nearly impossible. Many companies have responded by mounting a series of aggressive patent challenges in hopes of gaining Þrstto-Þle status and 180 days of marketing exclusivity. Although this strategy has provided several generics companies with impressive boosts in revenues, the legal Þghts are often long and complex and bring only ßeeting rewards. For investors and management alike, this litigation risk can be quite unsettling. As a result of the increasing competition and the unknowns surrounding patent challenges, generics manufacturers are turning to other strategies in their search for sustainable growth, increased pricing power, and reduced earnings volatility. Although most companies have chosen to pursue some form of diversiÞcation into branded drugs, others have selected very different strategies, including backward integration into active pharmaceutical ingredient (API) manufacturing and development of generic biologics. Against this backdrop, the pure generics manufacturer operating solely in commoditized market segments will likely go the way of the dinosaur. AN APPARENTLY HEALTHY GENERIC DRUG INDUSTRY At Þrst glance, the future has never looked brighter for the generic pharmaceutical industry. Demand is increasing, pipelines are stocked, and national health authorities are working to speed the ßow of generics to market. In 2002, according to

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IMS Health, generics and branded generics accounted for 51% of all U.S. prescriptions (17% of sales). Three of the top eight pharmaceutical companies by total U.S. prescription volume (Mylan Laboratories, Teva Pharmaceuticals, and Watson Pharmaceuticals) were generics manufacturers. In 2003, three generics manufacturers made Fortune magazine’s list of the “100 Fastest-Growing Companies” in the United States: aaiPharma (#19), Barr Laboratories (#50), and Sicor (#52). Twelve-month stock returns (January–December 2003) for industry leaders have been robust: Teva Pharmaceuticals (47.6%), Mylan Laboratories (63.5%), Watson Pharmaceuticals (62.7%), and Barr Laboratories (77.3%). Indeed, several trends would appear to point toward good times ahead for the $15 billion U.S. generic drug industry. Steady Product Flow A steady stream of patent expiries is in many respects the lifeblood of the generics industry. The 2003–2007 period is poised to provide ample product ßow, in the form of both Paragraph III and Paragraph IV generics. (Paragraph III certiÞcations apply to abbreviated new drug application (ANDA) Þlings that do not involve challenges to patents still protecting branded products. Paragraph IV certiÞcation applies when patent protection has not expired but the generics drugmaker claims that either the patent is invalid or that its product does not infringe the patent. Generics drug companies aim to be Þrst to Þle ANDAs with Paragraph IV certiÞcation because the rules make them eligible for a 180-day period of marketing exclusivity.) Accurately handicapping the outcome of litigation and the launch of speciÞc Paragraph IV generics is very difÞcult, but two key Þndings suggest there will be many more successes. First, the FDA has dozens of such challenges on Þle, many involving blockbuster drugs with patent expiries in the 2003–2007 period. Second, according to a study by the Federal Trade Commission, generics manufacturers (through June 2002) have won almost 75% of all Paragraph IV challenges. Even if this Paragraph IV success rate declines, the large number of branded drugs being challenged points to a robust ßow of popular drugs for the generic drug industry over the next Þve to ten years. Growing Demand for Generic Drugs On the demand side, the outlook for the generic drug industry just keeps getting better and better. Overall use of pharmaceuticals will increase as the population ages, and with the Medicare drug beneÞt enacted in the United States, greater use of prescription drugs by this population is likely. In addition to the demographic and economic forces driving demand for prescription drugs, several factors favor the preferential growth of generics at the expense of branded drugs. Pharmaceutical cost containment is a primary concern in most major pharmaceutical markets, and a popular tactic for addressing the problem is encouraging the substitution of generic drugs for branded drugs. The Medicare drug beneÞt can be expected to increase the role of active formulary management and its strategy of promoting generic over more-expensive

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branded formulations. The concerted efforts of regulators, physicians, and payers to improve the public’s perception of generics also appear to be Þnally bearing fruit. As a result of these forces, the Þrst generic versions of blockbuster drugs are achieving maximum penetration in the United States in a matter of weeks. An Increasingly Favorable Regulatory Environment Completing the generic drug industry’s apparently favorable outlook is an increasingly accommodating regulatory environment. In a September 23, 2003, speech in Mexico, FDA commissioner Mark McClellan unambiguously indicated the agency’s agenda as he declared, “improving our own generic drug industry has been one of my top priorities as FDA commissioner.” In a January 27, 2003, letter to the Generic Pharmaceutical Association (GPhA), President George W. Bush lavished praise on the trade group. He stated: “I commend the GPhA for your commitment to providing consumers with critical information, and I applaud your ongoing work to advance affordable, quality medicine. Your important efforts bring hope and healing to countless individuals and help to enhance the quality of life for all.” As a result, resources have been added and regulations modiÞed to improve the efÞciency of the FDA’s generic-drug review process. In another key move, the agency closed a legal loophole by limiting branded manufacturers to a single 30-month stay in a Paragraph IV patent challenge. Regulatory authorities in Europe have also demonstrated increased willingness to promote the use of generics, including in markets with historically low generic penetration (e.g., France and Spain). In addition, the expansion of the European Union will bring into the system many new Central and Eastern European nations with traditionally high generics penetration. CHALLENGES TO THE GENERIC DRUG INDUSTRY Despite all the factors suggesting a rosy future for the generics industry—or, in part, because of them—competition in many markets is intense, challenging both proÞtability and growth. In the United States, the “plain vanilla” oral generic (past the 180-day exclusivity period, if any) has become a commodity. Small and midsize generics producers are increasingly squeezed out of supply relationships by companies offering broader menus of products (e.g., Teva, Mylan, Watson, Sandoz, Alpharma). Making matters worse for the entire generics industry is the consolidation of U.S. wholesalers into three powerful companies (AmerisourceBergen, Cardinal Health, and McKesson) that represent 90% of the basic drug distribution. This small number of buyers creates considerable opportunity to play one generics company off another. Adding to the problem of growing wholesaler power is the consolidation of pharmacy beneÞt management companies (PBMs) into a similarly concentrated market (i.e., Caremark, AdvancePCS (target of a planned Caremark acquisition), Medco Health Solutions, and Express Scripts). The situation is no better in Japan or in the major European markets, where government agencies render the pricing power of generics virtually nonexistent.

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Competition is particularly acute in the year following the loss of patent protection or exclusivity for a successful branded drug (or Þrst-to-Þle Paragraph IV challenger). It is not unusual for 10–15 generics to win approvals for their ANDAs and enter the market almost simultaneously. Many of these entrants subsequently drop out in the face of Þerce price competition and overall market volatility; ultimately, the market may stabilize with Þve to six key generics and somewhat Þrmer prices. Increasing Competition for First-to-File Status The potential windfall from winning 180-day marketing exclusivity for Þrstto-Þle generics has created a powerful incentive for challenging drug patents. Barr’s generic version of Eli Lilly’s Prozac (ßuoxetine), for example, earned $350 million in the United States during the drug’s 180-day exclusivity period in late 2001. Given this strong incentive for successfully litigating a Paragraph IV challenge, competition to be Þrst to Þle among generics companies is signiÞcant. Furthermore, the number of major Paragraph IV opportunities is likely to decline in the future as innovators learn from past mistakes and build more-robust patent estates around promising molecules. Pressure from Generics Manufacturers in India The increasing presence of Indian generics manufacturers continues to keep downward pressure on prices in many commodity generics markets. Companies such as Dr. Reddy’s, Ranbaxy, and Cipla enjoy a signiÞcant competitive advantage over their counterparts in the United States and Western Europe for a variety of reasons. India offers a generally well-educated labor force coupled with low construction, manufacturing, and labor costs. Historically, Indian companies have excelled in reverse engineering and manufacturing chemistry. Adding to these factors is India’s weak intellectual property system, which has provided companies there with signiÞcant manufacturing and marketing experience prior to the launch of generics in the U.S. and Western European markets. By the time a drug’s patent has expired or a company has lost its exclusivity for that drug to allow entry into the United States and Europe, Indian generics manufacturers have already gained signiÞcant experience manufacturing the drug for its large domestic market and for emerging nations not part of the World Trade Organization’s Agreement of Trade-Related Aspects of Intellectual Property Rights (TRIPS). Although India began conforming to TRIPS in 2005 and thus lost its traditional head start, several Indian companies have had time to achieve “critical mass” and are able to compete effectively in the global generics industry. STRATEGIES TO SURVIVE THE GENERIC DRUG MARKET As a result of the forces just discussed, pharmaceutical companies that focus exclusively on commodity generics will face constant problems maintaining profitability and sustaining adequate growth. Indeed, most generics manufacturers

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have expanded their business models beyond the sale of commodity products using one or more of the following strategies: • • • •

Backward integration into the manufacture of active pharmaceutical ingredients (APIs). Manufacture of generic biologics. Development of competency and efÞciency in manufacturing “difÞcult” generics. DiversiÞcation into branded generics.

With these strategies come varying degrees of risk and reward (Figure 1). The result of these and other diversiÞcation strategies has been to preserve (or grow) margins and reduce the disruptive effects of volatile commodity generic pricing. API Sourcing Although the global API market is increasingly driven by entrants from India and China, several other generics companies have added or continue to build their capability in API manufacturing. Such backward integration can take two fundamentally different forms: manufacture primarily for internal use and contract manufacture for other pharmaceutical companies. In either case, retaining or adding API capability addresses a variety of Þnancial and operational issues.

FIGURE 1. Risks and rewards of generics manufacturers’ strategies

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In the Þrst year after a drug’s loss of patent protection (or exclusivity), generics Þrms rapidly enter the market, at times outstripping the supply of bulk ingredients. In some markets, reports have even surfaced of generics companies attempting to buy up all available API capacity in a series of exclusive partnerships. Supply constraints may lead to cost increases for companies that have failed to lock in secure supply agreements. Even worse, sourcing problems may have a ripple effect and delay deliveries to wholesalers, jeopardizing one of the few areas of differentiation still available to a generics company operating in the commodity segment. As a result of periodic supply constraints, API sourcing has evolved from a tactical concern to a strategic one. Backward integration into API manufacturing helps ensure a reliable supply chain and creates the possibility of contract manufacturing for other suppliers. Maintaining API production in-house may keep costs down, reduce time-to-market, and better control quality. DiversiÞcation into bulk manufacturing also provides another revenue stream for those years when patent expiries are fewer and so new (generic) product ßow is reduced. In the following paragraphs, we describe the different approaches companies have taken in the backward integration into APIs. Teva Pharmaceuticals. This Israeli generics company has systematically expanded its own API capacity as the company has grown into the generics industry leader, with revenue of $2.5 billion in 2002. By producing a signiÞcant percentage of its own API needs, Teva is able to avoid many of the sourcing problems that can hobble a large, rapidly growing generics producer. In addition to covering its own requirements, Teva supplies APIs to other generics and specialty pharmaceutical companies in the United States and Europe. This model provides Teva with both additional revenue and market intelligence and helps maintain the quality and competitiveness of internal API supply activities. Ivax. With a large portfolio of generics, Ivax has a truly global reach. For several years, the company has manufactured a signiÞcant portion of its own APIs, mostly in facilities in the Czech Republic. In a move seen commonly in the generic drug industry, Ivax recently achieved further backward integration by purchasing one of its major bulk suppliers. In January 2003, the company acquired Puerto Rican API manufacturer ChemSource, from Chemo Iberica and Quimica Sintetica, for $12.5 million in stock. Sandoz. In May 2003, Novartis consolidated its collection of generics companies (including Geneva, Apothecon, and BASF Generics) under the “Sandoz” brand. In an October 9, 2003, company presentation in London, Sandoz executives emphasized the role of “industrial generics” (including APIs) as one of the three key franchises—along with retail generics and biopharmaceuticals—forming the foundation of its generics strategy. Although Sandoz is already a major supplier of more than 80 different oral antibiotic, hormone, and enzyme intermediates, the company announced its intention to further expand its bulk manufacturing business into more “high-value” APIs.

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Sicor. Formed out of the 1997 merger of Gensia and Rakepoll Holding, Sicor (Irvine, California) has quickly become a leader in the manufacture of injectable generics and their component APIs, both for its own generics business (25 APIs) and for use by other generics manufacturers (36 APIs). Sicor’s production facilities in Italy and Mexico produce a variety of injectable corticosteroids and small-molecule chemotherapeutics. With a biopharmaceutical plant in Mexico and a newly acquired facility in Lithuania, Sicor has moved strongly into the manufacture of both Þnished dose and bulk proteins. On October 31, 2003, Teva Pharmaceuticals announced it had reached an agreement to purchase Sicor for $3.4 billion. Generic Biologics The Generic Pharmaceutical Association calculates that biologics with combined annual U.S. sales of approximately $10 billion face patent expiration by the end of 2007. However, the legal pathway to generic biologics remains unclear in the United States and the major markets of Western Europe. Although payer interest is obviously quite high, the regulatory hurdles are daunting. In addition, manufacturing a bioequivalent biogeneric is a far more complex task than for most small molecules. Nonetheless, biogenerics represents an enormous opportunity for generics manufacturers, which are moving forward with R&D and lining up manufacturing capacity. In many cases, emerging markets have already provided regulatory approval, giving early entrants a head start. In the following paragraphs, we discuss companies that either have already developed generic biologics for emerging markets or are rapidly building capabilities in this segment. Sicor. As noted previously, Sicor has developed both the technical skills and manufacturing capacity for the production of generic biologics. The company’s July 2001 acquisition of Biotechna UAB (Vilnius, Lithuania) gives it a large, modern current good manufacturing practice (cGMP)-compliant facility already actively producing APIs and Þnished injectable generic biologics for emerging Asian, Eastern European, and Middle Eastern markets. Among the products it has launched are recombinant interferon alpha-2b and human growth hormone. Generic versions of Amgen’s Neupogen (Þlgrastim) and Amgen/Johnson & Johnson’s Epogen/Procrit/Eprex (epoetin alfa) have been successfully developed and await clariÞcation of regulatory requirements in the United States and major European markets. Teva’s purchase of Sicor would give the Israeli company a key presence in injectable generics as well as in the newly forming generic biologics market. This addition would clearly add to Teva’s commanding position in the global generic pharmaceutical industry. Pliva. Croatian pharmaceutical company Pliva is attempting to use soon-to-disappear partnership income from the sale of PÞzer’s Zithromax (azithromycin) to rapidly remake itself into a global generics company complete with biopharmaceutical capability. Income from Zithromax (almost $200 million in 2002) has in part funded R&D of generic biologics, including an equivalent of Amgen/Johnson & Johnson’s Epogen/Procrit/Eprex (epoetin alfa). As with several Eastern European

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generics manufacturers, Pliva’s development of generic biologics has been aided signiÞcantly by Croatia’s weak intellectual property laws. Sandoz. As previously noted, Sandoz views generic biologics as one of the three franchises underlying the newly rebranded set of Novartis generics manufacturers. The generic biologics group will build on the company’s long experience in recombinant protein technology, and in the future, it will provide APIs for branded Novartis biologics. Dr. Reddy’s Laboratories. Already a major global player in API and smallmolecule generics, Dr. Reddy’s is moving to position itself in the emerging biologics market. This Indian powerhouse has developed Grastim, a generic form of Amgen’s Neupogen (Þlgrastim), and it claims to have “several” other recombinant proteins and antibodies in its pipeline. While the regulatory environment for generic biologics tries to catch up with that for small molecules, Dr. Reddy’s is building an infrastructure for the manufacture and marketing of diagnostic and therapeutic proteins. ‘‘Difficult’’ Generics Another approach to avoiding the commoditized generics market has been the development of “difÞcult” generics (i.e., generic equivalents of products with some feature complicating manufacture or marketing). By taking on additional developmental complexities, generics companies may create signiÞcant barriers to competition and enjoy a degree of de facto generic exclusivity. Gross margins on such products may be far better than those of “plain vanilla” generics. Perhaps the leading proponent of the difÞcult generic strategy is Barr Laboratories. The company describes its generics strategy as focusing on “products where the company believes that its product selection criteria may result in fewer competitors. Product candidates can include those where Barr has unique research and development skills or those that represent difÞcult-to-manufacture products where Barr has capabilities largely unmatched by competitors. As a result, many of the compounds in development are already without patent protection but currently have no generic competition.” In a prime example of the difÞcult generic strategy, Barr brought the Þrst generic of the oral anticoagulant warfarin to the U.S. market in 1997 and has built it into a $100 million product. U.S. patents on the molecule expired more than 40 years ago, but the drug’s formal classiÞcation as a narrow therapeutic index (NTI) drug effectively protected the branded form (Coumadin, a product of DuPont, DuPont Merck, and now Bristol-Myers Squibb) from competition. Over the years, DuPont Merck had successfully resisted the launch of generic warfarin by other manufacturers, but Barr’s persistence ultimately led to the compound’s launch in 1997. DuPont Merck continued to defend its Coumadin franchise by citing issues of bioequivalence and pharmacist substitutability. In an August 1, 1997, press release, for example, DuPont Merck accused Barr of a “lack of sensitivity to the patient safety issues associated with this narrow

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therapeutic index drug.” Ultimately, Barr was able to demonstrate that its warfarin formulation was equivalent to that of DuPont Merck and overcome the difÞculties of commercializing a generic NTI drug. Barr markets more than a dozen generic oral contraceptive pills (OCPs), and it is the number three company (by 2002 sales) in the United States in the OCP segment. The developmental, manufacturing, and regulatory hurdles faced in the commercialization of hormones such as OCPs can be signiÞcant, but Barr’s franchise is clearly beneÞting from a substantial experience curve. Barr’s focus began with the 2001 launch of generic forms of two key Ortho-McNeil Pharmaceutical products, Ortho-Novum 1/35 and Modicon-28. Since then, several generic OCPs have been added, typically marketed as Barr brands. Most recently, Barr added Seasonale (the Þrst extended-cycle OCP) and has signed a letter of intent to acquire the emergency oral contraceptive Plan B from Women’s Capital (Washington, DC). Perhaps the most sustained example of pursuing the difÞcult generic has been Barr’s pursuit of a generic equivalent of Wyeth’s oral hormone Premarin (conjugated estrogens). Premarin has been marketed in the United States since 1942 without patent protection, but no true generics are on the market today. Conjugated estrogen generics (including one from Barr) were available in the United States until the FDA pulled all generic formulations from the U.S. market in 1991 over concerns of bioequivalence with Premarin. Although related estrogen preparations have been approved since then (e.g., Barr/Duramed’s plant-derived conjugated estrogens Cenestin), Barr has continued to pursue a generic formulation of Premarin. Barr’s most recent attempt was derailed by a September 2003 district court ruling in which Barr’s supplier (Natural Biologics) was found to have misappropriated trade secrets from Wyeth. Barr is not the only company pursuing difÞcult generics. A typical case is the development of isotretinoin generics, Þrst by Mylan and then by Barr and Ranbaxy. Genericization of Roche’s Accutane (isotretinoin) for severe nodular acne has been hampered in part by the daunting regulatory environment and bad publicity surrounding this drug. Because Accutane use during pregnancy can cause fetal malformations, the drug carries a black box warning. In addition, a complex protocol is in place to obtain informed consent, provide patient information, and prevent pregnancy. The drug cannot be prescribed over the phone, and prescriptions are limited to a 30-day supply. In addition to concerns over teratogenicity, reports surfaced linking Accutane to adolescent suicide. Although a causal link has yet to be established, the negative publicity has been signiÞcant. Nevertheless, seeing opportunity in these barriers to entry, Bertek Pharmaceuticals (a subsidiary of Mylan Laboratories) launched its branded generic isotretinoin, Amnesteem, in late 2002. U.S. sales of Amnesteem will likely exceed $100 million. Recently, Barr and Ranbaxy launched generic versions of Accutane, but the barriers to entry in this market continue to protect it against complete commoditization. Other difÞcult molecules remain resistant to generic development, despite the lack of a protecting composition-of-matter patent (e.g., AstraZeneca’s Zoladex (goserelin acetate implant) for prostate cancer).

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Branded Products The most common response to the intense competition and limited growth prospects offered by the commodity generic market has been a movement into branded products. This movement has typically involved either newly developed branded generics or older, smaller marketed products, but it increasingly includes both early- and late-stage NMEs. Although some companies are just beginning to test these waters, others (e.g., Ivax, Teva) started on this path years ago and have aggressively launched a variety of branded products. DiversiÞcation into branded products creates a hybrid pharmaceutical company that combines low-margin commodity generics with higher-margin brands. However, with the notable exception of Novartis/Sandoz, no pharmaceutical company has successfully added a generics business to its existing branded business. Generics manufacturers face signiÞcant organizational challenges as they embrace branded products and the hybrid company model. Generics manufacturers’ R&D budgets are generally much smaller than those of major branded pharmaceutical companies. Any required increases in the R&D infrastructure must be initially funded from the relatively thin margins of the existing generics business. Salesforces must be built up and trained, contracted out, or gained through partnerships. New marketing and brand management experience must be brought in. Thought leaders must be recruited. Ideally, companies should develop some degree of therapeutic focus (Table 1). Nevertheless, if carefully executed, the shift to a hybrid model can be quite successful, yielding new growth prospects, higher margins, and potentially greater earnings consistency. Wall Street appreciates the hybrid’s better pipeline visibility and greater independence from cyclical variations in patent expiries. In the following sections, we discuss several strategies that generics companies are pursuing in their efforts to diversify into branded products. TABLE 1. Therapeutic Focus of Select Generics Manufacturers Company aaiPharma American Pharmaceutical Partners Barr Laboratories

Biovail Ivax Mylan Laboratories Pliva Sandoz Sicor Taro Teva Pharmaceuticals Watson Pharmaceuticals

Therapeutic Area(s) Pain, gastroenterology, critical care Oncology, infectious diseases Women’s health (including oral contraceptives and hormone therapy), infectious diseases, oncology Cardiology, central nervous system Respiratory, oncology Neurology, dermatology, cardiology Infectious diseases Cardiovascular, infectious diseases, central nervous system, gastrointestinal, hormones Critical care, anesthesiology, oncology, biologics Dermatology, pediatrics Central nervous system Nephrology, women’s health, pain

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Branded Formulations of Off-Patent Drugs. Generics companies are developing various types of branded formulations of off-patent drugs: •

•

•

New Oral Formulations. Several generics companies have developed (or are developing) new, branded oral formulations of off-patent molecules, with the goal of receiving three years’ new product exclusivity. Ivax has completed Phase II studies with Paxoral, an oral formulation of the successful intravenous chemotherapeutic agent paclitaxel. In 2002, Andrx launched Altocor, a once-daily formulation of the cholesterol-lowering lovastatin (Merck’s Mevacor). The statin market is crowded, but Andrx has targeted such pricesensitive segments as Medicaid recipients and the elderly. In late 2002, the company also Þled an application for an extended-release metformin (Metformin XT). This product is smaller and potentially easier to swallow than the rival sustained-release Glucophage XR (Bristol-Myers Squibb). K-V Pharmaceutical has taken a different approach to branded generics by using its MicroMask technology to develop better-tasting drugs (e.g., PreCare Chewables prenatal vitamins). New Transdermal Formulations. In April 2003, Watson launched Oxytrol, the Þrst transdermal formulation of oxybutynin for urinary incontinence in the United States. By bypassing the normal oral route of administration, the twice-weekly Oxytrol has the potential for fewer side effects than oral formulations, including J&J’s Ditropan XL. Initial uptake, however, has been slow. New Injectable Formulations. American Pharmaceutical Partners is developing Abraxane (ABI-007), a new formulation of Bristol-Myers Squibb’s blockbuster injectable chemotherapeutic agent Taxol (paclitaxel), which was licensed from American BioSciences. The Phase III Abraxane is based on a novel paclitaxel/albumin nanoparticle, which, theoretically, has the ability to reduce the side effects associated with Taxol’s Cremophor-based vehicle.

Acquired Brands. Other generics companies have taken the generics-branded hybrid model one step further by acquiring marketed brands, typically either niche products or older, “neglected” drugs receiving little promotion. Being already marketed, such drugs eliminate any approval risk. Their small revenue stream, a distraction for a large research-based pharmaceutical company, can be quite appealing to a smaller generics manufacturer for whom $50 million in U.S. sales may be a solid product. This strategy is becoming an increasingly common response to the commoditized oral generics market and its reliance on patent expiries for new product ßow. However, rising demand for these products is likely to increase the cost of similar transactions in the future. Here we review select examples of key players that have acquired branded products: •

Ivax . In April 2002, Ivax acquired the U.S. rights to 3M’s QVAR, a chloroßuorocarbon-free inhaled corticosteroid (beclomethasone dipropionate) for asthma. It had 2002 U.S. sales of $11 million. In October 2003,

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•

•

•

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Ivax acquired additional rights to QVAR in the United Kingdom, Ireland, France, Germany, Netherlands, Finland, Norway, Denmark, and Sweden. Watson. In 2001, Watson licensed nebivolol, a highly selective beta-1 antagonist, from Janssen. The antihypertensive product is available in several European markets but is only in Phase III clinical trials in the United States. Watson continues to move development of this product through the large pivotal trials typical of antihypertensive drug development, but whether nebivolol will be able to successfully compete in the highly genericized beta-blocker market is unclear. In February 2003, Watson acquired U.S. marketing rights to Fiorinal (butalbital, aspirin, and caffeine) and Fioricet (butalbital, acetaminophen, and caffeine) from Novartis for $178 million. These two older drugs (and their respective combinations with codeine) are indicated for tension headache. In 2002, Watson also purchased the U.S. rights to Actigall (ursodiol) from Novartis for $70 million. The gallstonedissolving drug was launched in 1988. Barr. In December 2002, Barr acquired the rights to three products from Wyeth: the glaucoma drug Diamox Sequels (acetazolamide) and two antihypertensives with relatively modest sales—Zebeta (bisoprolol fumarate) and Ziac (a bisoprolol/hydrochlorothiazide combination). Barr and Wyeth also signed agreements regarding several contraceptives, all of which were part of a settlement of litigation between Wyeth and Duramed (with which Wyeth merged in 2001). Biovail . Canadian generics maker Biovail purchased the U.S. rights to the anxiolytic benzodiazepine Ativan (lorazepam) and the oral antianginal Isordil (isosorbide dinitrate) from Wyeth for $130 million. U.S. sales of Ativan and Isordil exceeded $50 million in 2002. Biovail obtained similar rights to Merck & Co.’s once-powerful Vasotec (enalapril) franchise in 2002.

Development of NMEs. The approach with the greatest risks and potential rewards for generics manufacturers is developing NMEs, a strategy that requires adopting in part the business model of major research-based pharmaceutical companies. Implementation of this plan typically involves funding R&D with generics products’ revenue stream. This strategy, despite the risks and complexities of clinical development, has the potential to signiÞcantly strengthen a generics company: a portfolio of NMEs can provide valuable insulation from the competitive pressures on the commodity side of the generics market. In the following paragraphs, we brießy discuss the efforts of two generics companies that have followed this route. •

Teva. If Teva’s success as a hybrid branded-generics company is the envy of the generics industry, Teva’s Copaxone (glatiramer acetate) is a key reason for that envy. With 2002 U.S. sales of $410 million ($524 million worldwide), the multiple sclerosis drug is a major success with doubledigit sales growth. Copaxone is also far from a “one-hit NME wonder.” In September 2003, the company Þled an NDA in the United States for

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rasagiline (a monoamine oxidase-B inhibitor for Parkinson’s disease); it submitted a European Þling one month later. Teva also recently acquired the North American rights to GlaxoSmithKline’s Purinethol (mercaptopurine), an older oral chemotherapeutic without generic competition. The drug, acquired as part of a legal settlement with GlaxoSmithKline over Relafen (nabumetone) litigation, earned $70 million in the United States in 2002. Copaxone, rasagiline (assuming its approval), and Purinethol represent a strong line of NMEs to add to Teva’s 100+ line of generic products. Ivax . Ivax also maintains a strong NME pipeline in both oncology and the central nervous system. The company’s oncology drugs center on a platform of immunotoxins, including TP-38, a Phase I/II immunotoxin targeting the epidermal growth factor (EGF) receptor. Ivax is developing TP-38 primarily for malignant brain tumors. A variety of other immunotoxins as well as angiogenesis inhibitors are in late preclinical testing. The Ivax pipeline also includes the Phase II antiepileptic talampanel.

OUTLOOK In some sense, it is the fear of having to operate in this highly competitive “plain vanilla” generics market that spurs innovation in the biopharmaceutical industry today. In such a volatile commodity market, differentiation between products is virtually impossible and opportunities to add value (and support price increases) are scarce. Other than price, the major points of differentiation are increasingly reliability and operational efÞciency. The lines between branded and generics manufacturers will continue to blur. One by one, surviving generics companies will evolve into more traditional midtier pharmaceutical companies with focused portfolios of drugs targeted at smaller markets and/or enhanced drug delivery. Some will try to achieve this reinvention through organic growth, but for many, mergers and acquisitions will be the fastest route. Quite simply, the goal is to be the next Teva, viewed by many as the paragon of the successful hybrid generics-branded strategy. Along with consolidation will come more and more deal-making surrounding development and marketing, such as the key partnerships between Impax Laboratories and Teva (strategic marketing of controlled-release products), Watson and Cipla (Cipla will develop and supply various generics to Watson), and Eon Labs and Hexal (Eon will market Hexal-developed drugs in the United States). Pure-play commodity generics companies will fail in competition with lowercost Indian and Chinese manufacturers, particularly as these vertically integrated Asian entrants gain experience in U.S. and EU regulatory policy. As these newer competitors gain sales and marketing experience in the major Western markets, they will prove formidable competitors even for the more successful hybrid generics-branded companies. The only relief in sight for U.S. and European generics manufacturers—most likely too late to be real relief—is India’s acceptance of TRIPS patent intellectual property laws. Conforming to TRIPS will reduce India’s typical lead over most Western generics companies, but it will

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do little to alter Indian companies’ favorable cost structure and highly skilled labor force. Finally, biologics are likely to be the “next big thing” for the generics industry as the regulatory pathways are determined over the next Þve years. Companies with signiÞcant experience or production capacity in injectables are likely to be targeted for acquisition by larger generics producers. Sicor, perhaps the most obvious target, is already involved in a potential deal with Teva. Others such as Pliva and generic injectables specialist American Pharmaceutical Partners are likely to become the focus of future deals. Partnerships with specialized Indian producers of generic biologics (e.g., Shantha Biotechnics, Biocon India) are also likely. To successfully compete in the generic biologics market, capacity will assume almost as much signiÞcance as technical expertise.

The Indian Pharmaceutical Industry in Transition: Changes and Opportunities Ahead

SUMMARY The absence of strong patent protection for pharmaceuticals in India has shaped the development of the Indian pharmaceutical industry (IPI)—not only by making the lucrative trade in copy products possible but also by discouraging the industry from conducting original drug discovery. The IPI has focused its efforts on the production of known small molecules and conÞned its R&D efforts to reformulation and process engineering. On April 15, 1994, India signed the General Agreement on Tariffs and Trade, joining the World Trade Organization and becoming a party to the Agreement on Trade-Related Aspects of Intellectual Property Rights (TRIPS). India is therefore required to bring its patent laws into alignment with the standards of TRIPS. The changes will force Indian pharmaceutical companies to Þnd alternatives to their current copy-product business models. In this article, we discuss ongoing reforms in the regulation of pharmaceuticals in India and the challenges and opportunities these reforms present to the IPI. BUSINESS IMPLICATIONS •

Intellectual property and price control reforms in India pose challenges for domestic pharmaceutical companies, which have long beneÞted from a system of weak patent rights and of price controls imposed on foreignbased companies. Growing competition from multinational corporations and increasing limitations on their old business models (such as reverseengineering branded drugs and selling copy products) are forcing the Indian pharmaceutical industry to explore new business models and markets.

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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The largest Indian companies are becoming highly competitive in global generics markets and hope to develop into integrated specialty pharmaceutical companies. Sophisticated Paragraph IV challenges and 505(b)(2) Þlings are allowing these companies to launch generic formulations of branded drugs that once seemed safe from competition because of technical challenges, legal hurdles, or high development costs. In the near term, new drug launches by Indian companies will be limited to reformulations and life-cycle extensions of existing agents because the companies lack the capital needed to develop robust pipelines of novel compounds. With revenues of $6 billion a year, the Indian pharmaceutical industry would have to invest 13% of sales (twice the rate of investment at the most research-oriented Indian companies and nearly ten times the local industry average) to cover the cost of developing a single new chemical entity each year. As the Indian pharmaceutical industry grows and increasingly presses its cost advantages, Big Pharma will be compelled to partner with Indian companies, acquire them, build its own Indian facilities, or Þnd other means to lower its own costs.

INTRODUCTION Although India produces approximately 8% of the world’s drug supply and is the fourth-largest producer of pharmaceuticals by volume, it has historically focused on low-margin products, and its pharmaceutical companies have remained niche players in the global pharmaceutical market. In 2002, sales by the Indian pharmaceutical industry (IPI) totaled approximately $6 billion. By comparison, PÞzer’s worldwide sales of atorvastatin (Lipitor) alone reached $8.6 billion that year. The IPI has principally produced bulk active pharmaceutical ingredients (APIs), which are exported to international branded and generics drug companies for formulation, and low-cost copies of branded drug formulations, which are sold on the Indian market and exported to other countries that do not recognize the patents held on those products by Big Pharma companies. The absence of strong patent protection for pharmaceuticals in India has shaped the development of the IPI—not only by making the lucrative trade in copy products possible but also by discouraging the industry from conducting original drug discovery. In the absence of patent protection, Indian companies could not recoup the extensive costs of such R&D because competitors would immediately copy their products and drive prices down to commodity levels. Consequently, the IPI has focused its efforts on the production of known small molecules and conÞned its R&D efforts to reformulation and process engineering. Collectively, the IPI invests only an estimated 1.9% of its sales in R&D. By comparison, in 2003, Big Pharma invested approximately 14.2% of sales in R&D. Three major trade organizations represent Indian pharmaceutical companies. The Indian Drug Manufacturers’ Association (IDMA) was formed in 1961 to defend the interests of indigenous drug companies as major multinational

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corporations (MNCs) began to compete in the Indian market. The Organization of Pharmaceutical Producers of India (OPPI) was created in 1965 to represent the interests of both local Indian companies and the Indian subsidiaries of MNCs. The Indian Pharmaceutical Alliance (IPA) was formed in 1999 by eight of the largest domestic pharmaceutical companies and is principally concerned with promoting domestic R&D efforts. The IPA has 11 members (Alembic, Cipla, Dr. Reddy’s Laboratories, Lupin, Nicholas Piramal, Ranbaxy, Sun Pharmaceutical Industries, Torant Pharmaceuticals, Unichem Laboratories, Wockhardt, and Zydus Cadilla) and accounts for more than 90% of pharmaceutical R&D spending in India. In this article, we discuss ongoing reforms in the regulation of pharmaceuticals in India and the challenges and opportunities these reforms present to the IPI. IMPACT OF NATIONAL PATENT LAW ON THE INDIAN PHARMACEUTICAL INDUSTRY India’s historically weak system of intellectual property protection has beneÞted the IPI. Although India has had some form of patent rights since 1856, the Indian Patents Act of 1970 explicitly prohibited the granting of pharmaceutical product patents to facilitate broad access to medications and support the growth of the nascent domestic pharmaceutical industry. The Patents Act states that no patent shall be granted for “substances intended for use, or capable of being used, as food or as medicine or a drug” but only for the “methods or processes” of manufacturing them. In addition, the term of such pharmaceutical or chemical process patents was limited to the shorter of seven years from the date the patent application was Þled or Þve years from the date the patent was granted, while the term of patents on any other invention in India was fourteen years from the Þling date. The limitations imposed in 1970 have been a boon to the IPI, which has developed great expertise in reverse engineering drugs developed by Big Pharma and either inventing around the Indian process patents that cover them or simply waiting Þve years for those patents to expire. In many cases, these short-lived Indian patents will have already expired by the time a new drug completes clinical testing and receives its Þrst regulatory approvals. In the absence of product patents, the IPI has manufactured and sold copies of branded pharmaceuticals to the domestic Indian market and exported them to other countries where the drugs lack patent protection. On April 15, 1994, India signed the General Agreement on Tariffs and Trade (GATT), joining the World Trade Organization (WTO) and becoming a party to the Agreement on Trade-Related Aspects of Intellectual Property Rights (TRIPS). India is therefore required to bring its patent laws into alignment with the standards of TRIPS. These requirements include a minimum patent life of twenty years from the application Þling date and the availability of patents not only for processes but also for pharmaceutical products and methods of using them. These changes will force Indian pharmaceutical companies to Þnd alternatives to their current copy-product business models.

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Although TRIPS went into effect in 1995, developed countries were given one year to ensure that their laws and practices conformed to its requirements, and developing countries were given longer grace periods to establish their intellectual property protection systems. India’s grace period ended on January 1, 2005, so the Indian Patent OfÞce must now accept and review pharmaceutical product, process, and use patents and must extend the lifetime of all granted patents to twenty years from the Þling date. Although India is not required to grant pharmaceutical product patents until the grace period has elapsed, the “mailbox” provision of TRIPS requires India to accept applications for the newly allowed patents Þled from 1995 onward. During the grace period, these applications are not reviewed or granted, but they must be reviewed at the end of the grace period and, as a stopgap measure, India is required to grant exclusive marketing rights (EMRs) over these inventions to the Þler for Þve years from the date the drug receives marketing approval or until the application is Þnally granted or rejected. EMRs must be available for any drug covered by an Indian patent application Þled through the mailbox system if a corresponding application has been Þled and a patent granted in any other WTO member country after January 1, 1995. Consequently, there is already a large backlog of pharmaceutical patent applications on Þle with the Indian Patent OfÞce that effectively came into force in January 2005 as though a patent had already been granted. However, these EMRs apply only to actual commercialization of the drugs and do not prevent companies from manufacturing or conducting research with these drugs. India steadily implemented the changes necessary to comply with TRIPS in advance of the 2005 deadline. The 1999 amendment to the Patents Act included a provision instituting EMRs and setting up the mailbox system retroactive to patent applications Þled in other WTO countries since January 1995. The 2002 amendment to the Patents Act ofÞcially extended the term of Indian patents to twenty years from the Þling date. The new length applies to all patents still in effect at that time as well as to all new patent applications. Provisions for pharmaceutical product patents have remained conspicuously absent from these amendments, however, and India remains on a U.S. Trade Representative’s Priority Watch List owing to concerns over such remaining inconsistencies with international law. PRICE CONTROLS IN THE INDIAN MARKET The Indian government’s price control scheme has reinforced its historically weak patent policies by limiting the attractiveness of the Indian market for MNCs while providing competitive advantages to domestic companies. The Drug Price Control Order (DPCO) was introduced with the Patents Act in 1970 and originally placed limits on the proÞtability of companies selling pharmaceuticals in India. Since then, the Indian government has been slowly but steadily easing those restrictions. A 1979 revision shifted the emphasis to speciÞc caps on the prices of 370 widely used drugs. Subsequent revisions in 1987 and 1995 further reduced the scope of

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DPCO price controls to 74 drugs. Those 74 drugs, however, constituted 40% of pharmaceutical sales in India. This policy has depressed the value of the Indian market and made it difÞcult for MNCs to achieve attractive margins even on products that are covered by Indian patents and enjoy market exclusivity. The DPCO has deterred MNCs from launching many new drugs in India, leaving the market open for domestic companies. In addition to broadly reducing the attractiveness of the Indian market for MNCs, the criteria and exemptions used to select the price-controlled drugs were designed to give competitive advantages to domestic companies by providing ten-year exemptions for products that result from domestic R&D (on the active ingredient, formulation, or delivery system). Of course, this policy does not apply to the international blockbusters that are the cornerstone of the MNCs’ business, but the policy’s broad deÞnition of qualifying R&D also allows domestic companies to earn considerably more on reformulations of those drugs than the MNCs can earn on the original formulations. The Indian government’s 2002 pharmaceutical policy further reduced the scope of these price controls to 28 drugs that collectively account for 19% of the domestic pharmaceutical market. Although this policy change constitutes a considerable liberalization of price controls in India, the new policy is still based on a system of exemptions that favors products from domestic companies. New drugs developed through domestic R&D (whether novel drug discovery or the development of new formulations or production processes that circumvent the patents of MNCs) and patented under the 1970 Indian Patents Act are exempt from price controls until the expiry of their patents. Because such patents for novel drugs will generally have expired before the product reaches the market, these price controls effectively beneÞt only the copy products developed through process reengineering by local companies. If the product is also manufactured exclusively in India, the exemption is extended to Þfteen years from the date of commercialization. In contrast, new drugs developed through foreign R&D are exempt from price controls for only Þve years. Consequently, under the 2002 policy, MNCs still have considerably greater exposure to DPCO price controls than do local companies. OTHER LIMITS ON THE INDIAN PHARMACEUTICAL MARKET The Foreign Exchange Regulation Act of 1973 (FERA) and its replacement, the Foreign Exchange Management Act of 1999 (FEMA), imposed limits (generally less than 40%) on the equity stakes MNCs could hold in their Indian ventures, further diluting the value of Indian operations for foreign-based companies. Consequently, the domestic market share of MNCs in India has fallen steadily from 75% in 1971 to approximately 30% today, allowing the domestic industry to ßourish. The low purchasing power of the Indian population and the absence of comprehensive medical insurance further suppress Indian drug prices, even for products that are not subject to governmental price controls. Although India has one of

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the largest populations in the world (according to the 2001 census, in excess of 1 billion people), it also has one of the highest poverty rates (average per capita income is $400 per year), and India’s total health care spending in 2003 was less than $30 billion (approximately $26 per capita). Although the Indian government and some large private employers provide limited health insurance beneÞts, estimates suggest that only 2.2 million people in India have health insurance. Consequently, the largest single pharmaceutical brand in India today, Becosule (vitamin B complex), has annual sales of only $20 million. Even with the rise of market exclusivities and the fall of price controls, the underlying economic conditions in India will temper the potential of the Indian drug market for the foreseeable future. COMPETITIVE POSITION OF THE INDIAN PHARMACEUTICAL INDUSTRY The IPI has grown rapidly since 1970 by reverse engineering Big Pharma products that are unpatentable in India. In the process, it has built up considerable expertise in chemical synthesis and processing, API production, and formulation. These capabilities, coupled with considerable production cost advantages, are allowing Indian companies to expand rapidly into the international generics pharmaceutical market by bringing their existing products into regulated markets once the relevant patents have expired. Some of the largest Indian companies aim to expand into a specialty pharmaceutical role by reformulating existing drugs for improved clinical proÞles or niche applications. Current blockbusters and most drugs being launched in the near term are largely covered by patents Þled before 1995. Thus, the impact of the new intellectual property rights system will not come to bear on the portfolios of Indian pharmaceutical companies for several years. Although the commencement of patent protection and EMRs for pharmaceutical products in 2005 will have little impact on the IPI’s ability to market copies of current blockbusters, these measures will preclude it from reverse engineering newer drugs. This new restriction will cause the IPI’s copy-product pipelines to gradually dry up, forcing it to explore new business models. Compounding this threat are the availability of market exclusivity in India and the liberalization of price controls as well as other trade regulations, all of which will make India a more attractive market and boost competition from MNCs for domestic sales. As a result, many of the largest Indian companies are targeting API and retail formulation opportunities in foreign markets that leverage the skills and resources they have already developed. The industry’s key strategic assets are its low production costs and high concentration of technical skill gained through decades of reverse engineering and reformulating pharmaceuticals. India has a strong labor force with advanced degrees in medicine and pharmacology, high levels of complex chemistry and formulations skills, and low salary requirements. According to a recent survey conducted by Morgan Stanley, the cost of employing a chemist in India is between

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one-eighth and one-Þfth the cost of employing a comparable individual in the United States. Similarly, the cost of building a manufacturing facility in India is 20–25% of such costs in the United States and European Union. Although the IPI has clear cost advantages over the global branded and generics pharmaceutical companies, its expansion will be constrained by its relatively modest resources for investment. The revenues and margins on its generic and copy-product businesses are modest compared with those common among branded pharmaceutical companies. Furthermore, the IPI’s cost advantages are likely to erode as U.S. and European companies build manufacturing facilities of their own in India or partner with Indian companies for API production or even the production of Þnished formulations. U.S. and European generics companies (free from the conÞdentiality concerns that have made Big Pharma reticent to partner with Indian companies) already source many of their bulk active ingredients from Indian facilities. Expansion from API and generic drug production into higher-margin specialty pharmaceutical and biogenerics businesses will present additional challenges for the IPI. With its historical focus on reverse engineering and manufacturing small molecules, its expertise in molecular biology has not kept pace with its skills in basic chemistry and pharmacology. The production of biogenerics and forays into new drug discovery will require a signiÞcant expansion in these abilities. NEW STRATEGIES FOR GROWTH Launch Generic Drugs in the Regulated Markets The most readily accessible growth strategy for the IPI is to launch its existing portfolio of copy products in the major regulated markets as generics once the market exclusivities of the innovator companies expire. This strategy leverages the IPI’s existing skills, resources, and products without requiring large investments in new R&D, marketing, or other aspects of the pharmaceutical business with which it has limited expertise. This strategy should serve as an effective means of maintaining growth in the short term while laying the groundwork for more ambitious long-term changes to the IPI’s business models. With well-established development and manufacturing resources, the major hurdles for the IPI’s success in the international generics business are regulatory in nature. In the past few years, Indian companies have made progress in bringing their manufacturing facilities up to the standards required by U.S. and European regulatory authorities. They have also dramatically increased the volume of their generic approval Þlings and secured numerous marketing agreements with U.S. and European generics distributors. The launch of the Þrst crop of Indian generics in the Western markets has drawn attention to Indian companies’ complex chemistry skills, but more importantly, it has established their ability to navigate the legal and regulatory challenges that face generics players in the major pharmaceutical markets. The Þrst step for generics companies in building up a presence in the major markets is to Þle ANDAs with the FDA and to follow the analogous “abridged

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procedure” with the European Agency for the Evaluation of Medicinal Products (EMEA) for regulatory approval of their off-patent copy products. The surge in pharmaceutical patent expiries over the past few years has provided numerous such opportunities. Indian companies can then build on the regulatory expertise and distribution infrastructure they develop through these efforts to develop new generic drugs. Their large production capacity and base of technical expertise will make Indian companies formidable competitors to the large Western generics Þrms in developing and marketing generic formulations. However, the frequency of major drug patent expiries is expected to fall after 2007, and more aggressive approaches will be needed to keep portfolios and profits growing. Indian companies have increasingly taken advantage of the Paragraph IV challenge process in the United States. In a Paragraph IV challenge, the generics company Þles an ANDA for a drug that is still patent-protected but challenges the validity or scope of that patent. If the challenge is upheld in court, the FDA may approve the generic formulation immediately rather than wait for the original patent to expire. Indian companies have received the 180-day market exclusivity for being the Þrst generics company to successfully Þle Paragraph IV challenges on three drugs to date: Dr. Reddy’s ßuoxetine (Eli Lilly’s Prozac), Ranbaxy’s cefuroxime (GlaxoSmithKline’s Ceftin), and Ranbaxy’s amoxicillin/clavulanate (GlaxoSmithKline’s Augmentin). In January 2003, Ranbaxy Þled an ANDA with a particularly creative and aggressive Paragraph IV challenge to the key patents covering PÞzer’s atorvastatin (Lipitor). One patent covers a racemic mixture of atorvastatin isomers, and a subsequent patent claims the speciÞc isomer that is the active ingredient used in Lipitor. Ample legal precedent has been established for receiving a patent on a particularly useful speciÞc member of a group even after the whole group has been disclosed in a previous patent. Many speciÞc isomer patents have been upheld even in light of earlier racemate patents on the grounds that the racemate patent does not indicate which speciÞc isomer will be a therapeutically useful product. However, PÞzer applied for and received a Hatch–Waxman market exclusivity extension on the earlier racemate patent, a process that required PÞzer to stipulate that the patent covered the approved version of its drug. Ranbaxy argues that this stipulation runs contrary to the claims of novelty that PÞzer made in subsequently Þling for the speciÞc isomer patent. Ranbaxy thus claims that PÞzer defrauded the patent ofÞce by simultaneously arguing that the earlier patent did cover Lipitor (to extend that patent’s term) and that it did not cover Lipitor (to establish the novelty needed to get the second patent approved). Although this complex case is likely to remain in the courts for many more months, the circularity of Ranbaxy’s argument will make it difÞcult for PÞzer to defend both patents. Sophisticated challenges like this one have demonstrated that the IPI has developed legal and regulatory expertise to match its technical skills. By pursuing generic drug opportunities with high legal and technical barriers, Indian companies are focusing on products with less competition (and thus higher margins) than traditional commodity generics.

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Some Indian pharmaceutical companies are beginning to explore the Þeld of biogenerics. To date, two major factors have limited the growth of this Þeld. First, the FDA and EMEA have not yet developed a framework for approving biogenerics because the appropriate criteria for demonstrating bioequivalence among biologicals are still hotly debated. Second, even without the need for the original drug discovery steps, the cost of developing and manufacturing biologicals is considerably higher than that of small molecules, making it difÞcult for a generics company to earn acceptable proÞts while offering the price advantages that the market will expect from generics. Recently, however, both the FDA and EMEA have made commitments to develop approval processes for biogenerics. The cost advantages associated with Indian labor and production facilities leave the IPI well positioned to pioneer this business model. Several Indian companies—notably Biocon, Wockhardt, and Shantha Biotech —have established research programs to develop biogenerics. Early efforts are focusing on insulin, human growth hormone, and epoetin alfa (Amgen’s Epogen, Johnson & Johnson’s Procrit/Eprex). Insulin and human growth hormone have attracted the attention of Indian generics companies because these therapies’ patents have expired, and they were originally approved as new chemical entities (NCEs) through the traditional new drug application (NDA) process rather than the later-developed biologics license application (BLA) process for biologicals. Consequently, it is possible to Þle for approval of generic versions of these two agents using the existing ANDA process. In August 2003, Wockhardt launched a recombinant human insulin in India, making it the fourth company worldwide to develop and market a version of the drug. Wockhardt is likely to be the Þrst generics company to secure approval for recombinant insulin in the major markets because it has several good manufacturing practice (GMP)-certiÞed manufacturing plants, has already Þled for a U.S. patent on its yeast-based production process, and has indicated that it is pursuing regulatory approval in the United States and major European markets. With global sales of more than $6 billion and European patent expiry in 2004, epoetin alfa has also attracted the attention of would-be biogenerics companies and is likely to be the Þrst agent approved through any forthcoming biogenerics approval procedure. Wockhardt has already launched a recombinant human erythropoietin in India and is likely to be among the Þrst generics competitors to Amgen and Johnson & Johnson in the regulated markets. Invest in R&D and Develop Branded Specialty Pharmaceuticals The export of generics into regulated markets is likely to be the main growth area for the IPI for many years after the 2005 patent law changes, but the largest Indian companies intend to climb further up the pharmaceutical value chain. New generics launches will increasingly be paired with low-risk NDA Þlings for reformulations or follow-on compounds of existing drugs. The IPI’s expertise in process reengineering could make it a major player in the specialty pharmaceuticals arena, where the technical hurdles of extended-release and injectable formulations have so far constrained competition.

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Dr. Reddy’s Laboratories and Ranbaxy are at the forefront of Indian companies hoping to develop into fully integrated pharmaceutical companies (FIPCOs) with large R&D operations and rich pipelines of NCEs. The greatest challenges facing Indian companies looking to reposition themselves as FIPCOs are the long development times associated with new drug discovery and the large R&D investments required to bring even a single NCE to the market. To pursue the FIPCO model in the near term, Indian companies must build and promote proprietary product portfolios through in-licensing and reformulation strategies until their nascent drug discovery efforts begin to bear fruit. Dr. Reddy’s Laboratories, Ranbaxy, Cipla, and Wockhardt have all established development programs that leverage their existing formulation expertise to develop life-cycle extensions of existing drugs (e.g., extended-release capsules, depot injections, novel delivery systems). These efforts allow them to build up their R&D facilities and gain experience launching new drugs while mitigating the risks and pipeline gaps associated with new drug discovery. Dr. Reddy’s Laboratories has been leading the charge into the specialty pharmaceutical niche by developing proprietary modiÞcations of existing drugs rather than true generics. In addition to developing extended-release formulations of existing drugs or pairing them with novel delivery systems, Dr. Reddy’s has been exploring the Þeld of “non-AB-rated” generics. Such products are variants that are not precisely bioequivalent to the original drugs but are similar enough to rely at least in part on clinical data already collected and Þled by the innovator company. Because they are not bioequivalent, these drugs cannot be approved through the ANDA procedure but can make use of the 505(b)(2) Þling process (named after the section of the U.S. Food, Drug, and Cosmetics Act that established it). These Þlings require some original clinical trial work but less than would be required for a traditional NDA because they are allowed to refer to clinical trial data Þled by other companies on earlier formulations in previous NDAs. Although these specialty products require a much greater level of R&D than classic generics, this burden provides a measure of market exclusivity and prevents the product from becoming a commodity too quickly. Consequently, these products can maintain prices more in line with the original branded drugs, thus constituting a class of “branded generics.” In November 2003, Dr. Reddy’s Laboratories became the Þrst Indian company to receive FDA approval of a 505(b)(2) application. The product, amlodipine maleate (AmVaz), is a non-AB-rated variant of PÞzer’s amlodipine besylate (Norvasc). As a different salt, amlodipine maleate was deemed to fall outside some key PÞzer patents while being similar enough to amlodipine besylate to rely on some of the original PÞzer research demonstrating efÞcacy and safety in the treatment of hypertension and angina. Dr. Reddy’s did not have an opportunity to launch amlodipine maleate because an appeals court ruled in February 2004 that amlodipine maleate falls within the scope of PÞzer’s patent, overturning the original Þnding of noninfringement. However, Dr. Reddy’s is not prepared to abandon this regulatory strategy and has stated that it has already Þled a second 505(b)(2) application for an undisclosed drug.

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If successful in bringing non-AB-rated generics to the market, Dr. Reddy’s hopes that its price advantage and the publication of head-to-head studies showing equivalence to the branded drugs will not only drive the generics’ uptake among new patients but also draw market share through substitutions among patients using the original brands. A single success in navigating the legal and regulatory hurdles could open the door for a ßood of non-AB-rated Indian generics of Big Pharma products that were thought to have many years of market exclusivity left. Portfolios that combine generics and branded generics will allow the IPI to maintain revenue growth while it ramps up its R&D capacity. The long-term success of companies pursuing the specialty pharmaceutical or FIPCO model will, however, depend on their ability to develop their own patent-protected novel drug candidates. Ranbaxy and Dr. Reddy’s have made by far the greatest commitments in the IPI to in-house drug discovery. These two companies have already had some success in drug discovery and have even outlicensed novel drugs to Western pharmaceutical companies. Table 1 provides information on novel drug candidates in clinical development by the IPI. Nevertheless, the IPI has limited capital available for R&D investment. With total industry revenues of $6 billion per year and current estimates of approximately $800 million in R&D investment required to bring a single new drug to the market, the IPI would collectively need to invest 13% of its revenues to bring one new drug to market each year. The IPI currently falls far short of this R&D investment target: Indian R&D leaders Ranbaxy, Dr. Reddy’s, and Wockhardt invest only 4–8% of annual revenues in R&D (Table 2). (By contrast, the largest global pharmaceutical companies invest 12–16% of their revenues in R&D. In 2002, PÞzer and GlaxoSmithKline’s R&D investments totaled $5.2 billion and $4.4 billion, respectively—amounts that rival the entire IPI’s sales for that year.) To be competitive in new drug discovery, the IPI will need not only to drastically increase the fraction of its capital invested in R&D but also to pool its resources through research collaborations and industry consolidation. Among Western pharmaceutical companies, thin pipelines have been one of the major drivers of mergers and acquisitions. The IPI is likely to mirror this behavior over the next few years, leaving a handful of competitive specialty pharmaceutical companies. This trend of consolidation among Indian companies has already begun, as exempliÞed by the following: • • •

Dr. Reddy’s Laboratories merged with Cheminor Drugs and American Remedies in 2000. Wockhardt acquired CP Pharmaceuticals in 2003. Nicholas Piramal (itself the result of the 1988 merger between Nicholas Laboratories and Piramal Enterprises) has acquired several companies, including Roche’s Indian subsidiary, Roche Products, in 1993; Sumitra Pharmaceuticals in 1995; and Boehringer Mannheim India in 1996.

TABLE 1. Novel Drug Candidates in Clinical Development by the Indian Pharmaceutical Industry Company Dr. Reddy’s Laboratories Dr. Reddy’s Laboratories Dr. Reddy’s Laboratories Dr. Reddy’s Laboratories Ranbaxy

Compound Balaglitazone DRF-1644 DRF-4832 DRF-1042 RBx-2258

Ranbaxy

RBx-7796

Ranbaxy

RBx-9001

Wockhardt

WCK-771

a PPAR, peroxisome proliferator-activated receptor. b VLA-4, very-late-antigen 4.

Mechanism of Action PPARa

gamma agonist Topoisomerase I inhibitor PPAR alpha/gamma agonist Topoisomerase I inhibitor Endothelin-receptor antagonist VLA-4b antagonist Alpha-1 adrenoceptor antagonist Fluoroquinolone antibiotic

Target Indications Type II diabetes

Phase of Development

Benign prostatic hyperplasia Allergic rhinitis, asthma Benign prostatic hyperplasia

Phase II clinical Phase I clinical Phase I clinical Phase I clinical Phase II clinical Phase II clinical Phase I clinical

Bacterial infection

Phase I clinical

Solid tumor Atherosclerosis, syndrome X Cancer

Comments Licensed to Novo Nordisk in 1997

Licensed to Schwartz in 2002

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TABLE 2. R&D Investment by the Indian Pharmaceutical Industry, 2003 Company Dr. Reddy’s Laboratories Wockhardt Ranbaxy Sun Pharma Lupin Divi’s Laboratories Orchid Cadila Healthcare Aurobindo Nicholas Piramal Matrix Cipla

F2003 Sales (U.S. $millions)

F2003 R&D Expenses (U.S. $millions)

Percentage of Sales Invested in R&D

373 145 790 164 211 51 108 193 229 303 82 300

28 9 35 6 5 1 3 4 3 3 0 0

8 6 4 4 2 2 3 2 1 1 0 0

Notes: Financial year 2003 refers to the year ended March 31, 2003, for all companies except Ranbaxy and Wockhardt, for which the data reflect the year ended on December 31, 2002. An average F2003 exchange rate of Rs 48.39/US$ has been used in this table. Numbers have been rounded.

Supply Cost-Effective Contract Work for Global Pharmaceutical Companies A third option for the IPI is to avoid the FIPCO model and reposition itself as a cost-effective source of contract work for the global pharmaceutical industry. Several Indian companies, including Nicholas Piramal and Divi’s Laboratories, have already begun to position themselves as contract manufacturers of bulk APIs or Þnished formulations. Big Pharma has so far been reticent to contract with Indian Þrms for API manufacturing and formulation because of the risk that conÞdentiality breaches could expedite the launch of Indian copy products. However, the advent of strong pharmaceutical patent protection in India in 2005 greatly reduces this risk and may lead to an explosion of outsourcing deals. The increasing price pressures from companies such as Dr. Reddy’s and Ranbaxy may further drive Big Pharma and international generics companies to pursue the cost advantages of Indian manufacturing. Indeed, the success of some Indian companies pressing their cost advantages to compete as FIPCOs in the regulated market will open the niche for smaller Indian Þrms to offer the cost advantages of contract manufacturing in India. One critical requirement for this strategy (as well as for those discussed previously) is a large number of manufacturing facilities that have been inspected by Western regulatory authorities and certiÞed for compliance with current good manufacturing practices (cGMP). Although the FDA has approved several facilities to supply APIs and generic formulations to the United States, most Indian facilities are still geared toward domestic production and export to unregulated markets. The FDA’s large facilities-inspection backlog and the logistical complications of inspecting plants in foreign countries could be the major limiting factors on the IPI’s production capacity for the regulated markets over the next few years.

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Indian Þrms may also pursue business as contract research organizations. The cost of conducting clinical research in India is approximately one-tenth the cost in the United States, and the large population of treatment-na¨õve patients for most major indications could greatly facilitate recruitment and enrollment. However, relatively little clinical testing has been conducted in India to date. The IPI has had little need for clinical trial work because its novel drug development and testing efforts have been limited. MNCs have not exploited India’s potential as a low-cost clinical trial site owing to the same conÞdentiality concerns that have restrained their outsourcing of production and fears that the FDA and EMEA may be reluctant to accept Indian trial data. However, the FDA has begun to accept bioequivalence studies conducted in India—several approved ANDA submissions (notably Ranbaxy’s Þling for generic ßuoxetine) have relied on Indian trial data. This increasing receptivity from regulatory authorities combined with the improvements in patent protection may encourage MNCs to outsource more clinical research to the IPI. Consolidate Positions in the Domestic Market A fourth and more modest approach for Indian companies is to strengthen their grip on the domestic market. Owing to the remaining price controls and the time that will elapse before patents Þled after 1995 begin to cover major drug products, the level of competition the IPI will face from MNCs for control of the Indian market is likely to be modest in the near term. Many Indian companies that lack the resources to develop branded or generic products in foreign markets may focus on defending their current domestic role and extend this position through in-licensing or marketing collaborations. Several Indian companies, including Nicholas Piramal and Sun Pharma, have consolidated their positions in the domestic market by acquiring key local brands from smaller Indian Þrms. The acquisition of speciÞc brands and whole Indian companies to build portfolios focused on key therapeutic areas may allow midtier Indian companies to maintain revenue growth with little change to their fundamental business models. Successful domestic portfolios will focus heavily on antibiotics and other treatments for the tropical diseases that remain prevalent in India. Although companies focused on domestic opportunities cannot reverse engineer products patented after 1995, they can proÞt from those products by offering their local marketing and distribution infrastructure to MNCs that wish to sell their products in India but are not prepared to invest in building up their own presence there. For example, Cadilla Healthcare has already entered into agreements with Schering AG and Boehringer Ingelheim to be their preferred marketing partner in India. KEY INDIAN PHARMACEUTICAL COMPANIES Cipla Established in 1935, Cipla has attracted recent international attention for exporting the generic version of the anti-AIDS drug zidovudine (AZT) through Medecins

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THE INDIAN PHARMACEUTICAL INDUSTRY IN TRANSITION

Sans Frontieres (Doctors Without Borders) to more than 35 developing countries at retail prices of $1 per day. The company has a broad portfolio of generics and is a major exporter of CFC-free inhalant formulations. Additionally, all of Cipla’s bulk drug manufacturing facilities are FDA-approved and the company has become the preferred supplier of APIs to several Western generics companies. Dr. Reddy’s Laboratories Founded in 1984, Dr. Reddy’s became the third-largest pharmaceutical company in India after its merger with Cheminor Drugs and American Remedies in 2000. Dr. Reddy’s has made several efforts to build up its presence outside India. In 2001, it became the Þrst Indian drug company to win a 180-day market exclusivity for a successful Paragraph IV challenge and the Þrst to be listed on the New York Stock Exchange. In 2002, the company acquired U.K.-based BMS Laboratories and Meridian Healthcare. In May 2004, Dr. Reddy’s announced the acquisition of U.S.-based Trigenesis Therapeutics. Dr. Reddy’s has three novel drugs in clinical development. Nicholas Piramal India Nicholas Piramal was formed by the 1998 acquisition of Nicholas Laboratories by Piramal Enterprises. It has acquired several Indian companies since then to strengthen its position as a provider of retail formulations for the domestic market. Exports constitute only 4% of the company’s sales and consist largely of on-patent drugs produced under contract for the global companies that own them. Ranbaxy Laboratories Ranbaxy was established in 1962 and is now the largest pharmaceutical company in India and the ninth-largest generics company in the world. With manufacturing facilities in seven countries, the company produces more than 90 APIs and several hundred retail drug formulations. Ranbaxy has four novel drugs in clinical development and has begun to partner with larger Þrms for drug discovery. It developed, licensed, and is manufacturing Bayer’s once-a-day formulation of ciproßoxacin (Cipro) and, in October 2003, entered into a research collaboration for drug discovery and development with GlaxoSmithKline. Sun Pharmaceutical Industries Established in 1983, Sun has grown rapidly through a series of domestic acquisitions. Sun maintains three research facilities, one of which was established in 2000 to focus on the development of NCEs. The company’s pipeline includes a non-AB-rated generic luprolide depot formulation. Sun acquired Detroit-based Caraco Pharmaceutical Laboratories in 1996 to facilitate the launch and distribution of generics in the United States and has Þled ten ANDAs to date.

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149

Wockhardt Wockhardt was spun off from the larger Wockhardt Life Sciences in 1999 and is focused entirely on pharmaceuticals. The company announced a focus on R&D in 2001 and is developing depot injection and extended-release tablet technologies. It has a novel ßuoroquinolone antibiotic in clinical development and has already marketed a recombinant human erythropoietin formulation in India. The acquisition of U.K.-based CP Pharmaceuticals and Wallis Laboratories has made Wockhardt one of the ten largest generic drug companies in the United Kingdom. Wockhardt has also acquired Germany’s Rhein Biotech, with which it has jointly developed a hepatitis B vaccine. OUTLOOK As Indian pharmaceutical companies adjust to India’s stringent new patent laws that came into effect in 2005, industry consolidation, increased R&D investment, and the development of foreign marketing and distribution channels will be the principal trends reshaping the industry. Improved health care infrastructure and reimbursement, as well as a growing economy, will increase the overall value of the domestic market in the next few years, allowing modest revenue growth among both the established domestic companies and the increasingly competitive MNCs. Some smaller Indian companies may rely on the increasing utilization of older, unpatented drugs to drive their revenue growth, but the larger companies have been positioning themselves to exploit additional growth opportunities both in India and abroad. The largest and most aggressive members of the IPI are rapidly making the transition to global generics businesses and hope to develop into fully integrated, research-based specialty pharmaceutical companies. This transition will be slow and difÞcult because the IPI will lack the capital required to fund full drug discovery pipelines for the foreseeable future. Several of the largest Indian companies have been acquiring U.S. and European generics operations to facilitate their entry into those markets. Recent examples include Ranbaxy’s acquisitions of Aventis’s French generics operation and Basics (Bayer’s German generics Þrm), Dr. Reddy’s acquisition of the U.K.-based Meridian Healthcare and BMS Laboratories, and Wockhardt’s acquisition of Accumed in the United States. In most cases, Indian facilities supply the bulk active ingredients while the local facilities produce Þnished formulations that comply with local regulations and oversee distribution. The principal impact of these companies on global pharmaceutical markets over the next few years will be twofold. First, the production-cost advantages of the IPI will exert downward pressure on the price of bulk active ingredients and formulations. Second, sophisticated Paragraph IV challenges and 505(b)(2) Þlings by Indian companies will bring generic competitors to branded drugs that once seemed safe from competition because of technical challenges, legal hurdles, or high development costs.

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THE INDIAN PHARMACEUTICAL INDUSTRY IN TRANSITION

Patent reforms will reduce future copy-product opportunities, and competition for domestic sales among Indian companies lacking the resources to expand into foreign markets will lead to further consolidation. The midtier companies that survive are likely to become valuable partners for global pharmaceutical companies seeking cost-effective sources of manufacturing and clinical trial contract work, as well as marketing and distribution partners for their products within the Indian market. As the Indian pharmaceutical industry grows and increasingly presses its cost advantages, Big Pharma will be compelled to partner with Indian companies, acquire them, build its own Indian facilities, or Þnd other means to lower its own costs.

Spotlight on the Australian and New Zealand Pharmaceutical and Biotechnology Industries

SUMMARY With a combined population of approximately 24 million people, Australia and New Zealand are only slightly larger than the state of Texas and just under half the size of the United Kingdom. Nevertheless, both countries have high standards of living and operate government-funded monopsony systems that limit consumer costs for pharmaceuticals. In 2000–2001, the Australian government’s total health expenditure was approximately $20.2 billion, of which an estimated $2.3 billion was allocated to pharmaceuticals; the New Zealand government’s total health expenditure was approximately $4.6 billion, of which an estimated $303 million was spent on pharmaceuticals. Pharmaceutical Þrms based in other countries have found both countries attractive as export markets and as bases for subsidiary manufacturing and R&D operations. In this overview of Australia and New Zealand’s pharmaceutical and biotechnology sectors, we examine the countries’ current regulatory structures, government strategies to promote R&D, and partnerships with overseas pharmaceutical companies, and we proÞle Australia’s top Þve pharmaceutical Þrms.

BUSINESS IMPLICATIONS •

Australia and New Zealand are relatively small pharmaceutical markets, but their demand for pharmaceuticals exceeds their manufacturing capacity, and the aging population provides markets for several major drug classes.

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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AUSTRALIAN AND NEW ZEALAND INDUSTRIES

The Free Trade Agreement recently negotiated between the Australian and U.S. governments opens up more opportunities for U.S. Þrms to list their products on the Australian Pharmaceutical BeneÞts Scheme, whereby the costs of these products to consumers will be subsidized by the Australian government. Many biopharmaceutical Þrms are emerging in Australia and New Zealand. However, these Þrms are relatively small and therefore license their technologies to larger Þrms at an early stage. Most do not intend to bring their own products to market; instead, they aim to partner with larger Þrms once their inventions have passed early-stage trials. The range and scope of existing business and R&D alliances reveal that Australian and New Zealand biotechnology Þrms are attractive investment and alliance partners. Several overseas Þrms have negotiated alliances and other investments with these Þrms. The Australian medical and pharmaceutical research base is strong and is a favorable target for alliances, collaborative research programs, and licensing. New Zealand’s research base is focused more on agricultural biotechnology, but the government’s biotechnology strategy has highlighted the potential for medical devices.

INTRODUCTION Australia and New Zealand are relatively small players in the global pharmaceutical market; together, they constituted approximately 1% of the global market in 2002. With a combined population of approximately 24 million people, these two countries are only slightly larger than the state of Texas and just under half the size of the United Kingdom. Nevertheless, both countries—Australia in particular—have high standards of living and operate government-funded monopsony systems that limit the cost of pharmaceuticals to the consumer. Many Þrms based in other countries have found Australia attractive both as an export market and as a base for subsidiary manufacturing and R&D operations. In 2000–2001, the Australian government’s health expenditure totaled approximately $20.2 billion, of which an estimated $2.3 billion was allocated to pharmaceuticals. (All dollar amounts are expressed in U.S. dollars unless otherwise stated. Where necessary, dollar amounts have been converted from Australian dollars or New Zealand dollars at the applicable exchange rate for the date. The May 2004 exchange rate for Australian dollars to U.S. dollars was approximately US$0.71; for New Zealand dollars to U.S. dollars, the exchange rate was approximately US$0.62.) In the same period, the New Zealand government’s health expenditure totaled approximately $4.6 billion, of which $303 million was spent on pharmaceuticals. Like other developed nations, Australia and New Zealand have aging populations and therefore have a high demand for pharmaceutical products that address diseases prevalent in these populations. The pharmaceutical markets in both countries rely heavily on imports because local manufacturing capacity is insufÞcient to meet demand. Australia is

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a major importer of pharmaceutical products and consumes approximately 1% of global pharmaceutical output. Australia’s pharmaceutical exports have grown as well, increasing from $73 million in 1987–1988 to $810 million in 1999–2000. The prime export markets for Australian pharmaceutical products are Asia and Europe. Australia’s pharmaceutical industry is more developed than New Zealand’s industry. It constitutes approximately 2% of the entire Australian manufacturing industry and has been growing signiÞcantly in both manufacturing capacity and exports. According to government data, the pharmaceutical industry generates more than $8.5 billion annually and employs approximately 30,000 people; the pharmaceutical manufacturing sector alone generates $4.3 billion per year and employs 16,000 people. In this article, we present an overview of the Australian and New Zealand pharmaceutical and emerging biotechnology sectors, examine the current regulatory environment and government strategies to promote investment in R&D, and proÞle Australia’s top Þve pharmaceutical Þrms. REGULATION, PRICING, AND REIMBURSEMENT Australia In Australia, both federal and state governments purchase pharmaceuticals. The federal government purchases drugs through the Pharmaceutical BeneÞts Scheme (PBS) administered by the federal Department of Health and Aging—consequently, the federal government determines national drug prices. More than 90% of prescription pharmaceutical products are sold through the PBS, which subsidizes a select list of drugs. State governments purchase large quantities of pharmaceuticals through their hospitals. Private hospitals also purchase drugs. The PBS has been in place for the past 57 years and is the mechanism by which the federal government subsidizes the consumer’s cost of buying drugs from pharmaceutical companies. Consumers for whom drugs have been prescribed by an authorized physician purchase them for a fraction of the original price. The PBS subsidizes approximately 80% of all available drugs, thereby bringing consumers’ costs for PBS drugs down to approximately $2.78–17.39 per prescription. Only low-income consumers who have been granted a government health care card pay the lowest amount. In 2002–2003, the PBS cost the Australian government $3.3 billion in prescriptions, a 7.2% increase over the previous year. The government subsidized 84.2% of the total cost of prescriptions; patient contributions accounted for the remaining 15.8%. The Therapeutic Goods Administration (TGA), a unit of the Department of Health and Aging, is responsible for the approval and regulation of all therapeutic goods in Australia. Under the Therapeutic Goods Act of 1989, the TGA ensures the quality, safety, and efÞcacy of drugs and medical devices. To be included in the PBS, products must Þrst be approved by the TGA for inclusion in the Australian Register of Therapeutic Goods. After TGA registration, drug manufacturers apply to the Pharmaceutical BeneÞts Advisory Committee (PBAC)

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TABLE 1. Australia’s Pharmaceutical Regulatory Agencies Agency Name

Function

Pharmaceutical Benefits Scheme (PBS) Therapeutic Goods Administration (TGA)

Pharmaceutical Benefits Advisory Committee (PBAC)

Pharmaceutical Benefits Pricing Authority (PBPA)

The PBS is not technically an agency; it is the prime mechanism by which the Australian government subsidizes a select list of price-controlled drugs. The TGA is responsible for approval of new drugs; it assesses and monitors quality, safety, and effectiveness. Products must be listed in the Australian Register of Therapeutic Goods prior to inclusion in the PBS. The PBAC meets four times per year and advises the federal government on which drugs should be subsidized. After a drug is registered by the TGA, the manufacturer can apply to the PBAC to have the drug listed on the PBS. The PBPA reviews the pricing of drugs currently listed in the PBS and recommends pricing for new drugs after the PBAC endorses the drugs for PBS inclusion. The PBPA negotiates the final price with the manufacturer.

to have their products listed on the PBS. The PBAC advises the Pharmaceutical BeneÞts Pricing Authority (PBPA), which in turn advises the minister for health, who gives Þnal approval. These advisory groups must consider the public need for the medication, its effectiveness, and its value for money compared with other drugs. Table 1 lists Australian pharmaceutical regulatory agencies and their functions. Multinational companies are the main suppliers of PBS-approved drugs (Table 2). In 2003, the three top categories of prescription drug spending were TABLE 2. Top Multinational Pharmaceutical Suppliers to the Australian Pharmaceutical Benefits Scheme, by Sales, 2002–2003 Company Pfizer GlaxoSmithKline Alphapharm Merck Sharp & Dohme AstraZeneca Bristol-Myers Squibb Pharmacia (now Pfizer) Aventis Eli Lilly ´ Sanofi-Synthelabo Wyeth Amrad Roche Novartis

Prescription Drug Sales ($MMa ) 390 310 304 293 261 214 212 163 145 132 93 91 85 80

a U.S. dollars, calculated at exchange rate of AUS$1.00 = US$0.71.

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serum-lipid-reducing agents ($550.2 million), drugs for acid-related disorders ($322.3 million), and agents that act on the renin–angiotensin system ($271.6 million). The top Þve drugs for PBS spending in 2003 were PÞzer’s atorvastatin (Lipitor, $238.9 million), Merck’s simvastatin (Denan, $220.7 million), AstraZeneca’s omeprazole (Losec, $127.3 million), GlaxoSmithKline’s salmeterol and ßuticasone (Seretide, $104.9 million), and Lilly’s olanzapine (Zyprexa, $100.9 million). Free Trade Agreement On May 18, 2004, the United States and Australia signed a landmark Free Trade Agreement (FTA). The goals of the agreement include the elimination of manufactured goods tariffs and the increase of trade between the two countries. In accord with the agreement, Australia will implement changes to the PBS to increase transparency and accountability and will institute an independent process for the review of pharmaceutical product listings. SpeciÞcally, Australia has promised to implement the following changes: • • • •

To ensure that applications from companies seeking to have products added to the PBS are considered by the PBAC within a speciÞed time frame. To publish the procedural rules and guiding principles that govern the PBAC’s consideration of those applications. To provide applicants with an opportunity to provide comments to the PBAC during the process. To provide applicants with a detailed explanation of the PBAC’s consideration of their applications.

These changes will establish a new process that will allow companies to appeal decisions against inclusion of their drugs on the PBS provided that the drugs have been “substantially made or transformed in the United States or Australia.” Among other advantages, the agreement provides U.S. companies that have applied for PBS listing with the following opportunities: • • • •

To consult relevant ofÞcials prior to submission of an application for listing, including consultation on the selection of a comparator pharmaceutical. To respond fully to reports on, or evaluations of, the applications that are prepared for the technical subcommittees of the PBAC. To obtain a hearing before the PBAC while it is considering reports or advice from the technical subcommittees regarding applications. To expeditiously collect sufÞcient information on the reasons for the PBAC’s determinations on applications for PBS listing in order to facilitate any appeals to the PBPA.

Australia is also trying to reduce the time required for implementation of the PBAC’s recommendations, introduce provisions for more frequent revision and dissemination of pharmaceutical beneÞts, and expedite procedures for applications that do not require an economic evaluation.

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The provisions of the FTA have generated considerable controversy. Some analysts claim that the FTA will increase the inßuence of U.S. drug companies over the PBS and, as a result, increase signiÞcantly the cost of prescription drugs. In a report on the FTA and the PBS drafted by leading Australian academics, researchers estimated that although the full effects of the FTA may not be felt for Þve years, the agreement could eventually cost the government as much as $1.5 billion. Other analysts have predicted that average drug prices could rise by as much as 30%. Skeptics are also concerned that the FTA could hinder the addition of generic drugs to the PBS. The Australian government, however, insists that the FTA will have no effect on drug prices in Australia. New Zealand The New Zealand Medicines and Medical Devices Safety Authority (Medsafe) is the principal pharmaceutical regulatory authority. Medsafe is a unit of the Ministry of Health and is responsible for the regulation and safety of therapeutic goods in New Zealand. The New Zealand Pharmaceutical Schedule (the “Schedule”) lists approximately 2600 prescription medicines and related health products subsidized by the New Zealand government. It is administered by the Ministry of Health and monitored by the Pharmaceutical Management Agency (PHARMAC), which was established in 1993 to improve the government’s management of pharmaceutical expenditures. PHARMAC determines which drugs will be included on the Schedule and uses a variety of mechanisms to control pricing. For example, New Zealand employs reference pricing to categorize products into therapeutic classes. The Schedule is similar to the Australian PBS in that patients must pay part of the cost of a prescription medication. The amount paid takes into account such factors as the patient’s age (pediatric patients under the age of six receive free medication) and income. In some cases, medications are only partially subsidized by the government, so patients may also have to pay a manufacturer’s surcharge (the difference between the government subsidy and the manufacturer’s price) plus a pharmacy markup. Lists of major suppliers of drugs to the New Zealand Medsafe Scheme are not available. Trans-Tasman Therapeutic Products Agency On December 10, 2003, Australia and New Zealand signed the Agreement Between the Government of Australia and the Government of New Zealand for the Establishment of a Joint Scheme for the Regulation of Therapeutic Products. This treaty will transform the two nations’ current pharmaceutical regulatory structures by establishing a new Trans-Tasman Therapeutic Products Agency. The joint scheme evolved from the 1997 Trans-Tasman Mutual Recognition Agreement (TTMRA), which integrated the New Zealand and Australian economies but excluded therapeutic products.

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GOVERNMENT-SPONSORED INVESTMENT IN R&D Australia For several years, the Australian government has been attempting to increase the amount of R&D and value-added manufacturing undertaken by the Australian pharmaceutical industry. The Factor (f) Scheme, introduced in 1987, compensated companies for low PBS drug prices provided that they carried out R&D or drug manufacturing in Australia. In 1999, the Factor (f) Scheme was replaced by the Pharmaceutical Industry Investment Program (PIIP). Under PIIP, nine companies received shares of $190 million over Þve years (from July 1999 through June 2004) in return for commitments of at least $1 million each in additional R&D and production value-added. The aim was to partially compensate pharmaceutical manufacturers for price and volume constraints imposed under the PBS. Of these nine companies, three are based in Australia: CSL, Mayne Pharma, and Amrad. The remainder are subsidiaries of foreign multinationals: BristolMyers Squibb, Eli Lilly, GlaxoSmithKline, Janssen-Cilag, PÞzer, and Pharmacia (now PÞzer). In 2003, the Productivity Commission evaluated PIIP and determined that although the program was effective in enhancing R&D and adding value to production, it was not productive enough to make a real impact on Australia’s economy. In addition, the major reason for its introduction—to help counter the effects of low PBS prices on pharmaceutical activity—was not strong enough to justify a tax-funded program. Based on these Þndings, the Productivity Commission recommended terminating PIIP at the end of June 2004 and launching a new pharmaceutical R&D program, the Pharmaceuticals Partnership Program (P3), on July 1, 2004. P3 aims speciÞcally to increase pharmaceutical R&D activity in Australia throughout the entire value chain, including biotechnology, originator, and generic drug companies. To achieve this goal, P3 gives participating companies, over a Þve-year period, 30 Australian cents for each additional Australian dollar they spend on eligible R&D in Australia, up to a cap of $10 million (U.S. $7.1 million). The Þrst P3 participants were announced on April 22, 2004. In 2001, the Australian government invested in the promotion of scientiÞc research through the “Backing Australia’s Ability” program. The government initially funded $2.13 billion for Þve years (through 2005–2006). In May 2004, the government announced its pledge of an additional $3.7 billion to R&D over a seven-year period beginning in 2004–2005. Recipients of the new funding include the National Health and Medical Research Council ($141.8 million), the National Stem-Cell Center ($41 million), and the National Biotechnology Strategy and Biotechnology Australia ($14 million). Australia’s state governments also support the pharmaceutical industry through their development of biotechnology strategies, which largely aim to support the growth of pharmaceuticals-related biotechnology Þrms. State initiatives include the New South Wales BioFirst Strategy; the Queensland Bioindustries Strategy; the South Australia Bio Innovation SA strategy; the Victoria Biotechnology

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Strategic Development Plan; and the Australian Capital Territory Bio-Business Strategy. New Zealand New Zealand has no equivalent to Australia’s P3 scheme. However, the government has been very active in developing strategies for its indigenous biotechnology sector and related biopharmaceutical manufacturers. In May 2003, the Biotechnology Taskforce—a group convened by the Ministry of Research, Science, and Technology—released a comprehensive report addressing key infrastructure changes needed to support biotechnology growth. The report outlined an ambitious ten-year plan for biotechnology expansion and offered 28 recommendations categorized by people, funding, institutions, infrastructure, and global participation. Target goals for the next ten years included the following: • • • •

Tripling the size of the New Zealand biotechnology sector from 350 organizations to more than 1000. Increasing total biotechnology employment from 3900 to 18,000. Nurturing a Þvefold increase in the number of core biotechnology companies from 40 to 200. Improving performance both in public research agencies and in private companies to increase export values to $585 million per year (current levels are approximately $146 million per year).

In response to the Biotechnology Taskforce’s report, the ministry announced in September 2003 the establishment of a $7 million trans-Tasman biotechnology fund to be managed by New Zealand Trade and Enterprise. The fund provides partnership funding to New Zealand and Australian companies that propose working together on biotechnology development, manufacturing, and marketing. The government also allocated $0.76 million of support over three years for the newly formed industry association, New Zealand Bio, which was created from the merger of the industry groups BIOTENZ and the New Zealand Biotechnology Association. The New Zealand government also helps provide funding through Investment New Zealand, which supports direct investment to New Zealand companies and to overseas businesses that seek to establish operations in New Zealand. In one example of this fund’s commitment to the biotechnology sector, the Aucklandbased biopharmaceutical company Protemix received a $1.2 million government underwrite from Investment New Zealand in September 2003. The company also received an $11.1 million infusion from a private New Zealand consortium headed by Birnie Capital Partners. The funds will be used to support Protemix’s research into diabetes treatments. THE EMERGING BIOTECHNOLOGY INDUSTRY The Australian biotechnology industry is similar to that of the United States and the United Kingdom in that most new Þrms have emerged from research institutions. The number of biotechnology Þrms has grown rapidly and now

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stands at just under 400. About one-third of these companies are developing biopharmaceuticals; another 25% are developing human diagnostics. Most of these small biopharmaceutical Þrms do not intend to bring their own products to late-stage clinical trials; they plan to outlicense their molecules after reaching Phase I or Phase II. Despite the large number of Þrms, fewer than 20 Phase II clinical trials are under way for Australia-developed biopharmaceutical products, and only three Australia-developed biopharmaceutical products are on the global market: Zanamivir (Relenza), an inßuenza treatment developed by Biota and marketed by GlaxoSmithKline; the SIR-spheres micropolymer bead cancer therapy, developed and marketed by Sirtex Medical; and Promensil, a natural plant-based treatment for menopause developed by Novogen. The proportion of new spin-offs that are developing biopharmaceuticals appears to be rising: in recent years, about two-thirds of new Þrms have been aiming at this market (Figure 1). This increase is due to new mechanisms of early-stage funding introduced by government, greater attention to commercialization from universities and medical research institutes, and greater interest from venture capital Þrms in biotechnology investments that may yield high returns. Several major biotechnology-oriented venture capital funds were established in Australia in 2003, including Burrill Australia, Scientiaus, and Intersuisse. More than 100 publicly listed Australian Þrms are included in the Australian Stock Exchange’s Healthcare and Bioscience Index. Approximately 50 are developing biopharmaceuticals, and many are active overseas even though their products are not yet on the market. Table 3 lists several major commercial deals entered into recently by Australian biopharmaceutical Þrms.

FIGURE 1. Australia: Target sectors for biotechnology firms formed in 2002–2003.

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TABLE 3. Select Deals Between Australian Biopharmaceutical Firms and International Companies, 2003–2004 Australian Biopharmaceutical Company

International Partner

Date

Value (US$MM)

pSivida

QinetiQ (UK)

June 2004

41

Australian Cancer Technology Benitec

Galencia Pharmaceuticals (US) Avocel (US)

May 2004

5

May 2004

5.5

AGT Biosciences

ChemGenex Therapeutics (US)

April 2004

10

Mayne Pharma

aaiPharma (US)

March 2004

100

Ozgene

National Institutes of Health (US)

March 2004

8.5

Biota Holdings

Thermo Electron (US)

March 2004

—

Cellestis

Statens Serum Institut (SSI) (Denmark)

March 2004

—

Mayne Pharma

Ivax (US)

February 2004

—

Pharmatel

InKine Pharmaceutical (US)

January 2004

—

CSL

Aventis Behring (US)

December 2003

925

Comments Acquisition of 100% ownership of pSivida’s U.K. subsidiary pSiMedica in an agreement with QinetiQ, pSivida’s joint venture partner in pSiMedica. Acquisition of Galencia Pharmaceuticals by Australian Cancer Technology’s U.S. subsidiary Adjuvantys. Acquisition of RNAi-based therapeutics company Avocel and establishment of the U.S. subsidiary Benitec. Cross-border merger of AGT and ChemGenex to establish ChemGenex Pharmaceuticals. The merged company will focus on developing products for cancer, diabetes, obesity, and central nervous system disorders. Acquisition of aaiPharma’s injectable multivitamin business (MVI and Aquasol products). Contract to supply genetically modified mice to researchers at the NIH’s National Institute of Allergy and Infectious Diseases. Profit-share agreement on worldwide sales of the influenza kit FLU OIA A/B. Distribution agreement for the Cellestis tuberculosis QuantiFERON TB-Gold in vitro diagnostic test in Poland, Hungary, and the Nordic and Baltic countries. Supply agreement for marketing and distribution in Europe of the Ivax injectable paclitaxel product, Paxene. Licensing for the rights to register, manufacture, and market InKine’s gastroenterology drug Visicol for use in Australia and New Zealand. Acquisition of Aventis’s plasma therapeutics business. The business will be combined with CSL’s ZLB Bioplasma business and will be renamed ZLB Behring.

TABLE 3. (continued) Australian Biopharmaceutical Company

International Partner

Date

Value (US$MM)

Gradipore CSL

Serologicals (US) Pfizer (US)

December 2003 December 2003

18 126

BresaGen

Nexgen Technologies (US) GeneLabs Diagnostics (Singapore)

November 2003

0.275

September 2003

—

Select Vaccines

BGen (BresaGen subsidiary)

AlgoRx Pharmaceuticals (US)

September 2003

—

Mayne Pharma

NaPro BioTherapeutics (US) DaeWoong Chemical Company (Korea) Texas Christian University of Fort Worth (US)

August 2003

72

June 2003

3

June 2003

—

Genetic Technologies

Pyrosequencing AB (Sweden)

March 2003

0.5

Progen Industries

Sequella (US)

January 2003

—

Bio Pharma pSivida

Comments Acquisition of Serologicals’ therapeutic plasma business. Acquisition of CSL’s Animal Health business by Pfizer Animal Health. 4% stake in Nexgen and assignment of BresaGen’s cell delivery catheter licenses to Nexgen. Exclusive, worldwide, twelve-year license to GeneLabs for the manufacture and distribution of Select Vaccines’ hepatitis E rapid diagnostic kit. Acquisition of AlgoRx’s XeriJect drug delivery platform; BGen will use the technology to develop generic biopharmaceuticals. Acquisition of NaPro’s generic injectable paclitaxel business. Supply agreement with DaeWoong to provide chenodeoxycholic acid (CDCA). Revenue sharing relating to the intellectual property of self-assembling BioSilicon and polymer composite scaffolds for tissue engineering when commercialization through licensing activities begins. License to Genetic Technologies’ patents that cover noncoding DNA analysis and mapping for use with Pyrosequencing’s DNA-sequencing technology. Contract for Progen to manufacture the MPT64 protein for Sequella’s skin patch tuberculosis diagnostic test.

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AUSTRALIAN AND NEW ZEALAND INDUSTRIES

FIGURE 2. New Zealand: Target sectors for the biotechnology industry as of 2003.

The New Zealand biotechnology industry differs greatly from the Australian industry. Although New Zealand has a signiÞcant group of biopharmaceutical Þrms, agbiotech plays a relatively stronger role (Figure 2). Few biopharmaceutical Þrms are currently listed on the New Zealand Stock Exchange. THE RESEARCH INFRASTRUCTURE Australia Most of Australia’s basic research is carried out in public universities or funded through government organizations such as the National Health and Medical Research Council (NHMRC) and the Australian Research Council. Australia has an excellent public and university medical research infrastructure, a highquality clinical research capability, good access to the Asia-PaciÞc region, and low sovereign risk. By the standards of the Organization for Economic Cooperation and Development (OECD) nations, Australian support of health and medical research is low. In 1995, Australia spent only 0.115% of gross domestic proÞt (GDP) on health and medical R&D, 50% less than the GDP-weighted OECD average for developed nations. Following a major review of health and medical research, the government committed in 2000 to doubling its funding of health and medical research and named this Þeld as one of four national research priorities. In 2003, the Australian government allocated approximately $193.8 million to the NHMRC for health and medical research, a vast increase compared with the approximately $102.4 million invested in 1999. The review also led to the recognition of the need for a more coordinated approach to identifying national priority areas in health and medical research so

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163

that increased funding could be directed at particularly urgent research areas. The Strategic Research Development Committee (SRDC), responsible for directing NHMRC funds to urgent research areas, has identiÞed Þve areas of research priority: aging, chronic diseases, emerging/reemerging diseases, indigenous health, and socioeconomic determinants of health. The increase in funding and the enhanced focus on health research in these priority areas indicate that Australia has apparent strengths in several therapeutic areas, including cancer research, infection and immunity, and mental health and neuroscience. SigniÞcant research infrastructures for biotechnology and drug development R&D include the following: •

• •

•

•

Australian synchrotron, a particle accelerator being built at Monash University in Melbourne with funding from the Victoria government and other governments for commissioning in 2008. Australian Centre for Plant Functional Genomics, funded by the federal government through a national competition and headquartered in Adelaide. Australian Proteome Analysis Facility, based in Sydney at Macquarie University, one of several national facilities funded by the National Health and Medical Research Council. Australian National Stem-Cell Centre, located at Monash University in Melbourne and funded by the federal government in response to a national competition. In 2003, the center entered into an alliance with the U.S. biotechnology Þrm LifeCell. Institute of Molecular Biosciences, funded by the Queensland government under its state biotechnology strategy.

These research institutions and the research being undertaken at universities, the Commonwealth ScientiÞc and Industrial Research Organization (CSIRO), and medical research institutes are generating a growing number of biotechnology patents. The number of U.S. biotechnology patents that originate with Australian inventors has been increasing steadily, to approximately 80 per year. Most of these patents are being granted to three institutions: the University of Melbourne, the University of Sydney, and the Garvan Institute of Medical Research. Several international companies—including AstraZeneca, Aventis, BristolMyers Squibb, Johnson & Johnson, and PÞzer—collaborate with public sector research institutions or universities in discovery programs (Table 4). AstraZeneca and Johnson & Johnson maintain their own discovery laboratories in Queensland and New South Wales, respectively. These initiatives have been encouraged by government programs aimed at expanding R&D and manufacturing activity in Australia. However, international companies generally undertake the preclinical development of any lead compounds identiÞed in Australia in their established R&D facilities outside Australia.

164

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TABLE 4. Select Australia-Based R&D Activities for Multinational Pharmaceutical Companies Multinational Pharmaceutical Company

Australian Company Name/Location

AstraZeneca

AstraZeneca R&D Griffith University (Brisbane, Queensland)

Aventis

Aventis Pharma (Sydney, New South Wales)

Bristol-Myers Squibb

Bristol-Myers Squibb Australia (Noble Park, Victoria)

Johnson & Johnson

Johnson & Johnson Research (Eveleigh, New South Wales)

Pfizer

Pfizer Biometrics Centre (West Ryde, New South Wales)

Comments Natural product discovery alliance established in June 1993 focuses on lead discovery and screening of extracts from plants and marine organisms collected from Queensland’s rain forests and the Great Barrier Reef. The unit has found approximately 700 bioactive compounds from more than 35,000 specimens; about 40% of these compounds are new to science. Total investment from AZ to date represents $100 million. Investment of more than $12 million every year. More than 40 R&D studies are being conducted in Australia and New Zealand, including clinical trials for rheumatoid arthritis, breast cancer, and infectious diseases and for once-daily insulin (Lantus) to treat diabetes. International R&D hub. Since 1990, the Australian research program has included more than 95 different institutions and more than 200 investigators focusing on cardiovascular diseases, cancer, immune disorders, central nervous system disorders, and infectious diseases. Established in 1987, the company’s focus is to identify new product opportunities within Australia in the area of human health for early development of pharmaceuticals, biopharmaceuticals, gene therapy, and clinical diagnostics. In October 2003, Pfizer opened its $10.5 million Australian Biometrics Centre to design and manage clinical trials worldwide. Currently, the center is managing 20 research studies; the number will grow to 50 over the next two years.

R&D ALLIANCES WITH OVERSEAS PARTNERS

165

New Zealand The strengths of the New Zealand research sector are in agriculture rather than therapeutics. The 2003 report of the Biotechnology Taskforce listed the following key strengths: • • • • •

Large-animal-based biotechnologies (animal breeding, trait selection, animal physiology, high animal health status). Plant-based biotechnologies (unique germplasm libraries and expressedsequence tag libraries, plant physiology). Biomedical science and drug discovery (cardiovascular, neuroscience, asthma, diabetes, cancer, osteoporosis, clinical trials). Bioprocessing technologies and biomanufacturing (established infrastructure, bioengineering skills, software). Innovative foods and health (understanding of proteins, carbohydrates, and fats, especially in dairy, and marketing expertise).

These strengths derive from the New Zealand public sector R&D system’s concentration on agriculture through its Crown Research Institutes (CRIs), established in 1992. The nine CRIs cover agriculture, horticulture, industry, environment, geology, water/atmosphere, land care, forestry, and crops/foods. However, New Zealand has expressly identiÞed medical technologies and devices as a potential area of emerging strength, based on the work of several university and CRI bioengineering research groups and on existing examples of industry licensing and codevelopment of medical devices, including licensing to U.S. Þrms. Also, Australia and New Zealand have forged signiÞcant links—notably, an alliance between the New Zealand and Queensland governments and an alliance between the New Zealand and Victoria governments to strengthen scientiÞc relations and to cooperate in promoting biotechnology developments in medicine, agriculture, and biopharmaceuticals. New Zealand is also considering contributing to the cost of the Australian Synchrotron being built in Victoria. R&D ALLIANCES WITH OVERSEAS PARTNERS International and intercompany alliances, especially with major corporations, are a key success factor for Australian biopharmaceutical Þrms. Intercompany alliances made up approximately 3% of the international deals reported in 2002–2003. Table 5 lists select alliances announced in 2003–2004. New Zealand’s biotechnology and pharmaceutical Þrms, although smaller in number than Australia’s, are also forming international alliances for both commercial and R&D purposes (Table 6). Also, the New Zealand government recently signed an alliance with the state of Iowa to support cooperation in biotechnology research by facilitating contacts and reinforcing links between New Zealand and Iowa researchers and life sciences companies.

166 TABLE 5. Select Australia-International R&D Alliances, 2003–2004 Australian Company

Partner(s)

Date

Value ($US MM)

Bionomics

Perkin-Elmer (US)

May 2004

CancerVac

Biomira (Canada)

March 2004

15.4

QrxPharma

Shire Laboratories (US)

March 2004

—

Acrux

Vivus (US)

February 2004

13.3

CSL

Chiron (US)

January 2004

—

Cortical

Genzyme (US)

December 2003

0.75

Norwood Abbey

TAP Pharmaceuticals (US) Institutes for Pharmaceutical Discovery (US)

November 2003

—

November 2003

—

AGT Biosciences

—

Comments Collaboration to evaluate and discover ion channel compounds to treat epilepsy and anxiety through Perkin-Elmer and Bionomics’ high-throughput screening platforms. Licensing and development of CancerVac’s Mannan-MUC1 fusion protein cancer vaccine and 10% equity stake in CancerVac. Agreement for Shire to develop extended-release formulations of QrxPharma’s novel opioid product, Q8003. Licensing agreement for Vivus to develop and commercialize in the United States Testosterone MDTS and Estradiol MDTS, delivered using Acrux’s metered-dose transdermal system. Joint agreement to develop a therapeutic hepatitis C vaccine combining Chiron’s HCV antigens and CSL’s Iscomatrix adjuvant technology. Licensing and investment in Cortical to develop oral drugs for inflammatory diseases. Licensing of immunology intellectual property to TAP for use in Lupron Depot drug enhancement and $2 million investment in Norwood Abbey. Partnership to develop over-the-counter products to treat obesity.

TABLE 5. (continued) Australian Company

Partner(s)

Date

Value ($US MM)

Biota

Sankyo (Japan)

October 2003

—

Elan Biodynamics

Meditech Pharmaceuticals (US)

September 2003

—

Biota

USAMRIIDa (US) and NIAIDb (US) Domantis (UK)

September 2003

—

August 2003

—

pSivida

McComb Foundation (UK)

August 2003

—

Eiffel Technologies

Meridica (UK)

August 2003

—

Eiffel Technologies

Oriel Therapeutics (US)

July 2003

—

Amrad

Merck Sharp & Dohme Australia (Australia)

June 2003

112

EvoGenix

Comments Cross-licensing agreement whereby the companies will combine their relevant long-acting influenza drugs into a single pipeline of products, which will be offered to licensing partners for further development and marketing. Joint venture for the creation of Bio-D-Tech, specializing in the pursuit of bioterrorism solutions. The company will commercialize a biohazard detection system for toxic agents, including aflatoxin and anthrax. Three-year agreement for testing Biota’s antiviral compounds against the SARS virus. EvoGenix will apply its protein evolution and optimization technology to one of Domantis’s domain human antibody therapeutic leads. McComb will evaluate the use of pSivida’s BioSilicon as a tissue-engineering substrate for the treatment of wounds and burns. Collaboration whereby Eiffel will supply a reengineered asthma drug to Meridica for use in a new drug-delivery inhalation device. Eiffel will supply a reengineered and modified asthma drug to Oriel for testing with its inhalation device platform. Licensing and investment in Amrad for drug development in asthma, respiratory diseases, and oncology.

167

168

TABLE 5. (continued) Australian Company

Partner(s)

Date

Value ($US MM)

National Stem-Cell Centre

LifeCell (US)

June 2003

—

Amrad

Medarex (US)

May 2003

—

Virax

MedImmune (US)

April 2003

0.24

Prana Biotechnology

Shering AG (Germany) and Neuroscience Victoria (Australia) Medarex (US)

March 2003

4.5

March 2003

—

diaDexus (US)

February 2003

—

Apath (US)

January 2003

—

Oncomab (PrimaBioMed subsidiary) Queensland Institute of Medical Research BioProspect

a USAMRIID, U.S. Army Medical Research Institute of Infectious Diseases. b NIAID, National Institute of Allergy and Infectious Diseases. c IL, Interleukin.

Comments Licensing agreement and use of LifeCell’s tissue matrix technology with stem cells for tissue regeneration. LifeCell will have rights to commercialize resulting products. Licensing agreement whereby Amrad will use the Medarex UltiMAb human antibody development system to generate antibodies against Amrad’s IL-13c receptor alpha target for the treatment of asthma. Supply agreement whereby Virax will provide technology to MedImmune to develop future products. Licensing and funding agreement to discover new targets and diagnostics for neurological diseases. Codevelopment of human monoclonal antibodies for treating cancer and joint commercialization of any resulting antibody products. License agreement giving diaDexus exclusive development and commercialization rights to Testisin, a cell-surface serine protease cancer target. Collaboration for the screening of BioProspect’s natural plant extract library for potential antiviral therapeutics for hepatitis C virus and other pathogenic viruses.

TABLE 6. Select New Zealand-International Commercial and R&D Alliances New Zealand Company

Partner

Date

A2

GeneSeek (US)

January 2004

Virionyx

HauptmanWoodward Medical Clinic (US) and ZeptoMetrix (US) Danisco (Denmark)

June 2003

BLIS Technologies BLIS Technologies Douglas Pharmaceuticals

Astra Grace (Australia) Senetek (US)

November 2002 November 2002 November 2002

Endocrinz

Pharmacia (Pfizer) (US)

April 2002

Proacta Therapeutics

Onyx Pharmaceuticals (US)

March 2002

Genesis Research and Development Genesis Research and Development

Immunex (US)

February 2002 November 2001

SR Pharma (UK)

Comments Contract agreement whereby GeneSeek will provide high-throughput DNA analysis to genotype dairy cattle for production of A2 milk and other A2 dairy products. Codevelopment and collaboration on a vaccine to prevent SARS.

169

Marketing and licensing agreement for the BLIS K12 Throat Guard in Germany. Marketing and licensing agreement for the BLIS K12 Throat Guard in Korea. Marketing and distribution agreement for Invicorp, Senetek’s erectile dysfunction product, for New Zealand. Investment of $2 million in Endocrinz to support research into metabolic disorders, including diabetes and giantism. Codevelopment and collaboration to evaluate a novel cancer treatment to supplement Onyx’s anticancer viruses with Proacta’s prodrug technology. Proacta will conduct all research activities. Three-year joint collaboration to study immune modulator compounds. Joint drug development program for SRP299 and AVAC for the treatment of eczema (atopic dermatitis).

170 TABLE 6. (continued) New Zealand Company

Partner

Date

EPTTCOa

Onyx Pharmaceuticals (US)

August 2000

EPTTCO

Vion Pharmaceuticals (US)

April 1999

Bioengineering Institute, Auckland University

Johns Hopkins University (US), MIT (US), Oxford University (UK), Physiome Sciences (US), other research groups

Formally launched in 1997

Comments Multiyear licensing agreement to supplement Onyx’s anticancer viruses with EPTTCO’s prodrug-converting enzymes, designed to fight cancer without harming normal cells. Codevelopment of Vion’s TAPETb bacterial vector platform (genetically modified Salmonella bacteria), incorporating EPTTCO’s prodrug-converting enzymes to fight various cancers. The Physiome Project is a worldwide public domain effort to define and develop complex computer models of human cells, tissues, and organs and to simulate the behavior of diseases such as diabetes, hypertension, and asthma.

a EPTTCO (Enzyme Pro-drug Therapy Company) is a joint venture of Auckland Uniservices (commercial arm of the University of Auckland), the Institute of Cancer Research in

London, and UK Cancer Research Campaign Technology. The company has headquarters in the United Kingdom. b TAPET, tumor amplified protein expression therapy.

AUSTRALIA’S TOP FIVE PHARMACEUTICAL FIRMS

171

Many overseas pharmaceutical Þrms also conduct R&D activities in New Zealand. Some companies, however, have scaled back or restructured their operations, citing constraints imposed by PHARMAC and limited funding for drug development. For example, in late 2002, GlaxoSmithKline announced that it was restructuring its New Zealand operations to focus on a limited number of key products, and AstraZeneca moved its clinical trials to Australia because of the lack of support from the New Zealand government. We estimate that overseas pharmaceutical companies invest approximately $14.5 million in New Zealandbased pharmaceutical R&D. AUSTRALIA’S TOP FIVE PHARMACEUTICAL FIRMS The Australian pharmaceutical industry comprises approximately 120 locally owned or internationally owned companies. Most large, multinational, researchbased pharmaceutical companies maintain subsidiaries in Australia that may of total pharmaceutical sales. The exceptions are GlaxoSmithKline’s production of alkaloids in Tasmania (to the south), Janssen-Cilag’s manufacture of alkaloids, and niche manufacturing activities undertaken by IDT, an Australian Þrm that originated at the Australian College of Pharmacy. Australia also has a small number of locally owned pharmaceutical Þrms. As of June 8, 2004, the top Þve Australian Þrms—CSL, Mayne Pharma, Novogen, Peptech, and Sigma Pharmaceuticals—had a market cap totaling $8.2 billion. We brießy proÞle these Þve companies. CSL CSL was founded in 1916 as the government-owned Commonwealth Serum Laboratories. Its main role in its early history was the production of inßuenza vaccine. CSL was privatized and listed on the Australian Stock Exchange in 1994. Since then, it has grown and diversiÞed considerably and now manufactures and sells human pharmaceutical and diagnostic products and vaccines, products derived from human plasma, and cell culture and blood grouping reagents. The Þrm also acts as agent for a large range of other products. CSL is the largest Australia-owned pharmaceutical company and employs approximately 1300 people. Its head ofÞce and major production facility is in Melbourne, but it also has subsidiaries in the United States (JRH Biosciences and ZLBUSA), Switzerland (ZLB), New Zealand (CSL), the United Kingdom (ZLB), Germany (ZLB), Belgium (ZLB), Italy (ZLB), and Hong Kong (CSL). CSL’s pharmaceutical division develops, manufactures, and markets vaccines for human use, including vaccines for children, travel, respiratory illnesses, and antivenoms. Pharmaceutical sales earned revenues of $143.5 million in Þscal year 2002–2003. The Þrm recently introduced meningococcal C vaccine to the Australian market, on license from Chiron in the United States. CSL sold its Animal Health business to PÞzer for $131 million in March 2004. The aim of the sale was to provide funding for the acquisition of Aventis’s blood fractionation business (Aventis Behring) for $925 million. The acquisition was announced in December 2003 and the deal was completed on March 31, 2004.

172

AUSTRALIAN AND NEW ZEALAND INDUSTRIES

Mayne Pharma Mayne Group operates a generic pharmaceuticals manufacturing business and wholesale distribution business staffed by more than 1300 employees in 20 countries. In March 2004, Mayne Pharma acquired the MVI multivitamin business of United States-based aaiPharma for $100 million. This acquisition attracted some criticism in Australia owing to U.S. regulators’ investigation of aaiPharma for sales abnormalities. Mayne has defended the strength of its due diligence during the acquisition and has included several protections in the asset purchase agreement. The Þrm has manufacturing facilities in Australia, Germany, and Puerto Rico. Its product portfolio includes injectable therapies for the treatment of cancer, infectious diseases, cardiovascular disease, and neurological disorders; it also includes anesthesia, oral antibiotics, and pain control products. The Þrm also inlicenses improved chemical entities and generics. However, Australia’s low uptake of generic pharmaceuticals (compared with uptake in the United States) has been hampering Mayne Pharma’s generics business. On the other hand, Mayne also works with owners of chemical entities to develop new modes of drug delivery and thereby extend the patent life of products already on the market. Novogen Novogen was founded in 1992 to commercialize the work of its founder, Dr. Graham Kelly, a cancer researcher at an Australian medical school. The Þrm listed on the Australian Stock Exchange in 1994 and on NASDAQ in 1999. It has three divisions: pharmaceuticals (prescription pharmaceuticals), consumer health (overthe-counter (OTC) products), and Marshall Edwards, a wholly owned subsidiary. Marshall Edwards focuses on the development of cancer drugs and is listed on the United Kingdom’s Alternative Investment Market. Overall, Novogen’s broad areas of focus are anti-inßammatories, cancer, heart disease, rheumatoid arthritis, neurology, and autoimmune disease. Novogen’s main strength stems from intellectual property based on isoßavones, compounds found naturally in red clover. Novogen has developed synthetic pathways for its isoßavones and no longer extracts them from plants. Isoßavones can help regulate estrogen in women and may prevent osteoporosis and heart disease and help maintain normal prostate function in men. The company’s three major OTC products are Promensil, Rimostil, and Trinovin, all of them sold in Australia, New Zealand, North America, and The Netherlands. As noted earlier, Novogen recently won a place on the Australian government’s P3 program. Novogen stands to receive the maximum allowable grant of $7.3 million over Þve years, dependent on agreed R&D spending targets. Novogen’s funding will be used to accelerate the development of phenoxodiol, its lead anticancer compound, through Phase II clinical trials. Peptech Peptech originated as a spin-off from research at the Commonwealth ScientiÞc and Industrial Research Organisation in the mid-1980s and listed on what was

OUTLOOK

173

then the Second Board of the Australian Stock Exchange soon after. Peptech has several peptide-based patents targeted toward cancer, infectious diseases, and inßammation in humans and toward the regulation of fertility in animals. Peptech’s human health division aims to use antibodies and domain antibodies to address important clinical needs in the Þelds of immunology and cancer, targeting tumor necrosis factor (TNF) and a tumor-speciÞc protein. The company has studied the relationships between TNF and antibodies and has several patents in Australia, Þve European countries, Canada, and the United States. The Peptech patents highlight the parts of the TNF molecule that are critical to the design of a drug that speciÞcally blocks a given TNF function. The company’s TNF patents have been licensed to two global pharmaceutical companies that market antiTNF therapeutic antibodies. The patents also serve as a foundation for Peptech’s anti-TNF therapeutic domain antibody program with Domantis, a U.K. Þrm. In April 2004, Peptech announced a takeover bid for another listed Þrm, Agenix, to create a $306 million diagnostics, animal health, and pharmaceutical company; it will be Australia’s second-largest listed biotechnology company. The merged entity will retain the name Peptech, and the total combined revenue of the merged group is expected to reach $75 million within two years. Sigma Pharmaceuticals Founded in 1912 by two Melbourne pharmacists, Sigma Pharmaceuticals employs more than 1500 people in Australia. It was listed on the Australian Stock Exchange in 1999 and now has annual sales of $1.3 billion, boosted by the acquisition of two retail chemists’ product lines (Guardian and Amcal) in 1997 and 1998, respectively, and of Herron Pharmaceuticals in 2003. In 1999, Sigma also acquired the Australian pharmaceutical manufacturing business of SmithKline Beecham, based in the United Kingdom. The company also attempted a merger with Australian Pharmaceutical Industries in 2002, but the merger was rejected by the Australian regulators. Sigma is a major manufacturer and the largest pharmaceuticals contract manufacturer in Australia. Its contract manufacturing capacity includes liquid products, creams and ointments, and sterile and solid dose production. Sigma targets OTC remedies for pain and fever, bowel health, coughs and colds, hay fever, and cold sores. The company aims to expand this side of the business aggressively. Although pharmaceutical sales account for only 10% of revenue, they contribute more than 55% toward earnings before interest and tax. OUTLOOK Australia provides an attractive market for pharmaceutical Þrms for several reasons: • • •

High demand for imported pharmaceutical products. Transparent regulatory environment. New opportunities emerging through the Free Trade Agreement.

174 • •

AUSTRALIAN AND NEW ZEALAND INDUSTRIES

Rapidly growing number of biopharmaceutical Þrms that are seeking international partners. Strong research base and government funding priorities that target health and medical research.

In New Zealand, both the pharmaceutical market and the number of biopharmaceutical Þrms are smaller, but the government recognizes biotechnology’s potential and is making a strong push to enhance relationships with Australia, further harmonize the regulatory system, and help New Zealand companies Þnd overseas partnerships. Thus, the outlook for the biotechnology sectors in Australia and New Zealand is strong. There are opportunities for overseas companies to sell products in both markets as well as to form partnerships with Þrms in both countries. A growing number of overseas companies, particularly U.S. companies, have beneÞted from research and product-based alliances with Þrms in Australia and New Zealand. The creation of the Trans-Tasman Therapeutic Products Agency in July 2005 should help create a uniÞed market and enable companies to penetrate this region. The number of Australia- and New Zealand-based biotechnology Þrms is likely to continue to grow because government policy in both countries supports research commercialization. In particular, Australia’s strong research base will continue to yield new inventions that can be exploited commercially in global markets.

Overview of the World’s Major OTC Markets SUMMARY Although overshadowed by the prescription drug market, the over-the-counter (OTC) drug market should not be ignored. In 2003, sales in the United States, the Þve major European markets (France, Germany, Italy, Spain, and the United Kingdom), and Japan exceeded $40 billion (at retail prices), and the market has strong prospects for the future. Governments and other payers are increasingly looking to OTC drugs as a means of reducing their burgeoning expenditures on prescription medicines. The potential transfer of many drugs from prescription status to nonprescription status—a process known as Rx-to-OTC switching—could invigorate the self-medication market in the coming decade. In this article, we examine the current OTC environment in seven of the world’s major pharmaceutical markets (United States, France, Germany, Italy, Spain, United Kingdom, and Japan), and we discuss the trends that will likely fuel its growth. BUSINESS IMPLICATIONS •

With 2003 sales in the United States, the Þve major European markets (France, Germany, Italy, Spain, and the United Kingdom), and Japan in excess of $40 billion (at retail prices), the over-the-counter (OTC) drug market is by no means inconsequential. Furthermore, governments and other payers are increasingly looking at self-medication as a means of reducing their burgeoning expenditures on prescription drugs. The potential transfer of many drugs that were previously regarded as suitable only for prescription use to OTC status—a process known as Rx-to-OTC switching—could invigorate the self-medication market in the coming decade.

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

175

176 •

•

•

OVERVIEW OF THE WORLD’S MAJOR OTC MARKETS

In recent years, some U.S. health plans have added certain OTCs to their formularies as a way of containing their pharmaceutical costs. For example, following the switch of omeprazole (AstraZeneca’s Prilosec) to OTC status, some health plans began reimbursing the new OTC product and either altered the formulary status of prescription proton pump inhibitors or even excluded these drugs from coverage. Similarly, the switch of loratadine (ScheringPlough’s Claritin) to OTC status prompted some health plans to reassess the position of prescription antihistamines in their formularies. In 2002, the U.K. government published a radical list of more than 50 prescription-only medicines that could be switched to OTC status over the next Þve years. Most of these proposed switches are for indications previously considered unsuitable for self-medication—for example, stable angina, hypertension, prevention of stroke/myocardial infarction, chronic obstructive pulmonary disease, and erectile dysfunction. In July 2004, the United Kingdom became the Þrst country in the world to license an OTC statin: Zocor Heart-Pro, marketed by Johnson & Johnson MSD, is a 10-mg dosage of simvastatin. Other countries will observe the U.K. experience with OTC statins and will surely follow suit if they judge this experiment to be a success. Manufacturers may be able to fend off generics competition by switching a branded prescription drug to OTC status before its patent expires. By making a drug more accessible, OTC status may also boost sales by attracting new users.

INTRODUCTION The over-the-counter (OTC) drug market is often overshadowed by the prescription drug market. Sales of prescription medicines are far greater than those of OTC medicines and have grown much faster in recent years. The prescription drug sector frequently drives clinical innovation, whereas OTC drugs are sometimes regarded as dated drugs. Prescription medicines generally treat serious, even life-threatening, disorders, while self-medication is typically used for relatively minor ailments. Because the returns on prescription medicines tend to be much greater than those from self-medication products, R&D investment in new prescription drugs eclipses spending on OTC drugs. Nevertheless, the OTC market should not be ignored. With 2003 sales in the United States, the Þve major European markets (France, Germany, Italy, Spain, and the United Kingdom), and Japan in excess of $40 billion (at retail prices), this market is by no means inconsequential. Furthermore, governments and other payers are increasingly looking at the self-medication sector as a way to reduce their burgeoning expenditures on prescription drugs. The potential transfer of many drugs that were previously regarded as suitable only for prescription use to OTC status—a process known as Rx-to-OTC switching—could invigorate the self-medication market in the coming decade. In this article, we examine the current OTC environment in seven of the world’s major pharmaceutical markets (United States, France, Germany, Italy,

UNITED STATES

177

Spain, United Kingdom, and Japan). In each case, we consider drug registration and classiÞcation; consumer advertising; distribution, pricing, and reimbursement; Rx-to-OTC switching; and market trends. We conclude with a brief assessment of the outlook for the global OTC market. UNITED STATES The United States has the world’s largest OTC market by far: in 2003, sales totaled approximately $18 billion at retail prices. In terms of product classiÞcation, consumer advertising, distribution, and pricing, the U.S. market is one of the most liberal in the world, but the range of compounds available over the counter is much more limited than in many other countries. Drug Registration and Classification The United States assigns all medicines either prescription or nonprescription status. The default status is nonprescription. The Food, Drug, and Cosmetic Act limits drugs to prescription status only if they are not safe to use “except under the supervision of a practitioner licensed by law to administer such drug[s].” Dietary supplements are classiÞed as a subset of food. Manufacturers of dietary supplements may make one of three types of claims for their products: qualiÞed health claims, nutrient content claims, or structure/function claims. Health claims describe an association between a food, food component, or dietary supplement ingredient and a reduction in the risk of developing a given disease or health-related condition. Nutrient content claims deÞne the amount of a nutrient or dietary substance contained in a product. Structure/function claims explain how a product may affect the body’s organs or systems but may not mention any speciÞc disease. The manufacturer must provide the FDA with the text of its structure/function claims within 30 days of launching a new dietary supplement. Manufacturers of nonprescription drugs may apply for a marketing authorization from the FDA. Alternatively, some products may be “generally recognized as safe and effective” (GRAS/GRAE) by experts. Products that have GRAS/GRAE status do not require individual product licenses or authorizations but are subject to the general laws that govern all medicines sold in the United States (e.g., good manufacturing practice requirements, plant inspections, packaging requirements). Dietary supplements are not issued with individual licenses or marketing authorizations. Consumer Advertising The United States has arguably the most liberal environment for pharmaceutical advertising of any major pharmaceutical market in the world. Unlike most other countries, the United States permits direct-to-consumer (DTC) advertising of prescription drugs, subject to strict controls. Advertising of OTC drugs is permitted in all media. U.S. law prescribes no mandatory text for OTC advertisements. The

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OVERVIEW OF THE WORLD’S MAJOR OTC MARKETS

United States is also unusual in allowing comparative advertising of OTC drugs, whereby manufacturers assert the superiority of their products over rival brands. The U.S. government believes this practice stimulates competition. The Federal Trade Commission (FTC) is responsible for postpublication monitoring of advertisements for nonprescription drugs and dietary supplements. OTC advertising must comply with the same basic standards as advertisements for other consumer products: claims must be substantiated and advertisements must be neither deceptive nor unfair. Many states also have their own versions of the FTC Act and empower consumer protection authorities to take action against deceptive or unfair advertising. In addition, several self-regulatory systems for monitoring OTC advertising operate in the United States. Distribution Most countries restrict at least some nonprescription medicines to pharmacy distribution, and the purchase of certain products may require a pharmacist’s approval. In the United States, however, federal law permits the sale of OTC drugs in any retail outlet (e.g., pharmacies, mass merchandisers, food stores). Some states restrict the sale of a limited number of OTC controlled substances (e.g., products that contain codeine) to pharmacies. Mail-order and Internet pharmacies are permitted in the United States. Pricing and Reimbursement The United States does not subject OTC drugs to price controls and does not practice resale price maintenance (i.e., requiring all retailers to sell goods at a speciÞed price). Approximately one-third of states exempt medicines, including OTC drugs, from state sales tax. In recent years, some health plans have added certain OTC drugs to their formularies as a way of containing their pharmaceutical costs. For example, following the switch of omeprazole (AstraZeneca’s Prilosec) to OTC status, some health plans began reimbursing the new OTC product and either altered the formulary status of prescription proton pump inhibitors or even excluded these drugs from coverage. Similarly, the switch of loratadine (Schering-Plough’s Claritin) to OTC status prompted some health plans to reassess the position of prescription antihistamines in their formularies. Rx-to-OTC Switching The United States has no formal criteria for Rx-to-OTC switching; each case is judged on its own merits. In 1972, the FDA established a process known as the OTC review to evaluate the safety and efÞcacy of OTC ingredients, doses, formulations, and labeling used in nonprescription medicines. The OTC review sanctioned ingredient-related Rx-to-OTC switching, but no drugs have been switched under this system since 1992. More recent switches have been made on the basis of the new drug application (NDA) process and are generally product-speciÞc. However, if the drug in question has lost its patent or data

FRANCE

179

protection and faces competition from generic products registered by means of an abbreviated new drug application (ANDA), both the originator drug and the generics would be switched. The OTC review permitted manufacturers, the government, industry organizations, and other parties (by citizen petition) to initiate switch applications. For products approved by an NDA, it would be difÞcult for anyone other than the manufacturer to initiate a switch application. The United States has been more conservative than many European countries about Rx-to-OTC switching. Table 1 shows the number of compounds in various drug classes that are available over the counter in each of the major self-medication markets. Table 2 shows the legal status of select molecules in these markets. The range of compounds available without a prescription is much more limited in the United States than in the major European markets or Japan. However, the United States is unusual in permitting the OTC sale of insulin products and epinephrine-based asthma inhalers. Market The U.S. OTC market has stagnated in recent years. According to data from A.C. Nielsen, in 1999, sales totaled $18.9 billion, but they declined sharply in 2000 and have been slow to recover. In 2002, the U.S. OTC market was worth $17.2 billion at retail prices, a negligible increase over the preceding year. Among the best-selling OTC categories from 2001 to 2003 cough and cold remedies and related products were by far the largest category and experienced relatively strong growth. Analgesics ranked second, but sales declined in 2002 and remained static in 2003. Sales of digestives and intestinal remedies grew modestly in 2002 but declined marginally in 2003. FRANCE The OTC market in France is less developed than in some other countries. Because almost all residents have some form of complementary insurance that covers most of their prescription drug costs, they have little incentive to pay the full price for OTC products. Drug Registration and Classification Nonprescription medicines are divided into two categories in France: (1) sp´ecialit´es grand public (consumer products), which may be advertised to the public; and (2) products that are excluded from consumer advertising (e.g., codeine, nifuroxazide). It is possible to obtain most nonprescription medicines in France on prescription and to receive at least partial reimbursement for these drugs. However, nonprescription drugs that are prescribed may not be advertised to the public. Dietary supplements that make preventive or curative claims with regard to a human disease must satisfy the requirements for medicinal products and must receive a marketing authorization before they can be marketed.

180 TABLE 1. Number of Active Ingredients Available Over-the-Counter in Each Major Drug Class in the Leading Pharmaceutical Markets, October 2004 Drug Class Analgesics Antibacterials Antifungals Antihistamines Antivirals (topical) Corticosteroids Cough suppressants, expectorants, mucolytics Dermatologicals Gastrointestinal agents Lipid-regulating agents Local anesthetics Sympathomimetics Others Total

United States

France

Germany

Italy

Spain

United Kingdom

Japan

5 4 10 10 1 1 2 2 8 0 4 8 14 69

8 8 11 15 2 2 5 1 9 0 3 4 31 99

15 4 18 17 1 3 5 2 8 0 4 8 25 110

16 3 7 14 0 1 7 1 10 0 5 6 17 87

10 2 4 17 1 2 5 3 8 0 5 8 21 86

13 2 15 18 1 5 1 2 13 1 6 6 29 112

12 3 14 11 0 3 5 0 10 0 4 6 15 83

FRANCE

181

France’s drug registration process follows European Union (EU) standards (i.e., it is based on the fundamental principles of quality, safety, and efÞcacy). Approximately 20 widely used active ingredients, including acetaminophen, acyclovir, ibuprofen, ranitidine, and vitamin C, are covered by Þches substance (substance Þles), dossiers that provide exemption from the standard registration requirements. Most herbal medicines can be registered by means of an abridged procedure. Manufacturers of herbal medicines may generally choose a maximum of two indications from a list of “traditional therapeutic indications.” If a company wishes to market an herbal medicine that is not based on a traditional medicinal plant or is not to be used for a traditional therapeutic indication, it must conduct comprehensive clinical trials. Consumer Advertising Before a manufacturer may advertise a sp´ecialit´e grand public, it must obtain a visa grand public (consumer advertising permit) from the Agence Franc¸aise de S´ecurit´e Sanitaire des Produits de Sant´e (AFSSAPS; French Agency for the Safety of Health Care Products). This permit is valid for two years. Advertisements must be consistent with the terms of a drug’s marketing authorization and patient information leaßet. Comparative advertising to the public is not permitted in France. Distribution French law restricts the sale of medicinal products (including nonprescription drugs, herbal medicines, and homeopathic products) to France’s 22,689 pharmacies (as of 2002). Dietary supplements that are classiÞed as medicinal products are also restricted to pharmacy distribution. On the other hand, food supplements are technically excluded from sale in pharmacies, although in practice these products are widely available in French pharmacies. “Parapharmacies” are permitted to sell a limited range of health-related products (e.g., vitamins, minerals, antiseptics). These retail outlets are increasingly common in French supermarkets and as independent stores. Distance selling of pharmaceuticals is almost unknown in France. Under EU law, this trade is probably legal in France, but the law has yet to be put to the test. Pricing and Reimbursement In contrast to the strict controls on the pricing of prescription drugs, the OTC pricing environment in France is very liberal. Since July 1987, manufacturers have been free to set the prices of nonprescription medicines. Manufacturers may suggest a maximum recommended retail price, but they may not Þx the retail price or print prices on OTC packs. The Commission de la Transparence (CT; Transparency Commission) assesses drugs for their service m´edical rendu (SMR; medical beneÞt). Based on its degree

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OVERVIEW OF THE WORLD’S MAJOR OTC MARKETS

TABLE 2. Legal Status of Select Active Ingredients in the Major Pharmaceutical Markets, October 2004 Ingredient

United States

France

Germany

Italy

Spain

United Kingdom

Japan

Analgesics Codeine Diclofenac (oral) Ibuprofen Naproxen

Rx Rx OTC OTC

OTC Rx OTC Rx

Rx OTC OTC OTC

OTC OTC OTC OTC

Rx OTC OTC OTC

OTC Rx OTC Rx

OTC Rx OTC Rx

Antifungals Clotrimazole Terbinafine

OTC OTC

OTC OTC

OTC OTC

OTC Rx

OTC Rx

OTC OTC

OTC OTC

Antihistamines Cetirizine Loratadine

Rx OTC

OTC Rx

OTC OTC

OTC Rx

OTC OTC

OTC OTC

OTC Rx

Antivirals Acyclovir (topical)

Rx

OTC

OTC

Rx

OTC

OTC

Rx

Corticosteroids Beclomethasone Hydrocortisone

Rx OTC

OTC OTC

OTC OTC

Rx OTC

OTC OTC

OTC OTC

Rx OTC

Cough suppressants, expectorants, mucolytics Acetylcysteine Rx OTC OTC Carbocisteine N.R. OTC Rx Dextromethorphan OTC OTC OTC

OTC OTC OTC

OTC OTC OTC

N.R. Rx OTC

Rx OTC OTC

Gastrointestinal agents Loperamide OTC Omeprazole OTC Ranitidine OTC

OTC Rx OTC

OTC Rx OTC

OTC Rx Rx

OTC Rx OTC

OTC OTC OTC

OTC Rx OTC

Lipid-regulating agents Simvastatin Rx

Rx

Rx

Rx

Rx

OTC

Rx

Sympathomimetics Epinephrine

OTC

Rx

Rx

N.R.

Rx

Rx

Rx

Others Nicotine (nasal spray)

Rx

N.R.

Rx

N.R.

Rx

OTC

N.R.

N.R., not registered; OTC, over-the-counter; Rx, prescription-only.

of clinical utility, a drug is given an SMR rating of major, important, moderate, weak, or insufÞcient. In September 2003, the government unveiled plans for a three-stage program to dereimburse more than 600 products that have an “insufÞcient” SMR rating. In October 2003, the government dereimbursed a total of 84 products—analgesics, antacids, antibacterials, antiseptics, antispasmodics, combination antitussives, expectorants, muscle relaxants, and ophthalmic products. The second wave of dereimbursement was originally scheduled for July 2004 and the third for July 2005. The proposed second wave comprises 426 products considered “suitable to be used for self-medication”—mainly treatments for digestive disorders, mucolytics, magnesium, veinotonics, oligoelements, and plant-based medicines. The proposed third wave consists of “medicines that are

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183

not clinically useful but retain a certain social value because they have no more useful equivalent.” In February 2004, however, the government advised manufacturers of products that faced the possibility of dereimbursement in the second wave that the CT would reassess the medical beneÞt of each of these drugs before making a decision on dereimbursement. At this writing, it is unclear whether the government still intends to complete this initiative. Even if the second and third waves of dereimbursement are implemented, savings could prove to be much more modest than expected. A survey conducted by the market research company TNS Sofres in May 2004 found that, if their medications were dereimbursed, 72% of respondents would ask their physicians to prescribe similar drugs that were still covered. If these alternative drugs were more expensive than the dereimbursed medicines, the program could unintentionally increase drug reimbursement costs. This Þnding illustrates the difÞculties of persuading French consumers to use self-medication in place of reimbursable prescription drugs. Rx-to-OTC Switching Since 1996, 22 active ingredients—including acyclovir, cetirizine, diclofenac, famotidine, ibuprofen, ketoconazole, ketoprofen, levonorgestrel, loperamide, minoxidil, and ranitidine—have been switched to OTC status in France. Manufacturers generally initiate the switch process, although the government imposed the switch of the emergency contraceptive levonorgestrel. Market According to data from IMS Health, in 2003, the French OTC market was worth ¤1.572 billion ($1.775 billion) at retail prices, a 1.5% increase over the preceding year. (The U.S. dollar-to-euro exchange rate used in this report is the 2003 average rate: $1 = ¤0.8854.) However, the general trend in recent years has been downward. Cough and cold remedies, the best-selling drug category from 2001 to 2003, were the only one of the Þve categories whose sales grew. However, nicotine replacement therapies experienced even more vigorous growth: sales rose from $88.8 million in 2001 to $135 million in 2003, an increase of 52% in just two years. GERMANY The German OTC market is by far the largest in Europe. In 2003, OTC sales at retail prices totaled ¤4.257 billion ($4.808 billion), equivalent to 12% of the total pharmaceutical market. If prescription sales of nonprescription medicines (i.e., medicines that may be sold OTC or prescribed) are included, the nonprescription market was worth ¤7.068 billion ($7.983 billion), equivalent to 21% of the total pharmaceutical market. Recent reforms have boosted the OTC market but undermined prescription sales of nonprescription drugs.

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OVERVIEW OF THE WORLD’S MAJOR OTC MARKETS

Drug Registration and Classification All medicinal products marketed in Germany are governed by the Arzneimittelgesetz (Medicines Law). Herbal medicines are not distinguished from synthetic drugs. Vitamins and minerals may be classiÞed as medicinal products or foodstuffs, depending on their predominant purpose according to the common opinion or interpretation of the trade. The main criteria used to determine whether a vitamin or mineral product is classiÞed as a drug or foodstuff are composition, posology, and claims. A product’s dosage form, labeling, main retail outlet, and price may also inßuence whether the product is classiÞed as a medicine or a foodstuff. Because of Germany’s unusually large pharmacopoeia, the authorities there have struggled to comply with obligations to reregister all medicines in accordance with EU regulations. To expedite the reregistration process, in August 1994, the German government decided it would henceforth accept data on the “traditional use” of hundreds of herbal medicines, many of which are available OTC. Consumer Advertising Pharmaceutical companies may advertise nonprescription medicines to the public in all media. Advertisements must advise consumers they can obtain information on a drug’s risks and side effects by reading the patient information leaßet or consulting their physician or pharmacist. In addition, print advertising must carry the drug’s name, therapeutic indications, and (if applicable) warnings. With the exception of pricing information, comparative advertising to the public is not permitted under German law. Distribution Nonprescription medicines are divided into two categories in Germany, deÞned by their permitted channels of distribution. Apothekenpßichtige Arzneimittel (pharmacy-only medicines) may be sold only in the country’s approximately 21,700 retail pharmacies. Freiverk¨außiche Arzneimittel (free-sale medicines) can be purchased not only in pharmacies but also in some 5400 drugstores, 13,700 Drogeriem¨arkte (self-service drugstores), 2500 health stores, and 7500 other retail outlets (e.g., supermarkets, food stores). The recently enacted Gesetz zur Modernisierung der Gesetzlichen Krankenversicherung (GMG; Statutory Health Insurance Modernization Act) legalizes mail-order pharmacy in Germany, a practice that had previously been growing for several years without the blessing of the law. However, the GMG places strict conditions on mail-order pharmacies in Germany. These pharmacies must maintain the same high standards of quality and reliability as retail pharmacies. Trained personnel with qualiÞcations in pharmacy or medicine must be available to advise customers (e.g., by telephone). To deal with questions and problems, the mail-order pharmacy must be contactable around-the-clock. Medicines must be dispatched promptly and safely, and their quality and effectiveness must not deteriorate in transit. Drugs may be delivered only to the patient who ordered them or to a nominated representative.

GERMANY

185

Pricing and Reimbursement Manufacturers are largely free to set the prices of all medicines marketed in Germany. However, the German government is in the process of expanding the country’s reference pricing system to include many patent-protected drugs that are prescribed. Furthermore, in January 2003, the government imposed a 6% rebate on prescribed patent-protected drugs that were not subject to reference pricing. In 2004, this rebate was temporarily increased to 16% for prescription-only drugs outside the reference pricing system. (The rebate remained at 6% for prescriptions of nonprescription medicines.) OTC sales are exempt from the rebate. The GMG gives pharmacists in Germany the freedom to determine the retail prices of drugs sold OTC. This change is intended to reduce the prices for nonprescription medicines by promoting competition among pharmacies. Beginning January 1, 2004, the statutory health insurance system no longer reimburses the cost of most nonprescription medicines (i.e., drugs that are not prescription-only products) that are prescribed by physicians. However, the statutory health insurance system continues to cover the cost of nonprescription medicines that are prescribed to children under age 12, youths under age 18 who have development disorders, and patients who have serious conditions for which a nonprescription product is standard therapy. This reform has had an appreciable impact on the market for nonprescription medicines. According to IMS Health, in the Þrst six months of 2003, sales of nonprescription medicines totaled ¤3.41 billion ($3.851 billion) at retail prices. By comparison, in the Þrst six months of 2004, sales totaled ¤2.956 billion ($3.339 billion), a decline of 13.3%. OTC purchases increased by 8.6%, from ¤1.963 billion ($2.217 billion) to ¤2.131 billion ($2.407 billion), and private prescription sales grew by 47.4%, from ¤312 million ($352 million) to ¤459 million ($518 million). Conversely, sales within the statutory health insurance system fell from ¤1.135 billion ($1.282 billion) to ¤365 million ($412 million), a decline of 67.8%. Statutory health insurance funds have warned physicians not to try to circumvent the dereimbursement of nonprescription medicines by simply switching patients to similar prescription-only drugs that are still reimbursed. The government’s objective in dereimbursing nonprescription medicines is to force patients to bear a larger share of the cost of managing the less serious disorders that are typically treated by nonprescription medicines. Substituting a prescription-only drug, which may be considerably more expensive than the nonprescription product it replaces, undermines this cost-containment strategy. Physicians who switch patients from nonprescription medicines to prescriptiononly medicines without adequate justiÞcation—supported by documentary evidence—could be Þned. Rx-to-OTC Switching Germany has been one of the boldest exponents of Rx-to-OTC switching. Since January 1, 1996, the government has approved the switch of almost 20 active

186

OVERVIEW OF THE WORLD’S MAJOR OTC MARKETS

ingredients, including beclomethasone dipropionate, famotidine, fenticonazole, indomethacin, hydrocortisone acetate, miconazole, and ranitidine. Market The German OTC market has been static in recent years. Sales increased marginally in 2001, declined slightly in 2002, and grew modestly in 2003, reaching $4.808 billion. Over this three-year period, the market grew by just 1.7%, equivalent to 0.6% per year. Looking at the sales at retail prices of the Þve best-selling OTC drug categories in Germany from 2001 to 2003, cough and cold remedy sales experienced relatively strong growth, while sales of vitamins and minerals plummeted. The three other leading drug categories experienced little movement in their sales. ITALY OTC drugs have a relatively small share of the Italian pharmaceutical market. In 2003, they accounted for just 8.3% of total pharmaceutical sales. However, OTC sales are growing faster in Italy than in most other markets. Drug Registration and Classification In Italy, medicinal products are classiÞed as either prescription-only medicines or nonprescription medicines (i.e., drugs that can be obtained with or without a prescription). Nonprescription medicines include a category of products known as medicinali di automedicazione (self-medication drugs), products that the national health care system never reimburses. A drug can be classiÞed as a medicinale di automedicazione if it satisÞes the following criteria: • • •

• • •

It contains active ingredients or combinations whose efÞcacy and safety have been largely demonstrated. It has been widely used in therapy for at least Þve years in one or more EU member states. It is a treatment (typically offering minor symptomatic relief) for minor and temporary ailments with which patients can readily diagnose themselves without consulting a physician. It does not use a route of administration that requires the intervention of a health care professional (e.g., injection). Its dosage is within safety limits per dose and per pack. Its pack size is small and suitable only for short-term treatment.

Vitamins and minerals are classiÞed as “dietetic products” (i.e., foodstuffs) if they contain 0.3–1.5 times the recommended daily allowance (RDA) of the given dietary supplement. If their dosage exceeds 1.5 times the RDA or if they make medicinal claims, dietary supplements must be registered as medicinal products, following the standard procedure for obtaining a marketing authorization. Registration by reference or abridged registration are options for OTC products.

ITALY

187

Consumer Advertising Medicinali di automedicazione are the only medicinal products that may be advertised to the public. The Istituto dell’Autodisciplina Pubblicitaria (Institute for Advertising Self-Regulation) reviews OTC advertising in newspapers, magazines, and the radio prior to publication. No authorization is needed if a print advertisement contains a complete reproduction of the contents of the patient information leaßet and invites consumers to read the document. Distribution Italy’s 16,500 pharmacies have a monopoly on the distribution of medicinal products. Self-service within pharmacies is permitted for medicinali di automedicazione. Dietetic products may be sold in other retail outlets, but pharmacies remain the dominant channel of distribution for these products. Distance selling of medicinal products is illegal in Italy. Pricing and Reimbursement On January 16, 2003, the Italian government abolished the 50% reimbursement rate (known as “class B”) for pharmaceuticals and reassigned all drugs that had previously been in this category to either class A (100% reimbursement) or class C (no reimbursement). All nonprescription drugs belong to class C. Although manufacturers are free to set the prices of all drugs that are excluded from reimbursement without any prior authorization, they may increase their prices only once a year. At least 15 days before the price increase takes effect, the manufacturer must notify the Ministry of Health, the Comitato Interministeriale per la Programmazione Economica (CIPE; Interdepartmental Committee for Economic Planning), and the Federazione Ordini Farmacisti Italiani (FOFI; Federation of the Orders of Italian Pharmacists) of the proposed increase. Resale price maintenance is practiced in Italy, and medicines are sold for the same price throughout the country. Rx-to-OTC Switching The Italian pharmaceutical market was traditionally conservative about Rx-toOTC switching, but the authorities have become bolder in recent years. From 2000 to 2003, more than 20 compounds were switched to nonprescription status. This list includes cimetidine, dimethicone, ketoprofen (topical), minoxidil, lactulose, naproxen (topical), and piroxicam (topical). Drugs that are switched are automatically excluded from reimbursement. Market Unlike many of the other leading OTC markets, Italy’s market has grown steadily in recent years.

188

OVERVIEW OF THE WORLD’S MAJOR OTC MARKETS

Cough and cold remedies were the best-selling OTC drug category from 2001 to 2003 by far; their sales grew by 21.7% in just two years. In 2003, cold remedies accounted for 49.7% of sales in this category, cough remedies for 35.5%, sore throat remedies for 11.3%, and allergy treatments and other products for 3.3%. Digestives and intestinal remedies were the second-largest category in terms of sales in 2003. Laxatives accounted for 47.6% of total sales in this category, products for the stomach for 42.3%, antidiarrheals and electrolytes for 8.9%, and products for the liver for 1.2%. In the analgesics category, products for muscular pain accounted for 55.2% of total sales; general analgesics, 33.9%; products for oral pain, 2.6%; dysmenorrhea treatments, 2.3%; products for headaches, 0.7%; and other generic analgesics, 5.4%. Sales of OTC vitamins and minerals in Italy grew by 26.3% between 2001 and 2003. Mineral supplements accounted for 24.2% of 2003 sales in this category; multivitamins with minerals, 23.6%; vitamin B products, 20.8%; vitamin C products, 15.2%; and other products, 16.2%. In the dermatologicals category, antiseptics accounted for 26.8% of 2003 sales; antimycotics, 23.8%; treatments for skin irritations, 23.4%; wound treatments, 17.1%; and other products, 8.9%. SPAIN Spain is, by a substantial margin, the smallest OTC market among the seven pharmaceutical markets under study, but in recent years sales have grown much faster than in other countries. In 2003, OTC drugs accounted for 8.7% of total pharmaceutical sales in Spain (at retail prices). Drug Registration and Classification In Spain, nonprescription medicines can be divided into three main categories: • • •

Products that are reimbursed if prescribed by a physician but not reimbursed if purchased OTC. These products cannot be advertised to consumers. Products that are not reimbursed and cannot be advertised to consumers. Especialidades Farmac´euticas Publicitarias (EFPs; advertised pharmaceutical specialties), which are products that are not reimbursed but may be advertised to consumers.

Vitamins and minerals are classiÞed as food supplements if they do not exceed the RDA and do not make medicinal claims (e.g., prevention of conditions or a deÞciency). If they do exceed the RDA or make medicinal claims, they are classiÞed as EFPs. Spanish registration requirements for nonprescription medicines are generally similar to those for prescription drugs. The one major difference is that the data on pharmacological, toxicological, and clinical studies that are required for new prescription medicines may be replaced by bibliographic documentation.

SPAIN

189

The Spanish regulatory authorities generally do not favor combination therapies. Manufacturers are required to justify combinations, which may not contain more than two or three active ingredients (except for vitamins and minerals). Consumer Advertising As noted in the preceding section, EFPs are the only medicines that may be advertised to Spanish consumers. The Ministry of Health vets all advertising copy prior to publication. The Asociaci´on Nacional de Especialidades Farmac´euticas Publicitarias (ANEFP; National Association of Advertised Pharmaceutical Specialties), which represents manufacturers of EFPs, also reviews OTC advertisements before they are published and has long urged the Spanish government to introduce a system of industry self-regulation of OTC advertising. In 2003, self-medication manufacturers spent a total of ¤205.5 million ($232.1 million) on OTC advertising in Spain; 80% of that investment was allocated to television advertising. Distribution The 20,348 pharmacies in Spain have a monopoly on the sale of all medicines, including OTC products. Self-service is not permitted in Spanish pharmacies. Distance selling of pharmaceuticals is illegal in Spain. Pricing and Reimbursement In contrast to the strict controls on the pricing of prescription-only drugs in Spain, nonreimbursed OTCs can be priced at the manufacturers’ discretion. Spain practices resale price maintenance to ensure uniform pricing in all pharmacies. Rx-to-OTC Switching The ofÞcial positive list of EFPs is updated annually. A commission of representatives of the Ministry of Health and the ANEFP decides which drugs are suitable for Rx-to-OTC switching. Manufacturers generally submit individual switch applications, but the ANEFP can request the general switch of a particular ingredient if such an action would be in the interest of multiple member companies. Rx-to-OTC switching automatically precludes a drug from reimbursement by the Spanish health care system. Market The Spanish OTC market has grown vigorously in recent years. Sales at retail prices rose from $896 million in 2000 to $1.212 billion in 2003, an increase of 35.3% in three years, equivalent to 10.6% per year. Analgesics were the best-selling category in the Spanish OTC market in 2003, growing 14% in two years. Cough and cold remedies ranked second in terms of sales but achieved a slightly faster growth rate (15.4% in two years). Sales of dermatologicals also grew rapidly, whereas sales of digestives and intestinal remedies and vitamins and minerals remained static in 2002 and 2003.

190

OVERVIEW OF THE WORLD’S MAJOR OTC MARKETS

UNITED KINGDOM The United Kingdom has Europe’s second-largest OTC market in monetary terms, totaling £ 1.973 billion ($3.222 billion) in sales in 2003. (The U.S. dollarto-pound-sterling exchange rate used in this article is the 2003 average rate: $1 = £0.61229.) Self-medication accounts for a signiÞcantly higher proportion of total pharmaceutical sales in the United Kingdom (17.2% in 2003) than in any of the other major pharmaceutical markets under study. Drug Registration and Classification In the United Kingdom, medicines are assigned to one of three categories: •

•

•

Prescription-only medicines (POM$) require a prescription from a medical professional and may be dispensed only by pharmacies. These products are listed in the Prescription-Only Medicines (Human Use) Order, 1997. General Sale List (GSL) medicines may be sold in any lockable store. The General Sale List (The Medicines [Products Other than Veterinary Drugs] [General Sale List] Order; 1984—SI 1984/769) catalogues medicinal ingredients deemed suitable for distribution outside pharmacies, “where the hazard to health, the risk of misuse, or the need to take special precautions in handling is small and where wider sales would be a convenience for the purchaser.” GSL products include small packs of analgesics (e.g., acetaminophen, acetylsalicylic acid, ibuprofen), most antacids, simple cough remedies, antiseptics, and throat lozenges. Pharmacy medicines (P) are drugs that are neither POM nor GSL products. Pharmacy medicines are available without a prescription but may be sold only in pharmacies, under the supervision of the pharmacist. These products include larger packs of analgesics, cough remedies that contain cough suppressants, medicines that contain antihistamines or decongestants, and products that have recently been switched from prescription-only status. (In the United Kingdom, Rx-to-OTC switching is often called the “POM-toP” process.)

Herbal medicines may be assigned either P or GSL status. The United Kingdom does not have an ofÞcial list of plants that are considered medicinal products by function. The Medicines and Healthcare Products Regulatory Agency (MHRA, formerly the Medicines Control Agency (MCA)) deems plants to be medicinal by function if they have a pharmacological effect at the supplied dose and have no recognized nonmedicinal use. The MHRA supports an EU initiative to facilitate the registration of herbal and other traditional remedies on the basis of traditional use of these ingredients anywhere in the world. Vitamins and minerals are generally classiÞed as foodstuffs, regardless of their dosage. They can be licensed only if they can support a medicinal claim (e.g., correction of a deÞciency). The United Kingdom has no legally deÞned limits for vitamin dosing, but products should fall within recognized safety limits.

UNITED KINGDOM

191

Manufacturers are not permitted to make medicinal claims for foodstuffs, but they have considerable latitude to make health claims. Consumer Advertising Any nonprescription (i.e., P or GSL) medicine may be advertised to U.K. consumers in any media, even if the product can be prescribed and reimbursed by the National Health Service (NHS). Television, radio, print, Internet, and pointof-sale advertisements must all include the instruction: “Always read the label.” The United Kingdom’s OTC industry operates a system of self-regulation through the Proprietary Association of Great Britain (PAGB), the organization that represents self-medication manufacturers. PAGB members submit their advertisements to the agency for review prior to publication. In addition, media-speciÞc authorities review advertisements before they are published. In 2004, the PAGB, in collaboration with the MHRA, published new guidelines on the advertising of OTC drugs that are indicated for relatively serious conditions. The guidelines include the following advice: •

•

•

Advertising shall not discourage individuals from seeking advice from a health care professional. Advertisements should include information concerning the availability of professional advice, for example “Ask your pharmacist.” Advertisements for medicines with a speciÞc need for ongoing monitoring or review by an appropriately qualiÞed health care professional should make this clear. Advertising should not encourage people to discontinue the use of a prescribed medicine.

The new guidelines also contain speciÞc advice on the advertising of certain drug classes that either have been switched or are candidates for Rx-to-OTC switching (e.g., acetylsalicylic acid, statins and other antihyperlipidemics, nitrates for cardiovascular disorders and other diseases, such as asthma). The guidelines will be updated to reßect future Rx-to-OTC switches for serious conditions. Unlike most other European countries, the United Kingdom permits limited comparative advertising of OTC drugs. The condition is that speciÞc brands or competitors not be mentioned or be identiÞable in the advertisements. Distribution As of December 31, 2003, the United Kingdom had 12,198 pharmacies. Many of these pharmacies belong to extensive chains, including Lloyds (1322 outlets), Boots the Chemist (1304 outlets), and Moss (789 outlets). Pharmacies accounted for approximately 73% of all OTC purchases in 2003. Other than pharmacies, the main retail outlets for GSL products are grocery stores (including supermarkets), drugstores, and health food stores. Self-service of GSL medicines is permitted in all retail outlets that stock these products. Distance selling of GSL products is allowed in the United Kingdom but is uncommon.

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Pricing and Reimbursement The United Kingdom regulates the prices of branded prescription-only drugs by means of the Pharmaceutical Price Regulation Scheme. In contrast, nonprescription medicines are treated as consumer products and are therefore exempt from statutory price controls. Manufacturers are free to set prices as they see Þt. In May 2001, the United Kingdom abolished resale price maintenance for nonprescription drugs. Some retail outlets have used this new freedom to cut the prices of certain OTC drugs by as much as 50% at times, in an effort to attract additional business. The NHS reimburses a wide range of nonprescription drugs if these products are prescribed by a physician. (Patients have to pay a prescription charge—currently £6.40 ($10.45) per prescription item—unless they qualify for exemption.) In an effort to contain soaring prescription drug expenditures, in 1985, the Department of Health (DH) introduced the NHS Selected List, a catalog of products that could no longer be prescribed at NHS expense. The original list included analgesics for mild to moderate pain, antacids, benzodiazepines, cough and cold remedies, laxatives, and vitamins, tonics, and bitters. In August 1993, the DH dereimbursed a number of topical antirheumatics. In September 1994, four more drug categories were added to the Selected List: antidiarrheals, drugs used for anemia, drugs acting on the ear and nose, and topical corticosteroids. Rx-to-OTC Switching The United Kingdom has long been one of the foremost exponents of Rx-to-OTC switching. Since 1983, the DH has approved the switch of approximately 70 active ingredients, including acyclovir, cetirizine, cimetidine, famotidine, ketoprofen (topical), levonorgestrel, loperamide, loratadine, minoxidil, nicotine, omeprazole, piroxicam (topical), pseudoephedrine hydrochloride, and several antifungals. On April 1, 2002, the DH introduced a streamlined process for Rx-to-OTC switching. Approval times for Rx-to-OTC switching were reduced from approximately 18 months to 6–8 months. The government indicated that more than 50 prescription-only medicines could be switched to OTC status over the next Þve years. Manufacturers will be offered a three-month marketing exclusivity period as an incentive to pioneer compound switches. Most of the proposed switches are for indications previously considered unsuitable for self-medication (e.g., stable angina, hypertension, prevention of stroke/ myocardial infarction, chronic obstructive pulmonary disease, erectile dysfunction). However, if these drugs are switched, they would be subject to strict safeguards, including, in most cases, diagnosis by a physician and supervision of the sale by a pharmacist. Indeed, these switch candidates may represent a new category of drugs—a bridge between prescription-only medicines and true OTC products. On July 29, 2004, the United Kingdom became the Þrst country in the world to license an OTC statin. Zocor Heart-Pro, marketed by Johnson & Johnson MSD, is a 10-mg dosage of simvastatin, the most widely prescribed statin in

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the United Kingdom. The suggested retail price for a 28-tablet pack is £ 12.99 ($21.22), but pharmacy chains entered into an immediate price war. For example, the Asda supermarket chain, owned by Wal-Mart, is selling the product for £ 8.98 ($14.67), a 31% reduction on the suggested retail price. Reports indicate that, thanks to a large advance purchase by one major national pharmacy chain, Johnson & Johnson MSD achieved its sales target for 2004 by the end of August. However, it remains to be seen how consumers will respond to an OTC statin in the long term. Until very recently, selective cyclooxygenase (COX)-2 inhibitors appeared to be one of the most promising drug classes in terms of switch potential. However, the recent withdrawal of rofecoxib (Merck & Co.’s Vioxx) from the market and concerns about the safety of other selective COX-2 inhibitors will make it very difÞcult to switch drugs in this class. Market The U.K. self-medication market has experienced steady growth in recent years. Analgesics, which ranked Þrst in terms of sales of the main drug categories from 2001 to 2003, grew by 14.5% in two years. Dermatologicals and digestives and intestinal remedies experienced even more vigorous growth—in excess of 20% in two years. By comparison, sales of cough and cold remedies grew only slowly, and the market for vitamins and minerals remained static. JAPAN The Japanese OTC market is the second-largest in the world, surpassed only by the U.S. market. According to Japan’s Ministry of Health, Labor, and Welfare (MHLW), OTC drugs accounted for 14% of the total pharmaceutical market in Japan in 2002 (the most recent year for which data are available). Drug Registration and Classification Japanese law deÞnes prescription drugs as “medicinal products supplied for the purpose of being administered by physicians or dentists or of being used based on either the prescriptions or the directions of physicians or dentists.” Nonprescription medicines are “the medicinal products other than the prescription medicines.” Kampo products (i.e., traditional herbal medicines) have a long tradition in Japan, but sales have declined sharply in recent years. These products may be classiÞed as medicines if they satisfy the requirements of the Pharmaceutical Affairs Law (PAL). Vitamins and minerals may be classiÞed as OTC drugs, quasi-drugs, or foodstuffs. Manufacturers or importers of medicines must obtain both an approval and a license before they are permitted to market their products in Japan. The approval certiÞes the safety and efÞcacy of the product, while the license conÞrms that the manufacturer or importer meets the PAL’s standards for the sale of medicines.

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Consumer Advertising All nonprescription medicines may be advertised in any media in Japan, in the same way as other consumer goods. The self-medication industry subscribes to the Voluntary Code for OTC Medicine Advertising, a standard of practice that has the MHLW’s endorsement. This code sometimes requires that advertisements direct consumers to “read carefully the precautions for use before you take this medicine.” OTC advertisements are reviewed after they are published or broadcast. The Advertising Review Board, a committee that consists of representatives of the Japan Self-Medication Industry (JSMI) and independent experts, meets every other month to review all OTC advertisements that have appeared on television, radio, newspapers, and magazines. Another body, the Post-Review Committee, which comprises representatives of the MHLW and prefectural governments, meets three times per year to review OTC advertising. In 2003, the Japanese self-medication industry spent
181.3 billion ($1.563 billion) on advertising. (The U.S. dollar-to-yen exchange rate used in this article is the 2003 average rate: $1 =
157115.97995.) Television advertising accounted for 71.6% of this investment, newspapers for 17.2%, magazines for 6.1%, and radio for 5.1%. Distribution Japan’s Þve channels of distribution for medicines are as follows: • •

•

•

Yakkyoku (pharmacies) are the only outlets permitted to dispense prescriptions. As of March 2003, Japan had 49,332 pharmacies. Ippan-hanbai-gyo (drugstores with a pharmacist) may sell any medicines except prescription drugs. In March 2003, 12,397 such stores were active in Japan. Certain drugs may be distributed by mail order. Yakushusho-hanbai-gyo (drugstores without a pharmacist) must be managed by a person who has passed the Yakushusho QualiÞcation Test that is administered by prefectural governments. These stores are forbidden to sell prescription drugs and certain nonprescription drugs (as speciÞed by the MHLW). Japan had 15,151 of these stores in March 2003. Haichi-hanbai-gyo (household distribution) is a long-established Japanese custom of selling nonprescription medicines from door to door. Prefectural governors determine which medicines may be sold by this method, and a license is needed to engage in this trade. In March 2003, 11,440 people had licenses for household distribution.

Health care products that are not classiÞed as medicines are not subject to retail restrictions and may be sold anywhere. Pricing and Reimbursement Because nonprescription medicines are excluded from reimbursement in Japan, manufacturers have complete freedom in setting the prices of these products. Resale price maintenance is not practiced in Japan.

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Rx-to-OTC Switching Far fewer active ingredients have been switched to OTC status in Japan than in most European markets. In part, this situation reßects the fact that many drugs used in Europe are not marketed in Japan. However, the Japanese authorities have also been more conservative about the switch process. In 1997, three H2 antagonists (cimetidine, famotidine, and ranitidine) were switched to OTC status in Japan, as was the eye lotion sodium cromoglycate. The following year, sofalcone (a gastrointestinal remedy) was granted OTC status. In 1999, minoxidil was launched directly as an OTC treatment for male pattern baldness without ever having been marketed in Japan as a prescription therapy for this indication. In 2000, teprenone (a gastrointestinal remedy) was switched, and in 2001, nicotine gum (for smoking cessation) was granted OTC status. In 2002, pranoprofen (an antimicrobial eye lotion) and four athlete’s foot remedies (butenaÞne, amorolÞne, neticonazole, and terbinaÞne) were switched. No drugs were switched in 2003. Market Although the other major OTC markets have been relatively stable or experienced some growth in recent years, the Japanese self-medication market has been in steady decline. The Japanese OTC market also differs from the other major self-medication markets in terms of composition. In Japan, vitamins and minerals dwarfed all other categories. In 2002, sales of vitamins and minerals were more than double those of the next largest category, cough and cold remedies. The enormous success of vitamins and minerals is attributable to a peculiarity of the Japanese market: the popularity of OTC drinks and mini drinks—tonics that contain a mixture of vitamins, minerals, traditional medicinal herbs, and (in some cases) animal extracts. On the other hand, the market for systemic analgesics is much smaller in Japan than in the other countries. This situation reßects the fact that topical analgesics (e.g., patches) are much more widely used in Japan than in the other major OTC markets. OUTLOOK Although the worldwide OTC market has not grown as fast as the prescription drug market in recent years, it has strong prospects for future success. Governments in most countries want to stimulate demand for self-medication as a way of reducing their expenditures on prescription medicines. To that end, governments are likely to become much more radical in their attitude toward Rx-to-OTC switching. In the past, OTC medicines were generally reserved for self-limiting, relatively trivial diseases, but in the future, governments will probably approve the use of OTCs for more serious disorders and chronic conditions. To allay concerns about the safety of expanding the range of OTC indications, governments may

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introduce certain safeguards (e.g., initial diagnosis or prescription by a physician, mandatory consultation with a pharmacist, regular monitoring). Self-medication may also play an increasingly important role in disease management and other preventive strategies. The U.K. government has set a bold precedent by making low-dose simvastatin available OTC. It will be interesting to see if the United Kingdom switches some of the other drugs on its highly ambitious list of potential candidates and if other governments follow suit. Pharmaceutical companies may also show more interest in the OTC market in the future, especially if governments and other payers signal their will to stimulate greater use of OTC products. Manufacturers may be able to fend off generics competition by switching a branded prescription drug to OTC status before its patent expires. By making a drug more accessible, OTC status may also boost sales by attracting new users. In increasingly afßuent societies, the convenience of self-medication may be a considerable competitive advantage.

Biotech Hubs—Key Growth Engines for Drug Development

SUMMARY Although pharmaceutical industry research expenditure continues to grow, success rates for new drugs have fallen dramatically in the past Þve to seven years. A greater proportion of pharmaceutical research and development has therefore migrated to biotech “hubs,” or regional clusters of innovation where biotech-related activity tends to congregate. These hubs offer an especially fertile environment for development of biotech products because they provide strong academic research infrastructures, entrepreneurial spirit, an abundance of skilled labor, and government incentives to encourage further growth. The U.S. biotech industry is highly concentrated within the nine largest hubs, and pharmaceutical companies are positioning themselves to share in the opportunity for growth that these hubs offer. This article explores the phenomenon of biotech clustering, reviews key development factors that have fostered the success of the top nine hubs, and discusses implications for the pharmaceutical industry. BUSINESS IMPLICATIONS •

As a greater proportion of pharmaceutical R&D migrated to biotech, several biotech “hubs” emerged; these hubs are regional clusters of innovation where biotech-related activity congregates. They offer a fertile environment for development of biotech products because they provide strong academic research infrastructures, entrepreneurial spirit, an abundance of skilled labor, and government incentives to encourage further growth.

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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BIOTECH HUBS—KEY GROWTH ENGINES FOR DRUG DEVELOPMENT

The degree to which biotech-related activity in the United States has concentrated in the nine largest biotech hubs is staggering. They claim more than three-Þfths of all National Institutes of Health (NIH) spending on R&D, and nearly two-thirds of all biotech-related patents are held by academics or corporations located in these hubs. Statistics indicate that the nine key hubs account for three-fourths of the nation’s largest biotechnology Þrms, more than $8 of every $9 of venture capital invested in biotech, and more than 95% of all biotech research alliances. The emergence of biotech hubs and their growing social and economic impact on the health care industry attract much attention from state and local government. The U.S. Economic Development Association reports that 83% of local development agencies have made biotech one of their top two development priorities, and 41 states have drafted formal programs that outline speciÞc targets and strategies to encourage biotech growth. Although all successful biotech hubs share common traits, each of the nine recognized leaders in the United States has been driven by a different set of strengths and advantages. Among the top nine hubs, Boston and San Francisco stand out as the strongest biotech regions by a wide margin. Besides having “Þrst mover” advantage, these cities offer proximity to unusually active communities of world-class research institutions and rich sources of Þnancing and venture capital.

INTRODUCTION Innovation has made the pharmaceutical industry one of the most productive and proÞtable of all business sectors. Increasingly, however, many pharmaceutical companies face short-term operating pressures because the return on their research investment is declining. With current trends predicting a widening gap between research spending and NCE approvals, many pharmaceutical companies are now looking to the biotechnology sector to help bolster innovation and productivity in the research arena. Industry sources suggest that as much as 55% of products currently marketed by the top 20 pharmaceutical companies are inlicensed from biotech Þrms. Furthermore, products licensed or acquired from biotech companies have been gaining more FDA approvals in recent years than products developed through internal R&D efforts at “big pharma” companies. According to the Pharmaceutical Research and Manufacturers of America (PhRMA), of the 35 new medicines approved by the FDA in 2003, 14 were biologics. Few would dispute that biotech has only started to deliver on its full promise. According to the U.S. Department of Commerce, biotech R&D expenditure in 2001 accounted for approximately 10% of all U.S. industry R&D. This Þgure is likely to increase signiÞcantly over the next decade; consequently, the biotech industry’s inßuence on the changing nature of the research-based pharmaceutical industry will be substantial.

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As a greater proportion of pharmaceutical R&D migrated to biotech, several biotech “hubs” emerged; these hubs are geographical clusters of innovation where biotech-related activity tends to congregate. These hubs, which include Boston Cambridge, San Francisco/Bay Area, and San Diego, are fertile environments for development of biotech products because they offer strong academic research infrastructures, entrepreneurial spirit, an abundance of skilled labor, and government incentives to encourage further growth. The maturation of the biotech industry confers ever-growing importance on the hubs, which promise to be key growth engines for drug development in the future. Leading companies such as PÞzer, Merck, Aventis, Novartis, and AstraZeneca have already made aggressive moves to better position their operations within these hubs. The pharmaceutical industry as a whole must take notice and examine the evolving biotech landscape in order to plan strategically and share in this new opportunity for growth. In this article, we look at the nine major biotech hubs in the United States and consider the strengths and weaknesses of each. THE CLUSTERING PHENOMENON IN THE UNITED STATES A key 1998 paper by Harvard Business School professor Michael Porter deÞnes clusters as “concentrations of interconnected companies and institutions in a particular Þeld.” Why is the biotech industry concentrated geographically in certain hubs of activity? Although the emergence of biotech hubs is relatively new, the clustering phenomenon is not unique to biotech and has been well studied in such diverse Þelds as entertainment, Þnance, information technology, plastics, and aerospace and defense, among many others. Once a cluster begins to form (see the following section, “Key Factors in Development of a Biotech Hub”), a chain of events perpetuates its growth. The fast-paced sharing of information and ideas between academic and industry researchers increases the productivity of companies based in the area and drives the direction of innovation. This dynamic induces workers with ideas or relevant skills to migrate to the cluster from other regions, further expanding the pool of available talent. Companies compete for this labor, and the transfer of skilled workers, knowledge, and expertise bolsters the productivity of the area. Meanwhile, entrepreneurs take notice and either branch out from existing academic or corporate footholds or migrate to the region from other areas. All these factors stimulate the formation of new businesses, which expands and strengthens the cluster. Certain regions have already managed to capitalize on the biotech wave more successfully than others. In a 2002 survey of the nation’s 51 largest metropolitan areas, the Brookings Institution Center on Urban and Metropolitan Policy found that biotechnology centers in the United States were highly concentrated within nine key regional hubs: Boston, San Francisco, Research Triangle (North Carolina), San Diego, Los Angeles, New York/New Jersey, Washington, DC, Seattle, and Philadelphia.

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The degree to which biotech-related activity in the United States has concentrated in the largest hubs is staggering. They claim more than three-Þfths of all National Institutes of Health (NIH) R&D spending, and nearly two-thirds of all biotechnology-related patents are held by academics or corporations located in the nine key hubs. Statistics further indicate that the nine key hubs account for three-fourths of the nation’s largest biotech Þrms as well as three-fourths of the biotech Þrms formed in the past decade. This Þnding reveals that the biotech hubs that were coincidental locations for the earliest pioneering companies continue to be sources of innovation today. Biotech opportunities in these favorable business environments have attracted the Þnancial community as well. More than $8 of every $9 of venture capital invested in biotech is invested in the nine key hubs. According to a Stanford University study that examined investment during the period 1988–1999, more than half of all U.S. biotech Þrms received locally based venture funding. This Þnding suggests that Þnanciers themselves either migrate or set up operations in close physical proximity to biotech hubs, thereby providing additional beneÞts to local economies. Perhaps most important to the pharmaceutical industry is the fact that these biotech hubs attract the vast majority of partnering activity. More than 95% of all research alliances, including academic and industry licensing deals, occur within the nine largest biotech hubs. KEY FACTORS IN DEVELOPMENT OF A BIOTECH HUB Much attention has focused on the speciÞc conditions necessary to “seed” a future hub of biotech activity. Most academics believe that in order to achieve a successful clustering effect, a hub must offer a high degree of regional biotech competitiveness as well as a high innovation capability. Several key factors are necessary to create these conditions; we discuss each factor in the following subsections. Basic Research Presence Universities and specialized research centers are the driving forces behind innovation in every leading biotech region. Active federal funding, including NIH grants, a large number of patents granted, and an abundant supply of doctorate holders are all indicative of research strength within a particular region. Strong Physical and Information Infrastructure To facilitate communications with universities, specialized research institutes, and other potential collaborators, biotech companies need physical space that puts them in close proximity with these institutions. Also, location among other biotech companies is crucial to encourage competition for talent. Additionally, a low cost of doing business (wages, real estate, utilities), observed especially in some of the newer clusters, is a stimulus to hub development.

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Educated Workforce A strong educational system is necessary to develop local talent and attract outside talent. In the workforce itself, specialized talent and training are considered more critical than an abundance of labor. Quality universities, community colleges, and training programs often funnel talent and specialized skills into the local community. Mechanisms for Commercialization An entrepreneurial spirit and a strong Þnancial infrastructure are necessary if innovation is to translate to economic success. The availability of venture funds and capital markets is essential in helping start-up companies compete with established Þrms. Open communication with established Þrms is necessary to encourage research and development partnerships and product licensing opportunities. Support of State and Local Government The emergence of biotech hubs has attracted much attention from both state and local government. The U.S. Economic Development Association reports that 83% of local development agencies have made biotech one of their top two development priorities, while 41 states have drafted formal programs that outline speciÞc targets and strategies designed to encourage biotech growth. Favorable tax laws, research and development incentives, state-sponsored corporate recruitment, and subsidized training programs are all examples of ways in which a regional government can help nurture its biotech community. For biotech companies, the most important criterion in choosing a location is access to research institutes with good technology transfer policies. In some states, therefore, high-powered coordinating bodies that include key public ofÞcials and industry leaders have worked to facilitate collaboration and networking between industry, local academic institutions, and state agencies. The Biotechnology Industry Organization (BIO), a national coalition that comprises more than 1000 companies, academic institutions, and state and federal afÞliates, recently compiled a list of recommendations for local governments that describes best practices to support and encourage growth in the biotechnology industry. An expanding biotech infrastructure creates economic development opportunities for a region in a number of ways. Most directly, corporations contribute signiÞcant tax revenues when they base their operations in a particular region. In addition, the presence of academic and industry research laboratories, hospitalbased collaborations, and manufacturing facilities generates new jobs, often at high wages that reßect the specialized skills required. Furthermore, industry experts estimate that for every direct job created by biotechnology, roughly two additional indirect jobs are created in support services, such as business supplies and legal services.

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To date, however, even regions that have been most successful in recruiting the biotech industry have realized only modest returns to their overall economy. Most biotech Þrms are quite small. Nationally, according to the Institute for Biotechnology Information, only 44 Þrms have more than 1000 employees. Furthermore, because Þrms may outsource the production, marketing, and distribution of successful products, some potential beneÞts could be exported from the local community. However, the biotech industry is growing rapidly and expanding its capabilities. The urgency to incorporate biotech into local economic development plans lies in the widespread belief that biotech will be a powerful force in the future economy. LEADING BIOTECH HUBS Although all the successful biotech hubs share common traits, each one of the nine recognized leaders in the United States has been driven by distinct strengths and advantages. Within the top nine, Boston and San Francisco stand out as the strongest biotech hubs by a wide margin. In addition to having “Þrst mover” advantage, these cities offer proximity to unusually active communities of worldclass research institutions and to rich sources of Þnancing and venture capital. Other regions, such as New York and Philadelphia, have successfully built up their biotech capabilities on a local base of pharmaceutical industry leadership. North Carolina, San Diego, and Seattle have been aggressive in recruiting the industry. The greater Washington, DC, area has been helped by federal funding and its close ties to the NIH. Los Angeles is best known in the biotech industry as the birthplace of Amgen, the leading biotech company. The following sections highlight some of the key strengths and weaknesses of the top nine biotech hubs, including current issues and future challenges. Boston The biotech hub that has developed in Boston (and in nearby Cambridge), Massachusetts, is one of the key strongholds of the global biotech industry. BeneÞting from its close proximity to world-class research institutes, the hub is well networked with renowned universities such as Harvard and the Massachusetts Institute of Technology (MIT), as well as a plethora of afÞliated laboratories, specialized research centers, and leading hospitals, particularly in and around Boston’s Longwood medical area. These resources provide a solid foundation for basic scientiÞc and clinical research. Given its educational and basic research infrastructure, it comes as no surprise that the Boston hub stands out as a leader in developing scientiÞc talent. With more than 120 colleges and universities in Massachusetts, the Boston area produces more than 350 new doctorates each year and employs nearly 5000 life scientists. These numbers rival the totals of the San Francisco and Research Triangle biotech hubs combined. Recent history demonstrates that Boston is a leader not only in biotech research but also in commercialization. While leading all biotech hubs nationwide in total

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amount of NIH funding, academia and industry in Massachusetts have also forged more alliances than any other hub. The value of these alliances accounts for nearly two-thirds of the total deal value of all biotech hubs combined. Since its inception, the Massachusetts biotech industry has brought approximately 40 new products to market, including drugs, biomedical devices, and commercially available diagnostic tests. Of the 24 drugs approved by the FDA in 2003, one-third were developed by Massachusetts companies. High-proÞle drug approvals include alefacept (Biogen Idec’s Amevive), bortezomib (Millennium Pharmaceuticals’ Velcade), daptomycin (Cubist Pharmaceuticals’ Cubicin), agalsidase beta (Genzyme’s Fabrazyme), and laronidase (Genzyme’s Aldurazyme). Another key strength of the Massachusetts biotech industry is a strong central organization that supports many of the hub’s key activities. The Massachusetts Biotechnology Council (MBC) is the oldest and second-largest biotechnology association in the world. A nonproÞt organization composed primarily of inßuential biotech industry leaders, the group’s stated goals include the following: • • • •

Providing joint activities and services, such as consortium purchasing. Inßuencing legislative and regulatory policies. Strengthening industry’s community. Building public support for biotechnology.

A joint study conducted by the MBC and the Boston Consulting Group (MassBiotech 2010: Achieving Global Leadership in the Life-Sciences Economy), issued in 2002, found that employment in and around the Boston biotech hub grew at an annual rate of 10% between 1996 and 2001. The study projects that the total number of biotech jobs could increase from 30,000 in 2001 to 100,000 by 2010, provided that the state exploits the industry’s full potential. More than 80% of the 280 biotech companies in the state were founded in Massachusetts. The rest are out-of-state companies drawn by the region’s dynamic history and expectation of further growth. Most signiÞcantly, the list of companies now moving to the Boston hub includes many of the world’s largest biopharmaceutical companies. AstraZeneca, Merck, Wyeth, Amgen, and PÞzer, for example, have all begun to locate major research facilities in the metropolitan Boston area. One of the most aggressive investments has been made by Novartis, which has relocated its global R&D headquarters to Cambridge. Two Novartis facilities are expected to employ as many as 900 people and to contribute $4 billion to the Boston/Cambridge area economy by 2010. While the Boston hub has long been a hot spot for biotech research, it has not been a major center for clinical development or manufacturing. Many small biotech Þrms in the area lack the capability to conduct clinical trials, and the cost of setting up a manufacturing facility is prohibitive, given the high price of real estate in the region. Therefore, companies involved in these activities tend to move most of this work out of state. For example, Biogen Idec, one of the state’s oldest and largest biotech Þrms, elected to build the world’s second-largest manufacturing plant in North Carolina rather than locate these facilities with its R&D headquarters in Massachusetts.

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Because of widespread perceptions that Boston is an expensive place to do business, the region is challenged to persuade its own companies to stay within its borders. However, should the region start to retain functions such as manufacturing, the payoff would be substantial. Manufacturing jobs tend to be highly stable because the FDA requires approval of the manufacturing process and site as part of its overall drug-approval process. Therefore, little chance exists that a manufacturing operation would be moved once its product has been launched because the costs would be substantial. San Francisco With a rich tradition of science and technology innovation, the San Francisco Bay area has often been credited with nurturing the biotech industry in its earliest years. In the 1970s, research institutes such as Stanford University and the University of California at San Francisco and at Berkeley (UCSF and UCB) were among the Þrst to receive seed funding from the federal government to explore genetic engineering and other foundations of the biotech paradigm. In 1976, Genentech, the world’s Þrst biotechnology company, was founded in San Francisco through a partnership involving UCSF biochemist Herbert Boyer and venture capitalist Robert Swanson. The company made an initial public offering (IPO) in 1980 and generated $35 million; it was the Þrst of many biotech company IPOs made during that decade. Genentech was also the Þrst biotech company to manufacture and market a biotechnology drug—Protropin, a recombinant version of human growth hormone that was approved by the FDA in 1985. Recognizing San Francisco’s key role in the emergence of biotechnology, BIO hosted its 2004 annual meeting in the city under the title, “Where It All Began.” The event was the largest life sciences meeting in the industry’s history and drew record attendance of nearly 17,000 industry leaders, scientists, and government ofÞcials, representing 61 countries, 49 states (plus the District of Columbia and Puerto Rico), and 10 Canadian provinces. Today, San Francisco’s heavy biotech commercialization activity is its principal competitive advantage. Indeed, the region leads all hubs in terms of total established biotech companies having 100 or more employees, total biotech startups since 1990, and total amount of biotech venture capital invested over the past decade. Notably, in March 2004, Amgen announced its $1.3 billion acquisition of Tularik (based in South San Francisco), which specializes in small-molecule gene regulation therapies. The move gives Amgen—the Þrst biotech company to enter the top 10 Big Pharma tier—a key operational base in the San Francisco biotech hub. While still lagging Boston’s heavy concentration of academic research centers, the San Francisco hub is currently looking to expand its already well-developed infrastructure. State funding allocated in 2002 is expected to establish four new California Institutes for Science and Innovation. One of these new institutes, Bioengineering, Biotechnology and Quantitative Biomedical Research (QB3), will be located in San Francisco’s Mission Bay and will involve three campuses: UCSF, UCB, and UC Santa Cruz. In a cooperative effort with private industry,

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the institute is distributing $100 million in state funding among these three campuses. State government has done its part to foster commercial activity in the San Francisco hub. One example of state-level support is legislative backing for life sciences research, through both funding and enabling regulations of progressive areas of study such as stem-cell research. State funding of new biotech start-ups has also been a critical lifeline for many of the region’s ßedgling companies—for example, CalPERS, the California state pension fund, dedicates $500 million to investment in biotechnology through the local venture capital community. The state is also known for its favorable tax code, which grants liberal tax credits for R&D, including 15% credits for internal efforts and 24% credits for efforts outsourced to local organizations such as universities. Historically, San Francisco’s biotech leaders have made more signiÞcant local investments in manufacturing compared with investments made in the Boston area. Genentech, for example, currently controls about 30% of the world’s licensed capacity for the production of biological drugs; most operations are based at its manufacturing plants in South San Francisco and in Vacaville, 60 miles away. Looking to the future, however, biotech Þrms are eagerly examining other states and overseas sites to build new manufacturing capacity. The high cost of land and development in the Bay area, coupled with budgetary shortfalls, has limited government’s ability to subsidize future investments. Of particular concern to many in the life sciences business community is the 2003 expiration of the Manufacturers’ Investment Tax Credit, which had provided companies with a 6% tax credit to offset the cost of installing new equipment. A two-year bill that was reviewed by the state legislature in 2004 discussed the merits of reinstituting this credit and of pursuing many of the initiatives suggested in the BIO recommendations. Research Triangle Perhaps more than any other leading biotech hub, North Carolina’s Research Triangle is a product of meticulous planning and active corporate recruitment. The hub itself is concentrated in Research Triangle Park (RTP), a private, nonproÞt foundation located in the center of the regional triangle formed by the cities of Durham (Duke University), Chapel Hill (University of North Carolina), and Raleigh (North Carolina State University). In 1981, the state established and funded the North Carolina Biotechnology Center (NCBC). The NCBC aggressively recruits companies from outside the state by promoting the area’s strengths through outreach and marketing campaigns, facilitating processes such as site selection and permitting, and enlisting state and local government support in recruitment efforts. More than $135 million in state funds has been invested in the Research Triangle, and this investment has in turn stimulated more than $2 billion in direct, out-of-state investment through venture capital Þnancing, manufacturing investments, and federal research grants. Unlike most other biotech hotbeds, North Carolina has successfully developed a core strength in manufacturing in addition to R&D. The state demonstrated

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a strong commitment to biomanufacturing when it invested $42 million of its tobacco-settlement fund to expand the industry and tagged an additional $108 million for potential future investments. Numerous tax codes also testify to the region’s balanced support of both research and manufacturing: • • • • •

Sales tax of 1% or less for R&D equipment and an exemption for manufacturing equipment. Low-cost Þnancing for renovation and facility upgrade projects. A 7% tax credit for qualiÞed purchases of machinery and equipment. A 5% tax credit for R&D expenditures made in North Carolina. A $500–1000 tax credit for each employee who receives training; $500–12,500 tax credit for each new job created.

North Carolina is capturing an increasing share of the global biotechnology market owing to its established infrastructure, a solidly executed strategy for growth, and the Research Triangle’s favorable cost of doing business. San Diego San Diego is considered by many to be a fast-emerging biotech power that may soon rival Boston and San Francisco’s geographic leadership. A distinguishing characteristic of the San Diego biotech hub is its density. More than 35,000 people work at the 400 companies located within San Diego County’s 4200 square miles. By way of comparison, PriceWaterhouseCoopers estimates that this density of life sciences companies per square mile is 38% greater than in the San Francisco Bay Area region. Research centers such as the University of San Diego, the Salk Institute, the Scripps Institute, and the La Jolla Institute for Molecular Medicine highlight a long list of reputable academic institutions in San Diego. The physical proximity of these institutions creates a highly collegial atmosphere and helps to facilitate discussion and exchange of ideas through industry gatherings, lobbying efforts, and coordinated public education. This cross-fertilization of ideas is driven by an attractive working environment for area employees, who can easily pursue multiple job opportunities without having to change their residence. Such an environment helps drive entrepreneurialism, innovation, and job creation. San Diego has been particularly successful in helping its researchers translate their Þndings to commercial applications. A program called UCSD CONNECT is one key mechanism that links entrepreneurs in life sciences and other hightechnology industries with the resources they need for success, including technology, Þnancing, management advice, networking, and third-party support services. CONNECT is widely regarded as one of the nation’s most successful regional programs for fostering commercialization of promising research. Local biotech companies that have received FDA approval for new commercial products include Biosite, Idec Pharmaceuticals (now Biogen Idec), Agouron (now PÞzer), and Ligand Pharmaceuticals.

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In a joint study conducted by the Milken Institute and Deloitte & Touche, researchers identiÞed San Diego as the top hub of biotech activity. The study challenged conventional notions of biotech leadership in Boston and San Francisco, measuring each region’s strength in Þve categories: R&D inputs, risk capital, human capital, biotech workforce, and current impact. Notably, this study excluded all pharmaceutical and medical device activities; had these activities been included in the measurements, Boston would have ranked Þrst. Regardless of its precise ranking, there is no doubt that San Diego continues to gain visibility as a hub for biomedical discovery. The city has become a magnet for global pharmaceutical companies, outdoing its neighbor San Francisco in attracting new pharmaceutical investment. Novartis recently established two new research facilities in San Diego: the Novartis Agricultural Discovery Institute and the Genomics Institute of the Novartis Research Foundation. Johnson & Johnson just acquired biotech start-up Egea Biosciences. Other notable companies that have a foothold in the San Diego hub include Merck and PÞzer (through its acquisition of Warner-Lambert/Agouron). Los Angeles The Los Angeles hub is best known as the birthplace of Amgen—currently the industry’s largest biotech Þrm, with nearly 13,000 employees worldwide and $9 billion in annual revenues as of June 2004. The company was established in 1980 in the suburban area of Thousand Oaks, situated near the major research centers of the University of California at Los Angeles, the University of California at Santa Barbara, and the California Institute of Technology (Caltech). Private research institutions—such as the Cedars Sinai Medical Center, the City of Hope National Medical Center, and Huntington Medical Research Institute—augment the large infrastructure of biomedical research activity in this region. When compared with other leading biotech hubs, however, the Los Angeles region falls well behind on several important fronts. Although Southern California has numerous manufacturers of pharmaceuticals and medical devices, it has a relatively low concentration of strong biotech companies. The physical landscape, furthermore, is vast, and companies are distributed widely, resulting in a low degree of interaction between Þrms. Companies that spin off from the major research universities do not tend to stay in the Los Angeles region. Caltech has one of the best success rates: out of 32 biotech spin-offs created in the past Þve years, 31% remain in the area. Even the regional stalwart, Amgen, has elected to relocate key divisions elsewhere. The company is moving its cancer research division headquarters to Seattle, while locating a key manufacturing facility in Puerto Rico and an R&D facility in Boston. New York/New Jersey The greater New York and New Jersey areas are the traditional centers of the U.S. pharmaceutical industry. Emerging in the shadow of these pharmaceutical players is an active biotech research community. More than 200 academic

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research organizations, medical centers, and laboratories are located in metropolitan New York; some of the most prominent are Memorial Sloan-Kettering Cancer Center, Columbia University, New York Weill Cornell Medical Center, New York-Presbyterian Hospital, New York University, Rockefeller University, and the Albert Einstein College of Medicine. In 2003, universities and research institutions in and around New York City received more than $1 billion in research and training awards from the NIH. An additional amount exceeding $200 million was awarded by the NIH to research institutions in New Jersey. Although New Jersey ranks only modestly in terms of NIH funding, it is distinguished by the presence of 19 of the 25 largest pharmaceutical and medical technology companies in the world, including Aventis Pharmaceuticals, Bristol-Myers Squibb, Johnson & Johnson, Merck & Co., and Roche Pharmaceuticals. In addition, approximately 120 biotech companies cluster along a 48-mile research corridor stretching from northern to central New Jersey. Considering the large population of the metropolitan New York and New Jersey regions, the large number of well-known academic institutions, and the signiÞcant employment opportunities in life sciences research, it comes as no surprise that the New York/New Jersey region is home to a large number of highly educated workers. In fact, the total number of life sciences doctors who earned their degrees in the New York hub and are working there exceeds even that of Boston. Another key strength of this hub is its close proximity to the New York City Þnancial community, which provides easy access to venture capital Þrms, banking institutions, and other forms of private investment. Despite this seemingly critical advantage, the region is stronger in research than in biotechnology commercialization. In the past few years, the New York hub has actually lost national share in terms of total number of biotech companies. In part, this trend reßects faster industry growth in other hubs rather than attrition of companies in the New York/New Jersey hub. However, the high cost of doing business and relative lack of physical space are dissuading an increasing number of companies from conducting business in New York City, despite the abundance of life sciences discoveries ßowing from its universities and research institutes. State legislators estimate that the city’s top medical institutions launch approximately 30 start-up biotechnology companies every year, but few of these companies maintain their operations in the city. New Jersey has physical space and offers a more affordable cost proposition than New York City but lacks its neighbor’s high concentration of academic basic research centers. To encourage recruitment of the biotech industry, the Biotechnology Council of New Jersey has partnered with state government to enact an innovative and aggressive set of tax incentives—however, the state must combine tax incentives with other measures. Early accounts suggest that the region is on its way. Examples of recent progress include the establishment of a state-sponsored, $10 million Biotech/Life Sciences Venture Fund and aggressive pursuit of an academic center of excellence that produces biotech research.

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Philadelphia At BIO 2004, the life sciences industry’s national meeting, the greater Philadelphia region played a starring role. According to a report presented at the conference by the Pennsylvania Biotechnology Association, the state’s life sciences industry is a key driver of the regional economy, generating more than $5.5 billion in overall wages for its workforce in 2002. The Þve-county region that makes up greater Philadelphia accounts for nearly half of the total biotech-related employment in the state. Although Pennsylvania is not a particularly large recipient of venture capital investment, averaging just 2.5% of the total nationwide investment from 2001 through 2003, venture activity increased signiÞcantly in the Þrst quarter of 2004. Pennsylvania ofÞcials credit that growth partly to businesses and research generated by the state’s academic institutions, speciÞcally the University of Pennsylvania in Philadelphia and the University of Pittsburgh. Pennsylvania research institutions attracted more than $1.3 billion in NIH funding in 2003, making the state fourth in the nation for such grants. In a 2004 study entitled “Laboratories of Innovation: State Bioscience Initiatives 2004,” conducted by the Battelle Memorial Institute for BIO, Pennsylvania was identiÞed as one of only 12 states to have large employment bases in two specialized bioscience subsectors—drugs and pharmaceuticals and medical devices and equipment. The previously mentioned Milken Institute/Deloitte & Touche study lauded Philadelphia for having some of the key building blocks of a successful hub; in that study, the region ranked among the highest in areas such as total life sciences employment, total NIH funding, total biotech employment, and biotech graduate students per 10,000 residents. Pennsylvania has taken several steps in recent years to grow the state’s life sciences industry, including allocation of funds from the state’s tobacco industry settlement fund to biotechnology “greenhouses” that help seed and nurture earlystage companies. These greenhouses are sources of seed capital, but they typically do not provide start-up companies with physical space as business incubators do. The most prominent of these publicly supported greenhouses is BioAdvance, a $33.8 million fund focused on supporting biotech in southeastern Pennsylvania. Such initiatives, while leaving Philadelphia far below the top hubs in total commercial activity, demonstrate positive movement in a biotech community that will likely continue to grow. Washington, DC The Washington, DC, biotech hub has several strong assets, particularly in the area of research. The hub draws heavily on basic research performed at the NIH, which is located in nearby Bethesda, Maryland. The Washington academic community is also the beneÞciary of signiÞcant NIH investment. According to the Brookings Institute, the Washington/Baltimore region ranked third in NIH research funding, just behind the Boston and New York hubs. By a wide margin, the single largest recipient of NIH funding in 2003 was Johns Hopkins University,

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which earned well over $500 million in research and training grants, fellowships, and other awards. In addition to its research tradition, the Washington hub’s location in the political center of the United States has made it home to a number of important industry organizations, including BIO, which, as mentioned previously, is the largest and most inßuential biotech group in the United States. Furthermore, companies based in this region have ready access to key agencies, such as FDA, NIH, and the Patent and Trademark OfÞce, and to a rich talent pool of regulatory specialists and patent and IP specialists. Numerous biotech companies are based in the Washington area, including MedImmune, Human Genome Sciences, and Celera. However, in number of early-stage start-up companies, the Washington hub falls short compared with other leading hubs. The relative weakness in commercialization of new Þrms is due to low amounts of venture capital in the area and a comparatively lackluster technology transfer. The Washington hub ranks last among the top nine hubs in terms of venture capital investment. In addition, the number of R&D alliances between biotech and big pharmaceutical Þrms in Washington is far lower than in other leading hubs such as Boston, San Francisco, and New York. Another challenge facing the Washington hub is lack of commercial space for biotech laboratory facilities. What little space is available comes at a high price. Indeed, out of 180 metropolitan areas across the United States, the Washington region ranked 49 in housing affordability, according to the National Association of Home Builders. In comparison, the Boston area came in toward the bottom, with a ranking of 150, while San Francisco came in last at 180. Seattle Although less concentrated in biomedical research activity than other leading hubs, Seattle boasts some of the nation’s most important research centers. In 2003, for example, the NIH reported that the University of Washington received more than $440 million in research and training grants, the second-highest total of any public or private institution. Since 1974, the university has been the number one U.S. public university in terms of federal life sciences support. Another cornerstone of the Seattle hub is the Fred Hutchinson Cancer Research Center, one of only 35 comprehensive cancer centers nationwide. The center received approximately $30 million in R&D contracts from the NIH in 2003, among the highest of any nonproÞt institution in the United States. The Hutchinson Center and the University of Washington are the key technology foundations of Seattle’s biotech industry. Nearly one-half of the more than 190 biotechnology and medical device Þrms in the state are founded on technologies developed at these institutions. Perhaps the greatest example of past success is Immunex, which was founded by researchers at the Hutchinson Center and became the largest biotechnology company in the PaciÞc Northwest before being acquired by Amgen for more than $16 billion in stock and net cash in July 2002.

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Seattle’s biotech industry has been a major beneÞciary of the local Microsoft community. In 1990, Icos was established with $33 million in private Þnancing. At the time, it was the most signiÞcant biotech investment ever made, and the largest shareholder was Microsoft cofounder Bill Gates. In addition, the Bill & Melinda Gates Foundation has, over time, donated $70 million in grants for human genome research at the University of Washington. Microsoft’s other cofounder, Paul Allen, helped launch the Allen Brain Atlas project. The $100 million project will create a comprehensive map of the brain at the cellular level, illustrating the functional anatomy of the brain through a collection of gene expression maps, brain circuits, and cell locations. Allen is also currently lobbying to build a major biotech incubator in the Seattle area through his venture capital fund, Vulcan Ventures. In state support for its biotech initiatives, however, Seattle lags other leading biotech hubs. Although signiÞcant federal research grants help to sustain local biotech activity, the region has no history of state support for research and development. The Technology-Alliance, a nonproÞt consortium of industry leaders brought together by Bill Gates, reports that Washington ranks 46th out of 50 states in terms of state-level biotech support. Seattle’s thriving research institutions are threatened with growing competition for NIH funds, just one of the building blocks needed to construct a premier biotech hub. An initiative known as Bio21 is now under consideration to minimize the threat of competition. It outlines a plan that would commit $250 million from the state over Þve years, possibly doubling that sum with matching dollars from outside sources. The plan calls for establishment of a central biotech coordinating body that can help facilitate technology transfer and match early-stage companies to sources of investment capital. In addition, the state’s top research institutions could vie for competitive, peer-reviewed state grants distributed by a neutral, biotech-savvy commission. The Bio21 proposal took a back seat to a research and development tax credit in 2004, although it was considered an important step in the Seattle hub’s growth and development. OUTLOOK FOR EMERGENCE OF FUTURE HUBS Despite widespread interest in developing new biotech hubs, high barriers to entry confront regions that seek to develop them. Established concentrations of academic researchers and research institutions change slowly. Development of biotechnology-based products often takes a decade or more, and a very small number of patented biotech innovations achieve commercial success. Owing to the historically low odds of success and the long stretch of time associated with developing and securing regulatory approval for commercial biotechnology products, regions seeking to develop a biotech industry will need to invest a signiÞcant amount of time and resources. The established biotech hubs already offer a critical mass of talent, resources, and investment opportunities that attract the most desirable industry players.

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Although many other states have invested heavily in biotech or have unveiled plans for signiÞcant future biotech support, few start-up companies could afford to wait for these areas to develop. The failure of regions outside the top nine to catch up in the biotech race is best explained by recent trends. Nationwide, heavy investment and state-level support over the past ten years have increased competition for research dollars. As a result, research spending dispersed during that time, and the top nine hubs garnered a decreased share of total NIH funding and biotech-related patent Þlings. However, during the same period, factors related to the commercialization of biotech technology—venture capital investment, R&D alliances, and new companies—became more concentrated within the top nine hubs. IMPLICATIONS FOR THE PHARMACEUTICAL INDUSTRY Innovation and productivity within the research-based pharmaceutical industry have slowed. Industry executives are under increasing pressure to address the innovation gap through means such as reorganizing internal research efforts, inlicensing a greater proportion of products in development, and looking to biotechnology to help reÞll empty pipelines. In short, all of these approaches involve creating leaner, increasingly mobile, and more entrepreneurial organizations. Perhaps the most aggressive reorganization has been accomplished by GlaxoSmithKline (GSK), which restructured R&D into seven Centers of Excellence in Drug Discovery (CEDDs). Much like a discovery biotech company, each center is an independent, accountable, and entrepreneurial research organization that is focused on a speciÞc therapeutic area. In December 2003, two years after the initial reorganization, GSK presented the results of its redesigned research efforts. At that time, the company claimed 147 products in clinical trials—including 82 new chemical entities, a 46% increase in NCEs in two years—and 12 candidates in Phase III clinical trials, a 75% increase over the two-year period. While not all analysts were completely convinced that GSK had jump-started its pipeline, the company promised a record number of investigational new drug (IND) Þlings between 2004 and 2008, with a heavy focus on products culled from genomics and biotechnology efforts. The GSK model is but one example of how companies are beginning to integrate their divisional units into mini-businesses, each having a narrow focus and greater accountability. When splitting speciÞc business units from the broader corporate structure, it makes sense to position them in a fertile biotech environment. Proximity to the leading biotech hubs could facilitate the transfer of knowledge and technology and the recruitment of local talent and accelerate R&D output. Regions with active medical communities and research hospitals could even allow a pharmaceutical company unit to actively manage the closing phases of a drug’s life cycle. Provided that pharmaceutical units are given sufÞcient autonomy, each would be free to inlicense promising compounds, thereby tapping the regional hub’s

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abundant innovation in outside labs and start-up companies. At the same time, relative independence would allow pharmaceutical units to outlicense compounds that do not Þt the company’s needs, thereby providing focus and revenue to the entrepreneurial organization and improving morale. As the largest pharmaceutical companies continue to evolve, the importance of networking with leading biotech hubs continues to grow. The combination of strong academic research institutions, rich pools of scientiÞc talent, and ongoing cross-fertilization of ideas and energy will help drive productivity and innovation. Industry players that relocate key operations to the top biotech hubs in an entrepreneurial manner will be best positioned to share in this opportunity for future growth. REFERENCE DeVol R, et al. America’s Biotech and Life Science Clusters: San Diego’s Position and Economic Contributions. Milken Institute, June 2004.

Pharmaceutical Marketplace in Central and Eastern Europe

BUSINESS IMPLICATIONS •

•

•

Medium-term growth prospects are good for pharmaceutical markets in Central and Eastern Europe (CEE). Stable market economies characterize the region, and central to this stability is that more and more countries are becoming members of the European Union (EU). EU per capita gross domestic product (GDP) and pharmaceutical price convergence will gradually come, though at different rates across the CEE states. The region’s epidemiological profile suggests that strong potential exists for products to treat chronic conditions. However, although the intellectual property environment is increasingly stable, issues in this area remain—for example, a lack of bioequivalence information in the generics market and the longer-term issue of possible parallel export of products among EU markets. Reimbursement listing also will be a battleground because of government cost-cutting pressures. Further, although harmonization with European Commission (EC) regulations is progressing smoothly in general, the cost of required dossiers will shut down some local firms. Three types of company are active in CEE. Many traditional local firms face the daunting costs of meeting good manufacturing practice (GMP) standards as well as other costs of competing in an international marketplace. A small number of regional manufacturers will continue to succeed in both local and export markets. Western multinationals will increasingly dominate the market.

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright  2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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Overall, we expect that some issues related to transparency in reimbursement decision-making will create controversy and that health sector reform policies will contribute some uncertainty about regulatory systems. However, fairly stable politics and progressively stable market forces will make the region an attractive area of opportunity for the pharmaceutical industry.

CENTRAL AND EASTERN EUROPE’S CHANGING MARKET ENVIRONMENT Central and Eastern Europe (CEE), defined here as the ten countries that have just entered (or are about to enter) the European Union (EU)—the Czech Republic, Estonia, Hungary, Latvia, Lithuania, Poland, Slovakia, Slovenia, Bulgaria (in 2007), and Romania (in 2007)—encompasses a substantial pharmaceutical market with more than 100 million consumers.1 In addition, health spending as a percentage of gross domestic product (GDP) is growing in CEE. Major pharmaceutical companies are recognizing this market potential, and thus their activity in this region has increased in terms of joint ventures, mergers and acquisitions, and formation of subsidiaries. In this environment, however, companies must keep in mind that countries preparing for entry into the EU need to incorporate all European Commission (EC) legislation into their national law—including legislation pertaining to the pharmaceutical industry. As a result, they are in a state of flux as they reform legislation while promoting company merger and acquisition activity at a speed that exceeds the pace currently seen in Western Europe. Aspirant countries have already made great efforts to change their legislation, reflecting that joining the EU is the main focus of most CEE countries’ foreign policy. Poland was the first to do so, incorporating all of the pharmaceutical law established by the EC into its statutes as of October 2002. Also, Slovenia and Bulgaria both implemented intellectual property legislation during 2002. Intellectual property law and manufacturers’ obligation to comply with good manufacturing practice (GMP) rules will be the legislative changes that most affect pharmaceutical companies. In this article, we highlight the characteristics of the CEE countries, describe the pharmaceutical industry in this region, and profile four major players within the CEE region. We also expand on the pharmaceutical industry issues that countries are facing as a result of their efforts to become part of the EU. Finally, we provide an outlook for pharmaceutical markets in this region. CHARACTERIZING THE REGION Several key common features are notable across CEE: • • 1

A general embrace of capitalism has occurred. Political environments are stable and broadly democratic.

Plus Malta and Cyprus from outside the region.

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Prospective EU entry is a central tenet of economic, domestic, and foreign policies. Economic growth is currently rapid, but it is mainly rising from a low base. The public sector dominates the region in terms of providing coverage for growing health care expenditures, and its policies are subject to sporadic reforms.

Despite these commonalities, important differences exist between the CEE countries that will affect their pharmaceutical markets. For example, at one extreme, Slovenia already has average income levels and many health care institutions similar to those in EU nations, such as Portugal and Greece. Bulgaria and Romania, however, are far less developed in these areas. Also, although the CEE region’s epidemiological profile is generally similar to that of the EU, various countries exhibit dramatic differences. For instance, in Belarus, male life expectancy has fallen by 13 years since the end of Communist rule, while, in the Czech Republic, it has risen by 4 years. Demographically, low birth rates and emigration have skewed the population age in the CEE upward. According to the United Nations, the Czech Republic will soon have the world’s oldest population, with 41% of people older than age 60. This scenario suggests that demand is growing for products that target age-related chronic disease (especially osteoporosis, diabetes, and cardiac disease). Consequently, across the CEE pharmaceutical markets, double-digit percentage growth in pharmaceutical spending was the norm in 2001, with the largest market, Poland, growing approximately 17% in that year. Although the overall growth rate declined in 2002—in Poland, pharmaceutical market growth slowed to 8.5%—we believe growth is likely to continue in the medium term. IMS Health expects Bulgaria and Slovakia to have the greatest annual growth, but Poland is experiencing the most growth in absolute terms, surpassing $5 billion in pharmaceutical sales in 2005.

CENTRAL AND EASTERN EUROPE PHARMACEUTICAL INDUSTRY PROFILE Three types of pharmaceutical manufacturers operate in the CEE region: (1) traditional local generics (and branded copy) manufacturers, (2) Western multinationals, and (3) regional companies, a hybrid of the two aforementioned types. The traditional local companies are not likely to survive in an environment in which multinational and regional firms work to acquire them. Furthermore, these local companies will be required to comply with EU GMP standards—an undertaking whose high costs could put them out of business. The Western multinationals are gradually winning market share and will likely come to dominate the region. However, some regional firms may hold their ground as they are beginning to produce innovative products. All such regional firms, however, may not

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retain their independence because they will likely rely on Western multinational firms for capital and funding needed to take a drug candidate from early clinical trials to market. Traditional Local Firms Although rapidly decreasing in number, many local manufacturers still produce generics at a low cost. These manufacturers have the advantage of established distribution channels and, often, political support when they employ large numbers of local workers and/or are state owned. Nonetheless, these companies face some serious problems. First, they rely on traditional generics/branded copy products and therefore have little or no experience in R&D. Second, in many cases, production is ill-matched to demand, with marketing considered a novel concept after decades of command-and-control economics. Third, products may be removed from local reimbursement lists with little prospect of compensating new products or exports. Thus, the financial outlook for these local companies is bleak. Their products command low (often politically controlled) prices and declining market shares; at the same time, demands are rising for fresh capital to update outdated facilities, meet GMP compliance, and compete with professionally marketed rival products. Sometimes, even buying active ingredients for generics/copy products can be a challenge now that market prices are charged rather than Council for Mutual Economic Assistance (Comecon)2 administrative prices. For some local companies, such as Balkanpharma in Bulgaria, and some of the remaining large Polfa factories in Poland, there is good potential that foreign investment may rescue them. However, most traditional firms, including Poland’s approximately 300 family-owned pharmaceutical companies, are doomed in the changing market. Western Multinational Firms Several major North American, European, and Japanese pharmaceutical manufacturers are active in the CEE region. With a fluid environment that includes several national markets, various approaches and degrees of investment commitment are present. In two of the region’s larger markets, Poland and the Czech Republic, most international firms have set up subsidiaries. Incentives for establishing their own subsidiaries include receiving tax concessions; registration and reimbursement advantages; and gaining access to distribution, manufacturing facilities, and even research skills. Such incentives allow companies to gain maximum control and fit with other international operations. Companies that have established subsidiaries in the CEE region include Bristol-Myers Squibb (BMS) and Merck & Co. in the Czech Republic in 1991 and 1992, respectively. 2 Comecon is an intergovernmental body established in Moscow in 1949 to assist and coordinate the economic development of its members.

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In other cases, a major pharmaceutical company purchases a local firm to save time and/or takes advantage of specific existing resources (e.g., distribution channels, scientific expertise). In the past, occasional acquisitions in the CEE region were add-ons to deals with another focus. For example, in 1987, Abbott Laboratories bought BASF’s pharmaceutical business that included a subsidiary in Prague. However, more recently, acquisitions in the CEE region have been the focus of deals. This trend is increasing, with many deals taking place as part of CEE countries’ privatization programs. Examples of such acquisitions include the following: •

•

•

GSK (at the time Glaxo) bought the Polfa-Poznan production facility in Poland in 1997, paying $220 million for 80% of the stock and agreeing to invest a further $100 million over the following five years. (Employees of Polfa-Poznan received 15% of the stock, and the government retained 5% as an investment.) Ivax bought the Hungarian Institute of Drug Research (HIDR) in 1999. Ivax was attracted to HIDR’s research capability: established in 1950, HIDR had synthesized more than 5000 new molecules and collected over 1500 microorganisms to be able to produce medicinal substances via fermentation. Teva, the largest generics producer worldwide, bought two Hungarian companies: generics maker Biogal in 1995 and vaccine maker Human in 2000. Indeed, in 1999, Teva transferred the entire production of its U.K. generics subsidiary APS/Berk to Biogal, reflecting that it had met with success after acquiring Biogal.

Pharmaceutical companies are not the only entities to see the potential value in some of these local companies. In 1998, the U.S. venture capital group, Warburg Pincus, bought the Czech manufacturer Leciva for $110 million. Once a company has a subsidiary or has acquired a company, it can use it as a vehicle for further purchases in the same region. For instance, Sanofi-Synth´elabo bought Chinoin in Hungary and then Pharmavit from BMS at undisclosed price. The takeover of Pharmavit was announced in March 2002, and approval from Hungary’s Competition Office occurred in June 2002. Pharmavit will be used partly to export Sanofi’s products into Russia, according to Chinoin general manager Patrick Chocat. Emerging Regional Powerhouses Between the established multinationals and the traditional local firms is a select group of CEE manufacturers that have expanded to become regional, and even international, success stories. They have diversified sales well beyond their home markets, won access to capital through initial public offerings (IPOs), raised quality to GMP standards, and, although most are generics-based, they have also produced some successful innovative medicines. Four such companies have proved successful: Pliva (Croatia), Gedeon Richter (Hungary), Krka (Slovenia),

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and Lek (Slovenia). Table 1 shows their sales, exports, and profits in 2002. Several other firms, such as Leciva in the Czech Republic, are aspiring to reach this status. Many industry observers have been surprised by how these firms have thrived; they had expected these companies to quickly surrender their independence or even close once exposed to international competition. Instead, all four are now stronger than at any time in their post-1945 history. Their strategy is based on the recognition that domestic markets are too small and uncertain to rely on for substantial profit. For example, Lek and Krka have held a significant portion of market share in Slovenia—with 16% and 17%, respectively, in 2001—but they still achieved less than $50 million in sales each in that year. Their response was first to make efforts to obtain capital from shareholders. (In 2001, Lek and Krka were ranked as the top two companies on the Slovenian stock exchange for transparency and investor relations.) Then, they planned to gamble the capital gained in this way on major investments in quality improvement and export marketing, both westward into the United States and EU as well as eastward. Despite their growth, the four regional companies still have insufficient capital to carry new products beyond Phase I trials and are therefore highly dependent on licensing partners to bring their molecules to market. In addition, apart from Pliva, these companies still have much to do in terms of developing a strategy for direct international sales. Consequently, the question arises of whether they will be able to sustain their independence. In fact, Novartis acquired Lek in October 2002. The appeal of the other three companies as acquisition targets is growing for other multinationals because of their CEE distribution channels together with their production expertise. But, in the medium term, these companies are likely to remain successful as separate entities for reasons discussed in the following sections. Pliva. Pliva is the largest pharmaceutical company in the CEE region. Much of its recent success comes from its development of azithromycin, now one of the most widely prescribed antibiotics worldwide. Pliva has licensed azithromycin to Pfizer for all markets, except for Eastern Europe and some Asian countries. During late 2002, azithromycin received a boost after studies showed it can be used to treat cystic fibrosis. The new indication could help Pfizer extend the product’s patent expiry beyond the current 2006. TABLE 1. Pliva’s Corporate Acquisitions Company Polfa-Krakow Lachema Dominion Pharma AWD Pharma 2K Pharmaceuticals Sidmark

Country

Year

Poland Czech Republic United Kingdom Germany Denmark United States

1997 1999 2000 2001 2002 2002

Price (millions of U.S. dollars) 86.2 26.5 5 50 4 212

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The proceeds from azithromycin have allowed Pliva to act more unilaterally than any other CEE firm, expand more globally, and carry out a remarkable buying spree in the process. According to Pliva’s CEO, Zelijko Covic, in a recent interview, the company has reached its critical mass, and future growth is more likely to be organic, rather than through further acquisition. Mr. Covic also points out that Pliva’s ability to rely on azithromycin is declining, with the product accounting for 37% of sales in the first half of 2002, down from 44% in the same period in 2001. In addition, some key patents for the product expired in 2005, likely further contributing to declining sales. Other products in Pliva’s pipeline include a potential treatment for irritable bowel disease (Phase II); a thrombopoietin-like oncological therapy (Phase I); and a new antifungal (Phase II), inlicensed from Bayer. The company is also planning to launch generic albuterol sulfate in the United States shortly via its Sidmark subsidiary, following the FDA approval in October 2002. Most analysts are optimistic about Pliva—for example, Lazard Capital Markets expects Pliva to generate $1 billion in 2010 from three new proprietary products. An important part of Pliva’s R&D effort is a joint research project with GSK, in operation since 1999. The project is focused on developing a new macrolide antibiotic that is designed to overcome microbial resistance and treat candida yeast infection. The two companies shared responsibility up to Phase I; beyond Phase I, GSK is responsible for clinical development, and Pliva is charged with developing the manufacturing process and production of active ingredients. Lek. In November 2002, Lek became the first of the four regional majors to be acquired, when a majority of Novartis’s shareholders accepted Lek’s offer of the deal. Novartis was eager for this acquisition because its generics business geographically complements Lek’s. Novartis’s offer for Lek was CHF 1.3 billion ($860 million)—2.4 times Lek’s 2001 profits. Novartis was also interested in acquiring Lek’s capability to manufacture a generic version of the antibiotic amoxicillin/clavulanate potassium (Augmentin), which went off patent in the United States in December 2002. Other leading Lek products are ketoprofen (Ketonal; an analgesic) and amlodypine besylate (Amolpen; a calcium-channel blocker). Lek has developed generic omeprazole for the treatment of gastroesophageal reflux, peptic ulcer, duodenal ulcer, and esophagitis, and it is seeking FDA approval for the agent. Lek has already obtained FDA approval for its innovative anti-Parkinson’s product, bromocryptine (Bromergon). In the first half of 2002, Lek’s U.S. sales were up 200% over the previous year, with products sold through its subsidiary (also called Lek) in Wilmington, North Carolina. Novartis’s extensive distribution network in the EU will be particularly helpful for Lek’s products. In addition to its aforementioned investments in the United States, Lek has invested within the CEE region. A $12 million GMP-compliant antibiotics plant in Romania was completed in 2003. This plant expands the capabilities of local firm PharmaTech, which Lek bought in 2001. In 2001, Lek bought Argon (Poland)

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to gain access to the large Polish market. Lek also has subsidiaries in Austria, Bulgaria, Croatia, the Czech Republic, Macedonia, Nigeria, Pakistan, Russia, and Slovakia. Krka. Krka has focused on the CEE region, exporting approximately 80% of its generics products. Krka was established in 1954 in the midst of the Communist regime. According to Lazard Capital Markets analyst Martin Glen, Krka’s approach to the pharmaceutical business is less Western than that of the other three major companies in the CEE region; therefore, it is not as well known. In 2001, Krka invested a total of $51 million in its expansion efforts, completing plants in Poland and Slovenia as well as developing production and distribution facilities in Croatia and a factory in Russia (both completed in 2003). In 2002, the company opened another GMP-compliant plant in its hometown of Novo Mesto, Slovenia, that is able to produce 2.5 billion tablets per year. Krka subsidiaries now operate in Croatia, the Czech Republic, Germany, Hungary, Ireland, Italy, Macedonia, Poland, Russia, and the Ukraine. Krka’s strong position in the latter two markets would make it a particularly tempting target for Western multinationals. However, as Frances Cloud of investment bank Nomura International points out, Krka is protected against takeover by its shareholder structure, which provides its management (currently antimerger) with veto power. Leading Krka generics products are doxazosin (Kamiren), for high blood pressure and enlarged prostate treatment, and carvedilol (Coryol), for high blood pressure and congestive heart failure. With both agents, Krka was the first manufacturer to launch a generic version in the CEE region. Krka has also filed for registration of a generic version of carvedilol in the EU, where, in 2001, the company was granted marketing authorization for a generic version of another big seller, ciprofloxacin, to treat respiratory infections and anthrax. Gedeon Richter. The most well known of the four main CEE region firms is Gedeon Richter, founded in 1901 in Budapest. Like the other three companies, this firm recognizes that its domestic market will not be enough to sustain it—especially given current uncertainty in the industry–government relations in Hungary, as discussed in the later “Price Levels” section. Gedeon Richter’s strategy has been to increasingly focus on a limited number of market niches, in contrast to the broad-based product approach of the three competitors just covered. In particular, Gedeon Richter claims to be Europe’s number three player in the gynecological market, behind Organon (The Netherlands) and Schering (Germany). The company has proved its innovative credentials by developing a levonorgestrel-only emergency contraceptive, sold in the EU under license by Schering. Schering dropped its own contraceptive in favor of Gedeon Richter’s. Gedeon Richter’s other main products are generic enalapril, lisinopril, and fluoxetine in Western markets and the central nervous system (CNS) drug vinpocetine (Cavinton), which sells well in Russia. As a result of Gedeon Richter’s relatively even geographical spread in terms of pharmaceutical sales, in the first six months of 2002, only approximately

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$27 million of its exports went to CEE markets, $35 million to Russia, and $50 million to North American and EU countries. In October 2002, the Hungarian company won a Polish government tender for its Polfa-Grodzisk facility. Gedeon Richter already has manufacturing facilities in Romania and Russia. In the United States, Gedeon Richter has chosen to use a partnership strategy. It has a wide-ranging strategic agreement with Barr Laboratories covering active ingredient supply (Gedeon Richter supplies long-term steroid ingredients), product development, and marketing. Gedeon Richter also supplies long-term steroid ingredients to Johnson & Johnson. Gedeon Richter also is fairly well protected against takeover because the Hungarian government, whose privatization agency owns 25% of the company, is eager to keep the firm national. If the government’s position were to change, there is little doubt that foreign buyers (such as Teva) would be interested. DOSSIERS, PRODUCTS DEVELOPMENT, AND PATENT PROTECTION IN THE CENTRAL AND EASTERN EUROPE MARKETS Dossiers To meet EU rules, companies operating in the CEE region must update the legal dossiers of every product that they market locally. How many products this requirement will affect across the region is unknown, but, in Poland, for example, as many as 10,000 product dossiers will need updating, with almost half of these manufactured by domestic companies. If the dossiers are not up-to-date when a country becomes an EU member, products must be taken off the market (although some countries have won extensions). The significant expense of updating these dossiers is a problem for many companies; for traditional local companies that often operate on extremely small profit margins, it is not economically viable. New clinical trials must be conducted for each product, and, in many cases, the cost is greater than the potential return from a product’s continued domestic sales. Product Development Since the early 1990s, the range of products available in the CEE region has grown steadily. For example, in Hungary during 2001, there was a ratio of five new products to each withdrawal. The percentage of the market that imported products and over-the-counter (OTC) products account for is also growing, although generics remain prevalent. The continued importance of generics within the CEE markets varies. In Poland, low-priced generics account for approximately 65% of pharmaceutical sales; in contrast, in Slovenia’s relatively generous reimbursement system, generics account for less than 5% of sales. To date, there is little brand awareness among most CEE consumers. However, industry experts believe the situation is better than in Russia, where, according to a recent study from the Sechenov Medical Academy in Moscow, up to 40% of doctors, 25% of pharmacists, and 80% of patients are not able to distinguish between generics and proprietary drugs.

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Patent Protection Western levels of patent protection were entirely lacking in CEE when the Berlin Wall came down in 1990; hence, many CEE pharmaceutical manufacturers produced copies of branded drugs. Now, every CEE country has Trade-Related Aspects of Intellectual Property (TRIPS)-compliant legislation in place or pending as part of the region’s push to be accepted by multilateral organizations such as the EU, the Organization for Economic Cooperation and Development (OECD), and the World Trade Organization (WTO). For example, in Slovenia, home to Lek and Krka, all pharmaceutical copying was allowed until 1993; from 1993 to 2002, finished product patents were enforced but not pipeline protection. Now, data exclusivity rules are in place. This scenario is common, though in some countries rules may not be fully active until they become EU members. Note, however, that because patents are so recent in CEE, it will be several years before the copies of older products disappear from the market. Bulgaria’s legislature introduced a Roche-Bolar provision in 2002; such provisions allow generics companies to conduct development work on products still under patent. Hungary already has such a provision, Poland passed one in 2001, and Slovenia has a less well-defined form of one. These countries are thus in line with U.S. practice and contrary to the EU norm. The main reason that these CEE countries are not striving to follow EU rules for patent protection of generics is that Roche-Bolar provisions are an excellent way to assist indigenous generics manufacturers and attract investment from foreign ones (such as Novartis and Teva). Although these countries will need to conform to the EU norms when they are admitted to the EU, they can benefit from this practice in the meantime. PRICING AND REIMBURSEMENT IN THE CENTRAL AND EASTERN EUROPE MARKETS CEE governments are in a similar position to EU countries in terms of costcontainment issues: they face strong pressure to limit the spending of publicly funded health care systems, especially on drugs. Thus, their main objective is pharmaceutical cost control, with a constraint not to drive away foreign investment or to jeopardize EU accession. This position is becoming increasingly difficult because, despite government efforts, the amount spent on medicines is growing as a proportion of total health care spending. In Slovenia, for example, drugs accounted for 26% of the health budget in 2001, compared with just 12% in 1998. The range of measures to reduce drug spending is wide and similar to that in Western Europe: • •

•

Control of ex-manufacturer prices. Clawbacks if sales exceed limits. The term clawbacks denotes the government setting a value limit for a drug (or for all the drugs of a single company) that its public reimbursement system will cover. Values beyond this limit will be retained by the government in part or in full. Control of wholesaler margins.

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Control of prices for hospitals. Reimbursement decisions (e.g., on formulary inclusion). Reference pricing. Generics substitution. Patient copayments. Nonsupply of reimbursable products to pharmacies. (If a pharmacy does not have a product, no patients can purchase it and pass the cost to the government through reimbursement.)

Also, within the CEE countries themselves, there are still problems in clearly establishing generics whose properties are sufficiently similar to the original products. In many countries, little information exists on bioequivalence, and there is sometimes uncertainty regarding how to ensure that a generic is a quality product. Moreover, while less prevalent than in the ex-Soviet nations, counterfeiting of drugs (sometimes without active ingredients) is sometimes a problem. Also, research-based manufacturers in the EU are very concerned that CEE countries’ entry into the EU will herald a flood of cheap parallel imports, undercutting higher prices of certain drugs in EU markets. Several EU governments are pressing for at least a temporary ban on parallel export from new EU entrants. Price Levels Constant conflict surrounds pharmaceutical price levels, a key variable for governments as they reimburse so much medicine use. An example of such conflict in the CEE markets is the situation in Hungary. In 2001, the pharmaceutical industry association, the Association of Innovative Pharmaceutical Manufacturers (AIPM), and the government at that time signed a three-year price agreement. Under the deal, drug price increases for local firms were limited to 70% of the expected inflation rate. In 2001, the first year of the agreement, prices rose by 3.2%; in 2002, the increase was 3.0%. These increases essentially equate to no change; thus, the agreement has been upheld—to the dismay of pharmaceutical companies. However, state reimbursement spending is easily set to surpass its optimistic $578 million 2002 budget, as in many other countries in the CEE region. A new Socialist-led government stepped into this situation, with Judit Cseh´ak as health minister. Ms. Cseh´ak has implied that the pharmaceutical industry must choose between complying with the agreed price increases and having new products accepted for reimbursement, vital for most companies. Thus, a standoff has occurred. Given the balance of power, Ms. Cseh´ak should be able to dictate new agreement terms. However, she will be somewhat restrained by pressure from foreign investors/embassies and Gedeon Richter, the domestic pharmaceutical powerhouse. Despite these difficulties, Hungarian firms should be glad they are not in Slovakia, where local manufacturers have not been permitted a price rise in four years, despite 45% inflation during that time period.

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CEE government controls over drug prices vary. In the Czech Republic, for example, the government sets maximum prices even for nonreimbursed and OTC medicines (though we expect OTC drugs to be decentralized in late 2002). Poland, on the other hand, does not regulate nonreimbursed medicines. Also, governments sometimes treat local and foreign producers differently, both in terms of allowed ex-manufacturers’ prices and allowed distribution margins, but the trend is against this different treatment. Poland, for instance, eliminated such differentiation in 2001 and switched to a unified system. Some CEE countries explicitly use prices in countries with similar per capita GDP to determine their own prices. Poland has recently begun to do so. Slovenia sets innovative drug prices at 85% of the average price in France, Germany, and Italy—countries with a similar per capita GDP. Reimbursement Lists Even as the number of market-authorized products expands, in the interests of cost-cutting, CEE governments have been cutting the number of drugs on their health system reimbursement lists. Slovakia, for instance, cut 40 drugs from a 1000-product list in 2002, while Poland cut 380 drugs, though it introduced the first new additions in three years. As in the EU, approaches vary across the CEE region to gauging the economic benefit of new drugs and deciding whether they should be included on reimbursement lists. However, in 2002 Estonia, Latvia, and Lithuania all agreed to a common approach to cost–benefit assessment. Complaints are widespread among manufacturers (especially foreign ones) about levels of transparency in reimbursement list decision making. According to critics, inclusion criteria are often hard to define and administrative procedures are overly bureaucratic and fail to reward innovation. Foreign companies also sometimes allege that preference is shown toward local competitors. Pavol Mazan, executive director of the Czech chapter of the International Association of Pharmaceutical Companies (MAFS), went on record in 2002, asking for a new appeals body for reimbursement decisions because of the lack of transparency. The situation may be improving under EU influence. For example, to meet the EC Transparency Directive, in 2001, Poland passed regulations governing maximum time for setting prices (180 days), making reimbursement decisions (90 days), and updating the reimbursement list (annually). However, Wojciech Gryta, external affairs manager for Pharmacia, points to the continued lack of an adequate appeals procedure. Copayments All CEE countries apply patient copayments to at least part of their reimbursement lists. Hungary, for instance, reimburses 90% or 100% of the price of certain products for chronic and rare diseases. In 2001, 6 new chemical entities (NCEs) were added to those reimbursed at these rates, whereas 12 NCEs were included at lower 50% and 70% reimbursement rates. A total of 1161 generics were awarded fixed-fee reimbursement (i.e., a set amount of coverage was given for each drug).

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The higher subsidies may apply to either products that treat certain diseases or certain demographic groups, or both. In the Czech Republic, products for obesity, Alzheimer’s disease, and arthritis are all more reimbursed at the highest rate. In Poland, war veterans receive their drugs free (accounting for just 0.3% of the population, they consume 10% of the reimbursement budget). CEE patients are now well used to cost-sharing, although this practice was rare in Communist times. Apart from the official copayments, patients may also make unofficial “gifts” to physicians in some areas, on either a voluntary or nonvoluntary basis, partly in response to the very low level of physician salaries. The copayment structure also sometimes encourages fraud—some Polish war veterans may be obtaining free drugs for younger family members, or to sell. DISTRIBUTION The pharmaceutical wholesale and retail markets are two of the areas that have most radically changed since the Communist era. Private investment in setting up pharmacies has been substantial, though most has been in wealthier urban areas, with rural areas often neglected or left to a lower quality public system. Pharmacy chains are springing up where they are permitted, and pharmaceutical imports have often attracted more investment than have drugs manufactured domestically, especially at times of economic instability. Although CEE distribution markets are growing rapidly, foreign entrants have so far had mixed results in marketing their drugs. Pharmacies may gain in importance in the retail drug market if cost-saving measures that allow pharmacists to substitute generic equivalents for prescribed products are introduced. Many CEE pharmacists already advise patients on medicines, even prescription products. However, at the same time, they may lose some market share to other retail outlets—Poland recently legalized OTC sales outside pharmacies, for example. OUTLOOK Markets and Prices CEE pharmaceutical market performance will depend greatly on the local economies, as a small GDP change magnifies into a large change in the size of the pharmaceutical market. These GDPs, in turn, are influenced by the world economy, making precise forecasts difficult. But the underlying demand for medicines in the CEE region is strong—aging populations and high incidence of several chronic illnesses combine with societal consensus that an increased proportion of national resources should go to health care. However, the main driver of growth in the CEE region will likely be increases in average prices. These will gradually converge with those of the EU, even if the price growth rate will begin to slow because of public spending limits. One of the mechanisms promoting this growth is the growing trend among CEE governments, like Poland and Slovenia, to take into account EU prices when

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setting their own price levels. Conversely, CEE-produced generics will help put downward pressure on prices within the EU, as will parallel trading of patented products. Parallel trade is likely to be most visible across the Polish–German and Austrian–Hungarian borders. Corporate Change Analyst attention is likely to focus on the prospects for the remaining three independent regional powerhouse companies discussed earlier—in particular, whether they will follow Lek in being bought by Western multinationals. Ironically, the strongest of the three, Pliva, is the most probable takeover target because Krka and Gedeon Richter both have shareholder structures not disposed to selling. All four of these companies are likely to continue their recent success. Each has the possibility to supply a steady stream of generic versions of blockbusters once they come off patent. Some impressive R&D niches are also developing: Pliva with anti-infectives; Richter with CNS and gynecology; Pliva and Lek, both with cutting edge biotech generics. Thus, there will continue to be a thriving independent local CEE pharmaceutical sector, with perhaps some smaller new entrants to the ranks of the regional powerhouses appearing, now that a successful path has been demonstrated. But, inevitably, there will also be substantial consolidation across the CEE pharmaceutical manufacturing sector as a whole. An increase in competition in the generics industry will create a need for secure distribution and economies of scale as minimum survival requisites. GMP requirements for old factories present another challenge for the industry. The strongest consolidating force may be the EU’s dossier requirement, however. Unable to meet this cost, many smaller companies, like Poland’s 300-odd family-owned pharmaceutical companies, will dissolve. Regulatory Issues Strong local supply of generics will be supported on the demand side by CEE governments’ pushing of generics as a cost-containment measure. Reimbursement lists will also be more regularly updated for the same cost-cutting reason, possibly supplemented by greater use of treatment protocols. As a result, manufacturers will need to pay close attention to government policies. Although the private health care sector will grow, state health services will continue to dominate the CEE region. Although the CEE countries will need to comply with the EC Transparency Directive, market participants should expect continued occasional problems regarding reimbursement decision making. CEE legislators need only consult their counterparts in several EU countries to know that EC regulations may curb, but certainly will not eliminate, rule bending to cut costs or favor local players. Also, in the midst of its recent reforms, Poland transferred market authorization power from a semi-independent regulator to its Ministry of Health, which also now judges any appeals against its own decisions.

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Although EU harmonization is progressing well, establishing common protocols across the region will be a regulatory challenge. This cohesion is needed both for dossiers (to minimize their cost) and for mutual recognition purposes (i.e., if one country grants approval, the other EU countries will accept that drug). Although there is no problem in the EU’s European Medical Evaluation Agency (EMEA) licenses being immediately valid in CEE, mutual recognition procedures will not work while summaries of product characteristics (SmPCs) differ. Political Issues In some respects, the CEE political environment is positive for the pharmaceutical industry—the strong pro-EU commitment, for example, and near-universal recognition that more should be spent on health care. However, there are some political variables that will hinder the industry. One is health sector reform policies. CEE governments are constantly being advised by the World Bank, the EC, and private consultants and academics about the merits of reforming the financing system for health care, health care provider incentives, and the balance between the public and private health sectors. The Czech Republic, for example, switched in 1993 from a health ministry tax-base controlled finance system to a system of competing insurance funds. Poland decentralized control to public regional insurers in 1999, as a precursor to privatization, but it has now decided to reverse back to a single insurer because of several factors, including a change in government. Hungarian government-employed family doctors are currently receiving loans to help them privatize their practices. These shifts will continue, suggesting that industry participants should keep a close eye on political debate in the CEE and even involve themselves in it. A leftward political swing is currently occurring across the region. This shift and any future shifts to the right are all likely to be moderate. Nonetheless, a backlash remains against foreign ownership, following the massive privatizations of the 1990s. Nevertheless, participants in the CEE health sector should feel secure that they are in a fairly politically stable environment in which market forces will increasingly build.

Valuation Models for European Biotech

SUMMARY The biotechnology industry in Europe and the United States has underperformed as an asset class. Despite the promise of their underlying technologies, most biotech companies have delivered little value to investors thus far. After the stock price correction that burst the genomics bubble, few investors are willing to risk investment in biotechnology when undervalued blue-chip investment opportunities are available. Consequently, biotech companies suffer from lack of the follow-on funding they need to propel promising agents through clinical development. In this article, we examine the commercial and environmental drivers that are impacting the biotechnology industry, focusing particularly on European biotech and the negative impact of inßated company valuations. We emphasize the need for more precise valuations that will enable more efÞcient selection of investment opportunities and improved asset allocation, thereby helping to restore investor conÞdence and improve the industry’s long-term prospects, and we examine several analytical tools that companies can use to perform this essential task. BUSINESS IMPLICATIONS •

The European biotech industry needs development capital in order to bridge the funding gap between having potential drug candidates in early-stage development and establishing revenues in the form of product sales or licensing deals with big pharmaceutical companies.

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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VALUATION MODELS FOR EUROPEAN BIOTECH

In order for European biotech companies to aspire to a fully integrated biopharmaceutical business model capable of challenging Big Pharma, disinterested investors must be convinced that these companies can deliver suitable returns on investments. In many past valuations, less than rigorous project assessments and an excess of capital ßattered projects that were based on weak business models and represented poor investment opportunities. When the initial funding ran out, the weakness of many underlying business models was exposed, and many ventures ceased to operate because they could not attract additional sources of capital. More-precise assessment of intrinsic project value, enabled by reÞned analytical tools—discounted cash ßow (DCF) analysis (and the related net present value (NPV) approach), risk-adjusted NPV analysis (rNPV), Monte Carlo analysis, and real options valuation (ROV)—will generate more efÞcient selection of investment opportunities and improved asset allocation, which is essential to restoration of investor conÞdence in the European biotech sector.

INTRODUCTION The biotechnology industry in Europe and the United States has underperformed as an asset class. Despite the promise of their underlying technologies, biotech companies have delivered little value to investors thus far. The stock price correction that followed the genomics bubble caused nonspecialist investors and their capital to desert the biotechnology sector, and few private or institutional investors are willing now to bear the risk of investment in biotechnology when undervalued blue-chip investment opportunities are available elsewhere. The euphoria of the genomics era and the associated high company valuations have largely been replaced by the pragmatism of the proteomics, or post-genomics, era. Cash ßow dominates and Big Pharma appears to be in a position to control the biotechnology funding gap that has resulted from a combination of extended product development cycles, the need to maintain expensive R&D programs, and high cash burn rates. Consequently, biotech companies—many of them in early product development stages—suffer from restricted access to the follow-on funding, both equity and debt, that they require in order to propel promising agents through clinical development. The funding gap is driving consolidation in pursuit of solvency. In 2003, biotechnology sectors in the United States and, to a lesser extent, in Europe (where companies are Þnancially weaker) experienced consolidation through mergers of biotech companies and acquisition by pharmaceutical companies—which is likely to be increasingly frequent. Although the end goals behind biotech/biotech and pharma/biotech mergers or acquisitions differ, the primary driver of these events is the same: company valuations have plummeted. The European biotechnology sector is less mature than its U.S. counterpart and is therefore experiencing a more dire impact from current Þnancial trends. The European sector must recognize that if it is to break out of its limited

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role as supplier for the product pipelines of Big Pharma, it must acknowledge the current crisis in investor conÞdence. Venture capitalists (craving investment liquidity and driven by the needs of their institutional investors) and Big Pharma (on the lookout for both bargains and potential competitors) are remodeling the industry according to their own agendas. At the same time, industry analysts are urging European biotech ventures to seek near-term revenues by offering service products to Big Pharma—effectively, to function as an outsourced R&D arm. It is apparent that the biotech business model is under external pressure to change. If European biotech is to achieve a fully integrated biopharmaceutical business model capable of challenging Big Pharma, disinterested investors must be convinced that biotech companies can supply appropriate returns on their investments. In this article, we examine the commercial and environmental drivers that may account for these phenomena, focusing particularly on the European biotech industry, and we address the impact of these drivers on the evolution of the biotech business model. Also, we emphasize the need for implementation of more accurate project assessment and valuation tools if long-term prospects are to improve for the investor and for the European biotech industry. EUROPEAN BIOTECHS—THE STATE OF THE INDUSTRY The European biotechnology industry had revenues of $8 billion in 2001—less than one-third of the $25 billion in revenues recorded in the United States in the same year, and relatively few European biotechs are traded publicly. Many have little or no access to the licensing or product revenues associated with latestage development or marketed agents, and even fewer represent the blue-chip (i.e., low-risk/high-value) investment opportunities that would interest investors during a depressed economic period. Nevertheless, the European biotech industry needs development capital in the form of cash in order to bridge the funding gap between having potential drug candidates in early-stage development and establishing revenues in the form of product sales or licensing deals from Big Pharma companies. During these difÞcult times, large, cash-rich pharmaceutical companies are among the few investors willing to risk biotechnology investment. Big Pharma’s motivation to invest in a biotech, however, is more opportunistic than altruistic. Recognizing that it must provide appropriate returns to its shareholders, Big Pharma will look to biotech to populate its own barren pipeline. In the current environment, however, few product strategists or licensing executives would consider expensive inlicensing deals. Instead, Big Pharma is acquiring undervalued biotechnology projects that have high potential by offering restrictive licensing terms, by direct equity investment, or by acquisition of cash-strapped biotech companies. A bonus for Big Pharma is that this action may suppress the development of potential competitors. Indeed, industry analysts have suggested that Big Pharma can afford to be patient, “dripping” small amounts of funding into interesting biotechs before deciding, at some later date, to acquire these companies or to terminate the relationships (Smith Ewing E, 2003).

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With share prices depressed relative to the actual worth of the underlying companies (the intrinsic value), biotech companies are looking to mergers with their peers for salvation. The larger publicly held U.S. biotech companies are responding to the predatory activity of Big Pharma by forging multibillion dollar “paper mergers”—for example, the $6.6 billion merger of Biogen Pharmaceuticals and IDEC Pharmaceuticals can be viewed as a defensive response to the current investment trends in the biotech industry. The increase in market capitalization postmerger should move the new venture beyond the price that Big Pharma will pay to acquire its pipelines and product portfolios under current market conditions. Like their U.S. counterparts, European biotech companies are also consolidating; British Biotech acquired Vernalis for $80 million in 2003. However, the drivers for many proposed European biotech consolidations may be more prosaic than those in the United States. Economies of scope and scale achieved through R&D synergies and infrastructure cost savings, together with access to the cash reserves of potential partners, are expected to prolong the viability of merged European ventures, at least until the next round of funding can be identiÞed or product revenues can be generated. For much of the biotech industry in Europe, the funding gap appears wider than ever and prospects for commercially viable mergers with other biotech companies seem limited, either because of poor pipeline potential or poor market sentiment. For example, Cambridge Antibody Technologies (CAT; 2002 sales of $14.8 million) proposed a paper merger with OGS; however, following a decline in CAT’s market capitalization, the OGS board accepted acquisition by Celltech—while acknowledging that the price Celltech paid was substantially less than the intrinsic value of OGS. The fate of CAT demonstrates the plight of many European biotech companies. Despite a promising product pipeline and a business model that was until recently considered a paradigm for the sector (establishing licensing revenues with medium and Big Pharma partners and structuring the deals to include marketing and manufacturing rights), CAT is still struggling against the poor outlook that analysts forecast for biotech. If promising European biotech companies are to evolve from their current business model, they must repair the damage done by the excessive company valuations and subsequent stock price corrections associated with the genomics bubble. Only by providing attractive long-term investment opportunities will the sector attract the investment capital that it needs to progress beyond its current situation: excessive company valuations followed by stock corrections and lack of funding. A substantial step forward will be a reversion to sound business fundamentals. Managers must value their projects accurately according to realistic, auditable equity and project valuations—not sentiment and peer group comparable analysis. Ultimately, the biotech sector must provide fair shareholder returns by appraising future investment opportunities rigorously and accepting only opportunities that can maximize wealth. To make this scenario a reality, the biotech sector must continue to develop and stringently apply project assessment and valuation tools.

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ACCURATE VALUATION IS A NECESSITY Biotech management teams and institutional and private investors face the same investment question: How can limited funds be allocated to maximize investment returns? In biotechnology, as in any commercial setting, there is value in understanding and translating project attributes of uncertainty and risk into investment decisions. Financial theory suggests that efÞcient investment is based on valuemaximizing criteria—that is, a consistent system that assesses projects by their value and funds them with the intention of maximizing shareholder wealth. The initial rush to fund genomics-based biotech ventures and the high demand relative to supply led to excessive company valuations and a surplus of development capital. Considering many of the valuations of the past, it is obvious that less than rigorous project assessments and an excess of capital ßattered projects that were based on weak business models and represented poor investment opportunities. Once the initial funding ran out, however, the weakness of many underlying business models was exposed, and many ventures ceased to operate because they could not access additional sources of capital. Companies that accepted unrealistic equity valuations during the genomics bubble must now strive to generate substantial product revenues in order to provide fair returns to their investors. In many cases, however, the returns required to justify high company valuations are unlikely to be sustained by future cash ßows. The superßuity of target opportunities relative to demand continues to drive the cycle of wealth destruction. Furthermore, many biotech licensing deals or acquisitions are being completed for less than their intrinsic value. Setting up a valuation process is difÞcult in the biotech sector because of the complex timing and uncertain nature of project cash ßows, variations in risk throughout the life of a project, lengthy development times, low project salvage values, and high capital requirements. The forecast, or expected value of an investment, depends on two factors: the current value of the cash ßows associated with a project and the probability that those cash ßows will materialize—the risk of the investment, which is reßected in the required investment return (cost of capital), a critical consideration in the valuation process. The cost of capital represents the return that shareholders demand to compensate them both for the time value of money (the return that a risk-free investment would accrue over the same investment period) and the additional risk that their investments bear. Additionally, in companies funded with both equity and debt, the contribution of both to the projects’ funding structure should be reßected in the cost of capital as the weighted average cost of capital (WACC). The cost of capital is an opportunity cost, which is dependent on the use of funds rather than their source. The cost of capital will change from one investment to another depending on risk proÞle—as project risk increases, the cost of capital will increase. When the investor receives a return that is greater than a project’s cost of capital, wealth is created; if an investment returns less than the cost of capital, wealth is destroyed.

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Managers and investors should consider the cost of capital a hurdle rate for project approval. Projects that are not expected to return their cost of capital should not be funded unless there are compelling reasons to do so. Consequently, if management teams are to maximize shareholder wealth, they should rigorously assess and quantify the value of all investment opportunities; future project cash ßows, associated risks, and the cost of investor capital must all be forecasted accurately. Failure to assess and quantify the value of competing investment opportunities generated the poor investment decisions and valuation artifacts that are responsible for the current plight of the biotech industry. It is important to note that both positive and negative valuations destroy shareholder wealth in the long term. Many methods are available to facilitate stringent project assessment and funding according to value-maximizing criteria, but the potential for rapid growth and the risk associated with biotech makes many of them unsuitable for this sector. Indeed, the difÞculty of valuing biotech investments promoted the use of the comparable and peer group valuation techniques that are responsible for the genomics bubble and current valuation inaccuracies. Quantifying the Intrinsic Value of Biotech Projects The nature of the product life cycle complicates valuation because there are two distinct stages: the development stage and the product stage. The development stage can last more than ten years, and during this period, no sales revenue is generated. The product stage begins when a therapeutic agent or technology reaches the market. When return on investment Þnally results, the length of the product life cycle (the revenue-generating phase) will affect its value. The product stage is short compared with development time, sometimes as short as a few years (despite long patent-based monopolies). The payoffs for biotech are the licensing fees and royalties from sales achieved by Big Pharma and a marketing partner—or sales revenue, if the biotech retains those rights and can deliver a drug or technology to market. For therapeutic agents, these revenues may be estimated by using sales and market forecasts for drugs currently used to treat the target indications. Biotech project assessment and valuation have three basic drivers: •

• •

Developmental Age. The risk proÞle of an investment typically improves as the project develops; at later stages, there is less risk for the investor because the cost of capital is lower and the project is worth more. Length of Time to Market. The shorter the time to market, the greater the liquidity and, consequently, the value of the project. Terminal Value. The greater a project’s expected worth at completion, the greater the amount that an investor will be willing to venture in that project.

When quantifying the intrinsic value of a project, biotech management must assess the impact, implicit or explicit, of all three drivers on the “fair value” of the opportunity. The most appropriate tools for biotech valuations may be limited

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to discounted cash ßow (DCF) analysis (and the related net present value (NPV) approach), risk-adjusted NPV analysis (rNPV), Monte Carlo analysis, and real options valuation (ROV). Valuation Tools Discounted Cash Flow Analysis. DCF analysis enables the conversion of forecasted (i.e., uncertain) future cash ßows into values that can be directly compared with current investment needs; DCF analysis requires revenue and growth projections as well as projections of potential market share. Forecasted cash ßows are discounted back to current value terms by using an appropriate cost of capital as a discount rate. Figure 1 shows a model of DCF analysis along the drug-development time line. A beneÞt of this approach is that the discount rate can be modiÞed to reßect the changing risk proÞle of the venture. One study (Nelson TR and Mukherji A, 1998) suggests a model in which the cost of capital is adjusted to reßect the risk proÞle of the project at each stage of development. In this model, a project in preclinical basic research is discounted at a rate of 75%. As the project passes through the consecutive stages of clinical development, risk is mitigated: during Phase I, the discount rate is 40%; in Phase II, the rate is 35%; by Phase III, cash ßow discount rates are 20–25%. A general rule in using the DCF approach is that projections should not extend beyond ten years: because high discount rates are used, the model is sensitive to time. The long product development times that are common to early-stage biotechnology projects may make the standard DCF approach inappropriate for many valuation targets. However, for a biotech company seeking to value the licensing terms offered by a Big Pharma or Big Biotech partner (contingent on near-term milestones being met), DCF analysis may be suitable. A further criticism of this model applies to the accuracy of the discount rate. Many practitioners of DCF use subjectively estimated discount rates to reßect project risk. However, perceptions of risk may differ, thereby generating valuation variability. Indeed, some industry commentators speculate that during the genomics boom, equity analysts applied inappropriately low discount rates to Year 1 Preclinical

Year 15 Phase I

Phase II

Phase III

Sales of approved drug

CF1

CF2

CF3

Revenues from drug sales (cash in-flows) i

Present value = Σ CFi /(1+r)i t=1

Where CF = cash flow, r = cost of capital.

FIGURE 1. Model of discounted cash flow analysis.

CF4

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projected cash ßows and inaccurate estimates of the length of the FDA approval process (Papadopoulos S, 1998). This widespread failure to account for the risks and uncertainties associated with biotechnology investments—and the inappropriate valuations that resulted—was a substantial factor in driving the subsequent stock price correction. Financial theory suggests that cash ßows forecasted using DCF analysis should be discounted at the WACC to reßect the cost of both equity and debt; however, the cost of equity is difÞcult to estimate. The most appropriate solution may be to estimate the cost of equity by using the capital asset pricing model (CAPM). This model estimates the cost of equity by explicitly deÞning the relationship between risk and reward. One fundamental assumption of the CAPM, however, is that investors are rational—that is, they will seek to minimize their investment risk through portfolio diversiÞcation; therefore, the cost of equity estimated using this approach reßects only systematic risk, not projectspeciÞc or unsystematic risk. However, the addition of an investment-speciÞc risk premium to the cost of capital to reßect a project’s risk is theoretically unsound and will again generate valuation inaccuracies (James M and Koller TM, 2000). Risk-Adjusted Net Present Value Analysis. A better approach may be risk-adjusted NPV (rNPV) analysis. rNPV analysis couples classic DCF/NPV analysis with probability-weighted scenario analysis. In rNPV, analysts forecast a limited number of scenarios; initially, they construct a cash ßow proÞle for a base scenario. Subsequently, they design both optimistic and pessimistic scenarios to reßect, for example, high, medium, and low market penetration or delays to product launch, as shown in Figure 2. The critical phase of this analysis is the assignment of outcome probabilities to each scenario. The expected value of the investment is determined from these outcome probabilities and may then be High sales $250 million 2.5% probability Expected value = $0.625 million

i

Net Present value = Σ eCFi /(1+r)i t=1

Where eC F = expected cash flow, r = cost of capital.

Preclinical

Phase I

Phase II

CF1

CF2

CF3

Phase III

High sales, late launch $250 million 7.5% probability Expected value = $1.875 million

Sales of approved drug (brand) Low sales, late launch $50 million 20% probability Expected value = $1 million

Does not reach market: sales $0 40% probability Expected value = $1 million

Low sales, $50 million 30% probability Expected value = $1 million

Year 1

Year 15

FIGURE 2. Model of risk-adjusted net present value.

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237

discounted to current value terms by applying a cost of capital estimated by using the CAPM. Shortcomings of both DCF and rNPV analysis include the use of point estimates of valuation drivers—for example, future revenues, patient compliance, patient numbers, and market share (variables that have associated probability distributions)—in the valuation process. Consequently, a project’s expected value calculated using these approaches may not capture the value of all potential outcomes. Monte Carlo Analysis. In contrast, the Monte Carlo analysis uses computer simulation based on a large set of probability-weighted scenarios to determine a venture’s terminal value. Analysts determine a range of estimates for potential value drivers—for example, market size, product pricing, manufacturing rights, and time to market. Then, they forecast potential payoffs by estimating the probability of certain outcomes. Finally, computer simulation is used to predict a probability distribution for the value of the venture, based on simultaneous changes in variables; the expected value of these outcomes can be discounted to current value terms by using a WACC estimated by using the CAPM. Examples of probability distributions that could be generated are shown Figure 3. A common criticism of this approach is that, by the nature of the complex simulation, the value drivers underlying the model are not visible. The problem of model transparency may be addressed more effectively in the rNPV approach, where the critical assumptions and interactions used in the construction of the model are more visible. Having identiÞed the sensitivity of a model to different business drivers, management can endeavor to maximize company value by manipulation of those drivers. A more general criticism of discount-based approaches is that they value businesses statically, assuming that a company’s assets are held passively. None of these models account for attempts to maximize project value by active management, especially during preclinical stages. Furthermore, risks, time frames, and potential payoffs are extremely difÞcult to quantify for projects that are in preclinical stages. Therefore, discount-based approaches may be most appropriate for valuing projects that have entered the clinical phase of development, where analogue models can be used to more accurately establish likely time frames and outcomes. Real Options Valuation. For projects still in preclinical development, options valuation methods may more accurately reßect the value of new information and operational ßexibility. The ßexibility and uncertainty in terms of future cash ßows that are common to investment in early-stage biotech are also characteristic of Þnancial “call” options. A call option gives the bearer the right, but not the obligation, to purchase an underlying commodity. During early-stage investment in biotech, the initial investment in a project is effectively a call option on future investments: management has acquired the right but not the obligation to fund the project further at a later date. Such “real options” recognize and value uncertainty and operational

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Frequency

Frequency

40%

$3

100%

$8 Price per day

Compliance

Frequency

Frequency

1,000,000 5,000,000 Number of patients

5%

40% Patient share

Project value = sum of expected values Frequency

0 billion

5 billion Value ($)

FIGURE 3. Sample probability distributions generated by Monte Carlo analysis.

ßexibility, accounting for the value of being able to wait to invest further or to modify a project on the basis of better information or diminishing uncertainty. Financial option pricing tools such as the binomial option pricing model (BOPM) and the Black–Scholes–Merton (BSM) model have been developed to address the issues of investment irreversibility, timing, and uncertainty. The value of a Þnancial option is a function of the underlying commodity’s stock price; the option exercise price; the time to option expiration in years; the annualized, riskfree rate of return; and the standard deviation of the annualized returns on a stock (i.e., stock volatility, an accepted measure of risk). In the case of real options, the value of the option is a function of the current value of the expected cash ßows generated by the project, the current value of the

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expenditures required to undertake the project, the time required for the project to reach the market, the risk-free rate of return, and the risk associated with the underlying assets, which is best estimated using the standard deviation of returns for a typical biotechnology stock (Figure 4). Biotech projects face both technological and market uncertainty. In ROV terms, they are classiÞed as compound rainbow options—that is, they involve a sequence of options, each contingent on those preceding (e.g., to advance to stage II clinical development, a therapeutic compound must Þrst be successful in stage I)—and on multiple sources of uncertainty (e.g., market uncertainty, research uncertainty, and time-scale uncertainty) (Harrison M and Lerer L, 2002). ROV is considered suitable for early-stage biotech because it recognizes and values this uncertainty and operational ßexibility, thereby accounting for the value of being able to wait for new information before increasing investment in a project. Furthermore, in capturing the value of ßexibility, ROV may uncover additional value that can change recommendations for a project at several stages in its evolution—a characteristic that may improve investment efÞciency when assessing marginal opportunities. An additional beneÞt of ROV is that the effect of alterations to value drivers can be observed directly and used to maximize an option’s value prior to exercise

SE−dt * {N(d1)} − Xe−rt * {N(d2)} Where d1 = {ln(S/X) + {r− d +s 2/2}/s * √t Notes: S = Stock price; x = Exercise price; d = dividends; r = Risk-free interest rate; s = Uncertainty; t = Time to expiry; N(d) = Cumulative normal distribution function.

Scenario

Financial option

Real option

S

Stock price

Present value of expected cash flows

X

Exercise price

Present value of investment

T

Time to expiry

Length of delay until next investment

s2

Volatility of stock price

Uncertainty of expected cash flows

R

Risk free interest rate

Yield of a risk-less security of similar maturity

d

Dividends

Value lost over the duration of the option

FIGURE 4. Drivers in financial and real option values.

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VALUATION MODELS FOR EUROPEAN BIOTECH

(i.e., further investment), at which point the option should be “deep in the money.” ROV is especially valuable for projects that carry a high level of uncertainty but offer opportunities to dispel this uncertainty as new information becomes available, which is typical of preclinical development projects in biotech. In contrast, ROV is unsuitable for assessing projects with limited or no uncertainty or short time frames; DCF-based analyses, discussed previously, may enable more accurate assessment of these projects. CHOOSING THE APPROPRIATE VALUATION METHOD FOR A BIOTECH PROJECT No single valuation method can be applied universally to determine project value because every method involves assumptions compounded by assumptions. However, a combination of NPV and ROV may fulÞll the dual need to value ßexibility and to accurately and explicitly acknowledge the cost of biotechnology capital. In a modiÞed NPV analysis, as shown in Figure 5, the current value of an expected stream of cash ßows should exceed the cost of the investment by an amount equal to the value of the real option (Leslie JK and Michaels MP, 1997). Although individual project valuations based on this approach may remain noisy, portfolios of investment opportunities (such as the biotech sector) based on these valuations will be valued more accurately. Ultimately, a more precise reßection of intrinsic project value will generate the more efÞcient selection of investment opportunities and improved asset allocation required to restore investor conÞdence in the European biotech sector. The mature acceptance of project and equity valuations based on fair value will be a signiÞcant milestone in European biotech’s recovery from dependence on Big Pharma’s drugdevelopment capital.

Expected value = $0.625 billion Expected value = $1.875 billion Preclinical

Phase I

Phase II

Phase III

Sales of approved drug (brand) Expected value = $1 billion

CF1

CF2

CF3 i

Expected value = 0

Expected value = $1.5 billion

Net present value = ∑ eCFi /(1 + r)i t=1

Where eCF = expected cash flow, r = cost of capital. Project has a forecast NPV of $100 million in initial year but is in preclinical development. Option value should not exceed $100 million.

FIGURE 5. Modified net present value approaches.

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REFERENCES Harrison M, Lerer L. Real options for biotechnology valuation. Nature Biotechnology. 2002;20:223. James M, Koller TM. Valuation approaches in emerging markets. The McKinsey Quarterly. 2000;1:78–85. Leslie JK, Michaels MP. The real power of real options. The McKinsey Quarterly. 1997;3:97–108. Nelson TR, Mukherji A. Valuing biotechnology assets. Nature Biotechnology. 1998:16:525–529. Papadopoulos S. Quantifying the dream: valuation approaches in biotechnology. Nature Biotechnology. 1998;16: 55–56. Smith Ewing E. Pharma/biotech deals: are today’s collaborations tomorrow’s acquisition? Pink Sheet. June 2003.

Taking Global Action: Integrating Social Responsibility into Corporate Practices

SUMMARY The pharmaceutical industry is undergoing severe criticism from consumers, third-party payers, advocacy groups, and governments for a perceived failure in its social responsibility to provide affordable drugs to populations in need, especially in developing countries. These stakeholders say that pharmaceutical companies give higher priority to proÞts and shareholder demands than to humanitarian aid. Recent actions of the Bush administration—the call to implement a Medicare prescription drug beneÞt and the increase in funds to Þght AIDS in Africa—have intensiÞed the spotlight on the pharmaceutical industry. Consumers are beginning to view the industry with skepticism, and consumer perception can impact both legislative action and shareholder sentiment. As the level of negative perception rises, it will begin to impact bottom lines. In this article, we focus on the key issues that the pharmaceutical industry faces in fulÞlling social responsibilities in the United States and in developing countries, discuss actions that the industry has taken thus far to improve its reputation, and suggest other opportunities for integrating social responsibility into corporate practices.

BUSINESS IMPLICATIONS •

In the United States, Congress continues to debate legislative bills designed to contain escalating drug expenditure. Losing favor with key constituency groups at this time is a great danger for the pharmaceutical industry: effective lobbying from consumer groups could lead to passage of legislation that will

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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INTRODUCTION

•

•

243

markedly curtail the life of a drug by introducing generics to the market more quickly. Investors are starting to fear that a perceived lack of social responsibility, particularly in the HIV/AIDS crisis, is generating a growing negative public image that could eventually hurt pharmaceutical companies’ bottom lines. Investor action taken thus far seems a complete reversal of past strategies, namely, holding a hard line against the loosening of patent protection and pricing strategies for developing countries. Pharmaceutical companies have taken some measures to increase HIV/AIDS humanitarian efforts to developing nations. Between 1998 and 2001, for example, the industry donated more than $1.9 billion in Þnancial assistance and medicines for HIV/AIDS to the developing world. Although the actions that pharmaceutical companies have taken thus far have received some positive feedback from social stakeholders, the overwhelming message the industry continues to receive is that it is not doing enough to minimize public health crises in developing countries. We expect that the industry will listen more closely and respond to the calls for action because it faces increasingly negative public opinion.

INTRODUCTION The pharmaceutical industry has been greatly criticized by consumers, thirdparty payers, advocacy groups, and governments regarding the level of access to drugs among populations in need, such as the elderly population in the United States or the patient populations of developing countries. These stakeholders say that pharmaceutical companies are not doing enough to improve access to their drugs—for example, by lowering drug prices for needy populations. Moreover, stakeholders say that pharmaceutical companies give higher priority to proÞts and shareholders’ demands than to the humanitarian aid they should be providing. Improving access to pharmaceuticals is a critical issue for consumers in both the developed and developing worlds, even though their circumstances differ greatly. In the United States, overall health care is among the best in the world; still, many people do not have access to the medications they need. In the least developed countries (LDCs)—that is, countries with per capita incomes of less than $2000, such as the countries of sub-Saharan Africa—the health care demand is tremendous. Seventy percent (29.4 million people) of the world’s HIV/AIDS population are sick or dying of the disease in sub-Saharan Africa. The urgent need for pharmaceuticals in this region is complicated by lack of an adequate health care infrastructure. Also, manufacturing and distribution channels are not in place to deliver medications to populations in need. Aid to these countries must come from external sources—namely, global advocacy groups, governments of developed nations, and multinational companies. Recently, action taken by the Bush administration—speciÞcally, the call to implement a Medicare prescription drug beneÞt and the increase in funds to Þght AIDS in Africa—has intensiÞed the attention given to the global actions of the

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pharmaceutical industry. The question raised by some stakeholders is: How much can and should these companies be held responsible? The challenge of maintaining social responsibility is greater now than in the past because consumer awareness of these issues has increased. U.S. consumers have long been shielded from the costs of prescription drugs, but this situation is changing as their out-of-pocket expenditure grows and media coverage of escalating drug prices increases. Globally, there is growing awareness among consumers in developed countries of the health care needs of people living in the LDCs. Advocacy groups and nongovernmental organizations (NGOs) that represent populations afßicted with disease in developing countries are appealing to governments of developed nations and to the pharmaceutical industry to do more to improve health care in developing countries. One major problem is that developing countries are not commercially viable markets for pharmaceutical companies; therefore, diseases that afßict the populations of developing countries are not high on company priority lists. As demands for greater corporate social responsibility increase and pharmaceutical companies’ efforts to respond remain limited, the industry must face the consequences of the negative publicity it has been receiving: for the Þrst time, consumers are beginning to view the industry with skepticism, and because their inßuence can impact legislative action and shareholder sentiment, consumer perception is critical to the industry’s welfare. Pharmaceutical companies have always been concerned primarily with getting the greatest return on their investment to ensure sustained growth and funding for future projects. As the amount of negative publicity increases, it will impact company bottom lines—a point that shareholders are slowly recognizing and acting on. For the Þrst time, we see shareholders pressuring the pharmaceutical companies to take greater interest in expanding drug access for people in need. In reaction to the pharmaceutical companies’ initial, controversial response to the HIV/AIDS drug-pricing issue—which included Þghting for patent protection and minimizing drug discounts before improving drug access in order to reduce the threat of parallel trade—investors increasingly call on companies to protect their corporate image by demonstrating social responsibility in developing countries. In this article, we focus on the key issues that the pharmaceutical industry faces with respect to its social responsibility in the United States and in developing countries, the increasing connection between their actions and their proÞts, and the actions the industry has taken thus far to better its reputation. CRISIS IN THE UNITED STATES In the United States, the most public issue confronting the pharmaceutical industry is improved drug access for seniors, who consume 40% of all prescription drugs dispensed—on average, three prescriptions per senior. The issue came to the forefront in the mid-1990s as prescription drug expenditure began to increase at a double-digit rate. Because some of this increased expenditure is related to price (other reasons include new medicines on the market and a growing

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elderly population), pressure on the industry to minimize these escalating costs has continued to grow. Meanwhile, the Medicare prescription drug beneÞt debate continues in Congress. Escalating Health Care Costs: Who Is Responsible? In the United States, overall health care costs have been rising steadily since the 1960s. The perceived source of these escalating costs has shifted over time. Historically, when the predominant cost driver of health care has been identiÞed, stakeholders in government and the private sector have taken action to minimize its impact on health care spending. A concern for drug makers is that recently the tide of public perception seems to have turned against the pharmaceutical industry. During the 1970s, the prevailing belief was that physicians were charging too much for care. The response to the perceived increase in physician costs was the rise of the managed care industry. With managed care in place, physicians had to abide by strict cost-containment guidelines that aimed at lowering costs for physician services. Consequently, physicians have been restricted in the amount of time they can spend with each patient, the amount of service they are allowed to provide, and the referrals they can give their patients. Initially, consumers welcomed managed care, but soon these stakeholders decided that it was too restrictive for the amount of money charged to patients. Dissatisfaction soon shifted away from physicians to the managed care industry. Consumers and health care providers strongly opposed the restrictions the managed care industry was placing on physicians and on patient access to health services. These constituency groups believed that the managed care industry was implementing cost-containment strategies only to increase its own proÞts. During the mid 1990s, consumer and physician pressure forced the managed care industry to develop new products that offered patients greater choice—for example, preferred provider organizations and point of service plans. The availability of these options relieved some of the pressure on the managed care industry. Soon, the cost driver label shifted from the managed care industry to the pharmaceutical industry. As an array of managed care options became available, the cost of drugs began to increase. The managed care industry, one of the primary payers for prescription drugs, was the Þrst to notice. The managed care industry claims that prescription drug expenditure is the primary reason why health care costs are skyrocketing. Going one step farther, the managed care industry and other payers in the United States say that the pharmaceutical industry is putting its own proÞts and its responsibility to shareholders before the public good. Turning this perception around is critical to the pharmaceutical industry’s future, particularly when the U.S. government is working keenly to reduce prescription drug expenditure: currently, Congress is debating the Medicare prescription drug beneÞt and various other cost-containment strategies. Meanwhile, pressure from payers has already forced pharmaceutical companies to increase their performance of pharmacoeconomics studies and to minimize the production of “me-too” drugs.

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Medicare Prescription Drug Benefit (a Recent Historical Background) The continuing debate on Medicare relays a strong message that pharmaceutical drugs are too costly, a belief that received further attention when the proÞtability of pharmaceutical companies came under scrutiny by the mainstream media. If this reform improves seniors’ access to essential medications, it would lessen both criticism of drug prices and the pharmaceutical industry’s proÞtability. If the reform fails to improve seniors’ access to prescription drugs, criticism of the pharmaceutical industry will intensify. For example, in 2002, when four Medicare prescription drug beneÞt bills failed in the Senate, senators passed the Greater Access to Affordable Pharmaceuticals Act (GAAP) to speed the introduction of generics into the market. Another example of reducing drug costs for seniors came in July 2003, when the House passed H.R. 2427, a bill to allow the reimportation of pharmaceuticals from Canada. The historical background is as follows: The congressional debate on a Medicare prescription drug beneÞt ended unsuccessfully in the summer of 2002. However, the debate strongly relayed the message that pharmaceutical drugs are too costly, a belief that received further attention when the proÞtability of pharmaceutical companies came under scrutiny by the mainstream media. During the summer of 2003, Congress worked again to pass bills for a prescription drug beneÞt. The House of Representatives and the Senate each passed its own version of a bill. The two houses must now compromise on the points of contention—namely, how the program will be funded and run. If this reform is enacted and it improves seniors’ access to essential medications, it would lessen both criticism of drug prices and the pharmaceutical industry’s proÞtability. If the legislation is not enacted, however, or if it is enacted but fails to improve seniors’ access to prescription drugs, criticism of the pharmaceutical industry will intensify. For example, in 2002, when four Medicare prescription drug beneÞt bills failed in the Senate, senators passed the Greater Access to Affordable Pharmaceuticals Act (GAAP) to speed the introduction of generics into the market. Another example of reducing drug costs for seniors came in July 2003, when the House passed H.R. 2427, a bill to allow the reimportation of pharmaceuticals from Canada. This bill is likely to fail because of abundant opposition in the Senate; however, it makes the point that Congress will continue to attempt different strategies to reduce drug costs. Losing favor with key constituency groups at this time would pose great danger to the pharmaceutical industry. Effective lobbying by consumer groups could lead to passage of legislation that will markedly curtail the continued evergreening of patents that drug companies have relied on to sustain product revenues. With increased government intervention, the threat of price controls in the United States becomes greater. Furthermore, pharmaceutical companies must recognize that the industry needs to ally itself with health care consumers and payers, both private and public, because alliances with these groups increase the likelihood that they will

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support the pharmaceutical industry. Therefore, the industry’s active support of a Medicare prescription drug beneÞt will work in its favor. The pharmaceutical industry is undergoing severe criticism from consumers, third-party payers, advocacy groups, and governments for a perceived failure in its social responsibility to provide affordable drugs to populations in need, especially in developing countries. These stakeholders say that pharmaceutical companies give higher priority to proÞts and shareholder demands than to humanitarian aid. Recent actions of the Bush administration—the call to implement a Medicare prescription drug beneÞt and the increase in funds to Þght AIDS in Africa—have intensiÞed the spotlight on the pharmaceutical industry. Consumers are beginning to view the industry with skepticism, and consumer perception can impact both legislative action and shareholder sentiment. As the level of negative perception rises, it will begin to impact bottom lines. In this article, we focus on the key issues that the pharmaceutical industry faces in fulÞlling social responsibilities in the United States and in developing countries, discuss actions that the industry has taken thus far to improve its reputation, and suggest other opportunities for integrating social responsibility into corporate practices. BUSINESS IMPLICATIONS •

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In the United States, Congress continues to debate legislative bills designed to contain escalating drug expenditure. Losing favor with key constituency groups at this time is a great danger for the pharmaceutical industry: effective lobbying from consumer groups could lead to passage of legislation that will markedly curtail the life of a drug by introducing generics to the market more quickly. Investors are starting to fear that a perceived lack of social responsibility, particularly in the HIV/AIDS crisis, is generating a growing negative public image that could eventually hurt pharmaceutical companies’ bottom lines. Investor action taken thus far seems a complete reversal of past strategies, namely, holding a hard line against the loosening of patent protection and pricing strategies for developing countries. Pharmaceutical companies have taken some measures to increase HIV/AIDS humanitarian efforts to developing nations. Between 1998 and 2001, for example, the industry donated more than $1.9 billion in Þnancial assistance and medicines for HIV/AIDS to the developing world. Although the actions that pharmaceutical companies have taken thus far have received some positive feedback from social stakeholders, the overwhelming message the industry continues to receive is that it is not doing enough to minimize public health crises in developing countries. We expect that the industry will listen more closely and respond to the calls for action because it faces increasingly negative public opinion.

Consumers Want to Benefit from Better Drugs at the Best Cost U.S. consumers of prescription drugs have been and continue to be strong proponents of the research and development conducted by drug manufacturers. For

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consumers, new has always been synonymous with better. As consumers learn more about their health care options through tiered copayments, disease management programs, increased use of the Internet, and direct-to-consumer (DTC) advertising, they are becoming more involved in their health care decisions, particularly decisions about pharmaceuticals. They are more apt to seek multiple sources of information about wellness or disease, and they are better able to question medical practitioners about their health concerns. To date, this trend toward increased consumer empowerment has served the pharmaceutical industry well, because the result has been increased retail sales of medicines. However, this same trend may now be responsible for some erosion of consumer support for pharmaceutical companies. Consumer support has begun to waver because out-of-pocket drug expenses have increased steeply over the past Þve to seven years. On average, Medicare beneÞciaries pay out-of-pocket expenses of almost $2000 per year for prescription drugs. Some seniors spend more than $300 every month on their medications. Before prices soared, consumers wanted drug treatment at any cost; now, they seem to be moving toward wanting better drugs at the best cost. Tiered copayment systems, in particular, have increased consumers’ pharmaceutical price sensitivity. Third-party payers in the United States, most notably private insurance companies, have used tiered copayments as a cost-containment strategy, intending to sensitize consumers to the cost of prescription drugs and their cost–beneÞt trade-offs. Payers establish formularies, or lists of drugs for which they provide coverage. The Þrst tier of a formulary typically contains generic drugs, the second contains preferred branded drugs, and the third contains nonpreferred branded drugs. In tiered copayment systems, which are utilized by more than 95% of U.S. managed care organizations, consumers pay a fee based on the type of drug used. Consumer costs increase with each tier—typically, $8 for Þrst-tier drugs, $15 for second-tier drugs, and $20 for third-tier drugs, according to research by the Health Insurance Association of America. Prior to implementation of tiered copayments, consumers would pay a ßat fee for prescription drugs, whether generic or branded, or a minimal price differential—typically $5 for generic drugs and $10 for branded drugs. In the three-tier system, the increased price differential between tiers intensiÞes consumer involvement in the decision-making process because the choice of drug directly affects their out-of-pocket expense. Consequently, they are more likely to understand the difference between generic drugs and branded drugs, and this understanding tends to reinforce consumers’ belief that pharmaceutical companies are more concerned about their responsibility to shareholders than about their social responsibility. The Pharmaceutical Industry’s Response in the United States To date, the pharmaceutical industry has done little to counter the negative publicity it has received in the United States. This unresponsiveness will only reinforce the complaints of health care stakeholders. The industry must expand and continue its efforts toward social responsibility if it wishes to avert the

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imposition of price controls in the United States, the threat of which will become greater as government intervention there increases. The United States is the most lucrative market for pharmaceutical companies because it is the only majormarket country that has no price controls in place. Recently, in an effort to counter negative opinion, the Pharmaceutical Research and Manufacturers of America (PhRMA) released a statement on drug costs based on a Centers for Medicare and Medicaid (CMS) study that found that 10 cents of every health care dollar goes to prescription drugs. Pointing to the CMS Þndings, the PhRMA statement said that prescription drug expenditure is a small percentage of total health care spending and that any claim that prescription drugs are the cause of skyrocketing health care costs is therefore unsubstantiated. Unfortunately for the pharmaceutical industry, health care consumers and payers are likely to view the PhRMA statement as self-serving and diversionary. As consumer opinion continues to evolve, shaped by increasing exposure to information, the pharmaceutical industry will likely beneÞt from making efforts to target the consumer constituency with information of its own, designed to prove the cost-effectiveness of prescription drugs. Otherwise, contending with legislative bills (such as those aimed at bringing generics to market faster) may become an even greater challenge for the industry as more consumer groups and legislators support such legislation. Another strategy for the industry is to proactively offer consumers greater discounts in order to preempt the possibility of legislation that might impact bottom lines more severely. One such effort to alleviate the burden of prescription drug costs began at the end of 2001, when timing of the passage of a prescription drug beneÞt was unclear. A few leading pharmaceutical companies decided to implement their own programs to beneÞt the low-income elderly by giving them discounts on their products. GlaxoSmithKline led the way by introducing a prescription drug discount card. Other pharmaceutical companies have followed suit, including Novartis, PÞzer, and Eli Lilly. More such proactive responses will help to improve the industry’s wavering reputation. Reactionary responses tend to damage the industry’s credibility because their intentions are most often interpreted as self-serving. Pharmaceutical companies must also consider these potential outcomes as they take action globally. A HEALTH CRISIS IN THE DEVELOPING WORLD The populations of developing countries suffer mainly from infectious disease. The diseases responsible for the highest death rates are diarrheal disorders, tuberculosis, malaria, and, most predominantly, HIV/AIDS—the only disease for which drug therapies are still patent-protected, which is the major reason why stakeholders are imploring pharmaceutical companies to step in. For the remaining diseases mentioned above, some generics are available, but manufacturing and distribution capabilities in the LDCs inhibit access to these drugs.

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Drug Development for Developing Countries Each year, 30–50 new medical entities (NMEs) are introduced worldwide. Almost all are directed at diseases that afßict the populations of developed countries, diseases such as diabetes, obesity, osteoporosis, and hypertension. By one estimate from the humanitarian medical aid agency M´edicins sans Fronti`eres (MSF), only 1% of NMEs that reached the market between 1975 and 2000 were aimed at diseases that mainly afßict developing nations—and only 10% of global health research funds are dedicated to diseases that account for 90% of the global disease burden. Also, the drugs that do exist have poor efÞcacy and safety proÞles—for example, melarsoprol, a 50-year-old drug used for treatment of trypansomiasis, is effective in only two-thirds of patients and can cause death in up to 10% of patients. This situation is unfortunate but inevitable: because innovation is difÞcult and therefore extremely expensive (approximately $500–800 million to bring a single drug to market), companies must allocate their research and development (R&D) resources to therapeutic areas that will give them the greatest return on investment. Most of the diseases that need to be addressed in developing countries are not found in the developed world. Groups such as the MSF have criticized pharmaceutical companies for not investing R&D resources in these diseases, saying that they consider only the commercial viability of their products. Because the prospect of pharmaceutical companies undertaking drug development for diseases of developing countries is bleak, MSF has taken matters into its own hands and organized the Drugs for Neglected Diseases Initiative (DNDI). The goal of this organization is to bring to market six to eight drugs speciÞcally for diseases of developing countries. The organization currently has $250 million that it intends to use for ten years of drug development. It has set up collaborations with other global organizations such as the Indian Council of Medical Research, the Pasteur Institute, and the Kenya Medical Research Institute. MSF has been hesitant to set up direct collaborations with the pharmaceutical industry, given their contentious relationship, but the potential for partnership exists. The funding that MSF has put together may be enough to start developing drugs in these areas, but it is most likely not enough to cover all expenses. Pharmaceutical companies should take advantage of opportunities to collaborate with MSF to speed delivery of drugs to countries in need. Separately, neither the MSF nor the pharmaceutical industry can bear the burden of such great expenditure, which the World Health Organization has estimated at $3 billion. Collaborations will spread the risk among several players, thereby minimizing the cost of failure. One recent example of such a collaboration is GSK’s alliance with the WHO and the ministries of health in countries whose populations suffer from lymphatic Þlariasis. GSK is a founding partner in the alliance and is donating its albendazole tablets free of charge until the disease is eliminated, a twenty-year commitment estimated at 5–6 billion tablets, or $1 billion. Still, pharmaceutical companies have much work to do to improve both their relationships and their reputations with such organizations.

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Pricing in the Developing World Drug companies are further criticized for not offering enough discounts on drugs that would beneÞt needy populations, most notably HIV/AIDS drugs. Companies have always charged prices that markets would bear, provided they derive at least some proÞt. In practice, the prices companies set depend greatly on per capita income, the nature of the health care system, local attitudes toward medicines, and national regulatory policies. For new products that only developed nations can afford, companies set high prices—for example, pharmaceutical prices in the United States are the highest in the major pharmaceutical markets that we cover. In most poor countries, health care expenditures account for 3–5% of the gross domestic product (GDP) and are funded mainly by public sources and out-ofpocket consumer payments, although nonproÞt organizations such as missions and international aid groups also play an important role. On the other hand, the United States spends upward of 15% of its GDP on health care expenditures; major pharmaceutical markets in Europe spend, on average, 10% of their GDPs on health care. Clearly, the amount that developing countries dedicate to health care is not meeting the demand. One option is to allocate more of the GDP to health care, but most of these countries cannot afford to allocate the large percentage of the GDP that would be required to produce appreciable results. Therefore, aid must come from governments of the developed world and from the multinational companies that operate there. Decisions to discount drug prices must come from individual companies. One of the greatest concerns for drug makers is the parallel trading that would likely occur if they lower prices of patent-protected drugs. To increase the number of pharmaceutical companies that offer discounts and the dollar amounts of discounts given, governments of the developed and developing countries would need to ensure protection against parallel trade and delivery to the targeted populations. Investors: A New Source of Pressure Investors are starting to fear that pharmaceutical companies’ perceived lack of social responsibility, particularly in the HIV/AIDS crisis, is strengthening a negative public image that could eventually hurt the companies’ bottom lines. The investor action taken thus far seems to be a complete reversal of past strategies, such as the hard line held against loosening patent protection and pricing strategies for developing countries. In April 2003, the California Public Employees Retirement System (CalPERS), which holds $760 million in GSK stock, voted unanimously to send the company’s CEO a resolution that urged GSK to reevaluate its humanitarian programs for AIDS drug access and generic antiretroviral drug licensing policies. Similarly, in March 2003, 12 leading European Þnancial institutions, representing approximately $945 billion in investments, wrote a letter to more than 20 European drug companies—including GSK, Novartis, and AstraZeneca—calling for better practice in corporate social responsibility. The pharmaceutical industry must heed such warnings because inaction on its part may have dire consequences—such as losing investors.

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Investors are now encouraging wider use of differential pricing, a strategy that allows developing nations to purchase drugs at prices lower than prices in developed countries. Additionally, investors want pharmaceutical companies to inßuence European and U.S. governments to increase their Global Health Fund donations for grants to the LDCs. Investors are concerned that a minimal response to the HIV/AIDS pandemic from the industry will do further damage to its reputation, harming future profitability and the ability to attract talented staff. Furthermore, as public criticism mounts, the HIV/AIDS drug policy dilemma could undermine any valid arguments for patent protection, thereby hindering research and development of new drugs. The Pharmaceutical Industry’s Response in the Developing World Heeding such warnings, pharmaceutical companies have taken measures to increase HIV/AIDS humanitarian efforts in developing nations. Between 1998 and 2001, for example, the industry donated more than $1.9 billion in Þnancial assistance and medicines for HIV/AIDS to the developing world. In 2001 alone, pharmaceutical companies collectively donated $546 million for humanitarian aid to LDCs, according to a report published by PhRMA. Several companies have set up departments to address humanitarian issues and programs that beneÞt consumers directly. Companies have also increased awareness of their efforts by including program descriptions in their annual reports. Social pressures continue to mount, however, and often the pharmaceutical industry’s efforts have been deemed inadequate to meet worldwide demands. Pharmaceutical companies should consider more collaborations with one another and with advocacy groups, NGOs, and governments, as GSK has done (described next). Joint efforts will enable faster results while dividing the burden of cost and risk among several players. Building relationships with organizations outside the industry may help to lessen the criticism these organizations typically inßict on pharmaceutical companies. GlaxoSmithKline: A Case Study in Corporate Social Responsibility After receiving the greatest level of negative publicity for its limited response to the HIV/AIDS crisis in Africa, GSK has implemented a series of initiatives to improve its image with respect to social responsibility. In 2002, GSK released a report on social responsibility that highlighted its $380 million donation to charitable organizations, including 6 million tablets of its HIV/AIDS therapeutic zidovudine and lamivudine (Combivir), a substantial increase from 2.2 million tablets in 2001. Additionally, GSK now offers differential pricing for all its antiretrovirals to not-for-proÞt organizations in 55 LDCs and has announced plans to launch several new products to combat diseases in the developing world. On April 28, 2003, the company further reduced not-for-proÞt prices on many of its HIV/AIDS medicines, including a 47% reduction in the price of Combivir to 90 cents per day in all LDCs. Also in April 2003, GSK announced that it will work toward launching new products for the developing world over the next Þve years.

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GSK intends to conduct R&D in HIV/AIDS, malaria, and tuberculosis—the top-priority drugs for LDCs, as listed by the WHO. Oxfam, a global advocacy organization, has recognized GSK’s humanitarian efforts. Still, GSK has not wholly escaped the critics: the AIDS Healthcare Foundation (AHF) brought a lawsuit against GSK for patent piracy of the AIDS therapeutic zidovudine (Retrovir). Originally Þled in July 2002, the case was dismissed in March 2003, but the AHF reÞled the case in late spring 2003. GSK attempted to dismiss the reÞled case, but in July 2003, the court rejected GSK’s motion to dismiss. MOVING FORWARD WITH SOCIAL RESPONSIBILITY The pharmaceutical industry’s efforts to alleviate global health crises have earned some positive feedback from health care stakeholders, but the industry continues to hear that it is not doing enough to minimize public health crises in both developed and developing countries. We expect the industry to listen more closely and respond to the stakeholders’ call to action because the public’s negative perception is impacting the industry greatly and investors are stepping in. The industry’s failure to act will result in consequences ranging from consumer dissatisfaction to implementation of price controls in the United States and loss of investors. Pharmaceutical companies can augment their social responsibility efforts by taking a more proactive role. Proactive behavior will enable companies to better shield themselves from adverse publicity. Large companies, particularly those that have widespread name recognition through DTC advertising, are at great risk of exposure to negative publicity. When companies conduct humanitarian programs, which should include collaborations with advocacy groups and government agencies, they must send clear messages to the stakeholders that they are working to fulÞll their commitment to social responsibility—and make sure that conßicting messages are minimized.

Global Branding Strategy: From the Blockbuster to the Targeted Model

SUMMARY The pharmaceutical industry is at an inßection point: the traditional mass-market strategy emphasizing blockbusters is losing ground to the new paradigm of personalized medicine, which will lead to targeted therapies. The blockbuster brand model will be increasingly challenged by the targeted brand model that addresses genotype-speciÞc segments. Adoption of the targeted model will require a fundamental reorganization of the biopharma value chain, from discovery to manufacturing and marketing. Companies must also rethink brand economics: they are accustomed to pursuing high-volume blockbusters that require huge salesforces and marketing expenses, but their portfolios will gradually shift toward smaller, higher-margin targeted therapies with lower sales costs and potentially higher return on marketing investment. In this article we explore these marketing challenges, providing examples of companies that are successfully applying the dual-branding models. BUSINESS IMPLICATIONS •

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The focus of R&D in the postgenomics era is shifting from populations to individuals. This change presents a challenge to the traditional blockbuster brand model, which must make way for the targeted brand model, marketed globally to genotype-speciÞc segments. Bioscience is driving innovation across industry sectors and leading to the emergence of biobrands, which range from bioengineered tissue to molecular diagnostics and drug/device combinations. These new biobrands will

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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increasingly rely on evidence-based marketing (based on clinical performance and hard clinical end points), although experience-based marketing (which highlights emotional perceptions) will continue to play a role. The choice of an evidence-based or experience-based marketing approach will depend on the therapeutic area. Products that address critical therapeutic areas such as oncology will require evidence-based marketing, while products addressing primary care diseases such as allergies may beneÞt from a combination of evidence- and experience-based marketing. Both for targeted therapies and for mass-market products, successful global branding depends on an early interface of marketing and R&D, a fast worldwide rollout, and a balance of centralized components (strategic positioning and a key message) and localized elements (channels, packaging, media mix, and advertising execution).

A PARADIGM SHIFT The pharmaceutical industry is at an inßection point: the dominant mass-market strategy (i.e., the blockbuster brand model) is losing ground to the new paradigm of personalized medicine (i.e., the targeted brand model). As postgenomic science shifts its focus from populations to individuals, the blockbuster brand model is increasingly challenged by the targeted brand model, marketed globally to genotype-speciÞc segments. Big Biotech operates on the targeted model for most major therapies, but Big Pharma still largely follows the mass-market model for its blockbusters (products with more than $1 billion in annual sales). Despite the increasing importance of the targeted model, an absolute shift to this model is unlikely; several factors will continue to drive mass-market blockbusters: •

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The fully capitalized development cost for a prescription drug now averages $897 million, according to a May 2003 report by the Tufts Center for the Study of Drug Development; this cost must be offset by global sales. Postgenomic technologies have a long payoff and are expected to raise R&D cost and risk in the short term (the next three to Þve years) as druggable targets rise from the current 500 to more than 5000. Patents are due to expire on several drugs worth $100 billion in this decade, so companies will feel pressure to develop new blockbusters. Megamergers have increased scale and growth needs. With annual sales above $10 billion for the top ten Þrms, double-digit growth is generally estimated to require at least $1 billion in new sales each year—that is, at least three new products annually versus the average of only one or two. Marketing costs have increased since the late 1990s, driven by such factors as the use of direct-to-consumer advertising in the United States. Sales costs are also escalating and yield diminishing returns as salesforces have grown faster than the physician population and successful calls become scarcer.

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While the pressure to produce megabrands has risen, research productivity has declined. In the early 1990s, an annual R&D budget of $15 billion produced 50 new chemical entities (NCEs); the industry now spends more than $35 billion to produce 30 new compounds. The escalating cost of clinical trials, as well as the increased risk and longer approval times for novel drug targets, account, at least in part, for the shift. As a result, many blockbusters are inlicensed. In 2002, as many as 35 of the 78 new medicines approved by the FDA came from biotechnology companies, and nearly 400 biologics are currently in clinical development. As biologics gain a greater share of Big Pharma portfolios, companies will have a greater need for a targeted approach. Adoption of the targeted model will require a fundamental reorganization of the biopharma value chain, from discovery to manufacturing and marketing. Furthermore, companies must rethink brand economics—they are accustomed to pursuing high-volume single blockbusters requiring huge salesforces and marketing expenses, but they now must Þnd a way to optimize portfolios of smaller, higher-margin, targeted therapies with lower sales costs and potentially higher return on marketing investment. (This paradigm shift and its implications for business models are analyzed at greater length by Francoise Simon and Philip Kotler (2003).) BIOLOGICS ARE DRIVING INNOVATION Boundaries are fading between the biotech and pharma sectors as top-tier biotechs have turned into full-ßedged biopharmaceuticals: Amgen’s market capitalization tops those of Roche and AstraZeneca. Amgen’s leading product, epoetin alfa, is the industry’s second best-selling drug (after PÞzer’s Lipitor, which garnered $7.9 billion in 2002); sold as Epogen by Amgen and outlicensed to Johnson & Johnson under the brand name Procrit, it reached nearly $6.6 billion in 2002. In 2002, eight biologics were in the billion-dollar club, and many have a pattern of sustained high growth, given that they target critical therapeutic areas and have limited competition. Johnson & Johnson’s arthritis therapy inßiximab (Remicade) grew by 80% from 2001 to 2002, and its anemia drug epoetin alfa (Procrit) still registered a 25% annual sales increase in 2002, a decade after its launch. In addition to biologics, rationally designed small molecules such as imatinib (Novartis’s Gleevec), a leukemia therapy whose annual sales potential tops $1 billion, will play an increasing role in Big Pharma portfolios. The increasing proportion of biologics in Big Pharma portfolios will have several implications for how those companies approach drug development and branding. •

Time frames must be adjusted. According to the Tufts report mentioned previously, biologics had faster approval times in 2000–2003 than they did in the 1980s, but clinical development time has greatly increased since 1980 as a consequence of close regulator scrutiny of these novel therapies and greater product complexity.

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Challenges in manufacturing must be overcome. Dedicated protein production facilities must be developed early in the development cycle. Biomanufacturing facilities are in short supply, however, and they take four to Þve years to become fully operational. Monoclonal antibodies (MAbs) pose the greatest supply problem, given that they are used in fairly high doses for long-term chronic diseases. Approximately 300 MAbs are currently in the pipeline; industry analysts expect that more than 50 of them will be approved by 2010. This expansion will require a Þve- to sixfold increase in cell culture capacity (Dvorin J, 2001). Marketing organizations must accommodate the coexistence of two distinct models: mass-market blockbusters and targeted therapies, each with different distribution channels, pricing, and communication strategies.

CHALLENGE TO BIG PHARMA: ADOPTING THE TARGETED MODEL Advances in genomics have opened the door for the development of medications designed to target speciÞc genotypes (for greater efÞcacy and tolerability), and they have yielded tools that change the nature of drug development such that chemical screening is giving way to rational drug design. These shifts will require a rethinking of the concepts of health, disease, and therapies. The concept of biobrands, developed by Simon and Kotler in Building Global Biobrands, encompasses a wide range of products from bioengineered tissue to diagnostics, devices, drugs, and cosmeceuticals. This will lead companies to shift from drugs to integrated solutions (drug/device or drug/diagnostic combinations). Redefining Disease: Molecular Subtypes In therapeutic areas such as oncology, a “single” disease is fragmenting into its molecular subtypes. Although the molecular basis of some diseases has long been known, therapies did not immediately follow. For instance, the molecular basis of sickle cell anemia was discovered more than 40 years ago, but that Þnding did not yield a deÞnitive treatment. Therapies based on a molecular association have emerged only recently; for example, in 1998, Genentech launched its monoclonal antibody trastuzumab (Herceptin) for a subset of metastatic breast cancer characterized by HER-2 overexpression. The subdivision of diseases into their molecular types would lead to market fragmentation, but it would also optimize drug responses and minimize adverse events. A meta-analysis of 39 prospective studies from U.S. hospitals over 32 years showed a 6.7% incidence of serious adverse drug reactions (ADRs); for 1994, more than 2.2 million inpatients were estimated to have serious ADRs, and 106,000 died, placing ADRs between the fourth and sixth leading causes of death (Bates D, 1998; Fremont-Smith K, 1998; Lazarou JB, 1998). The fallout from an ADR can have a strong negative impact on the drug maker. In 1998–2000 alone, seven drugs were recalled (Morris L, 2001): terfenadine (Seldane), mibefradil (Posicor), bromfenac (Duract), cisapride (Propulsid),

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troglitazone (Rezulin), and alosetron (Lotronex). The economic costs of drug recalls have skyrocketed, given that they include the opportunity cost of lost sales and greater liability costs. Redefining Therapies: Biobrands The targeted model will allow for a better longitudinal analysis of disease that includes diagnosis. Therefore, a paradigm shift to personalized medicine involves a multistep approach: 1. Identifying the molecular cause of a disease. 2. Using specialized diagnostics to learn which patients have that molecular abnormality. 3. Linking genotypes and response to a candidate drug. 4. Proving that the drug effects an actual molecular change. The link between detection of a molecular abnormality and treatment of that molecular abnormality in the targeted model creates the potential for biobrands, which include a diagnostic/drug combination. Genentech pioneered this concept with its monoclonal antibody Herceptin and its related assay HercepTest, designed to identify and block HER-2 overexpression in metastatic breast cancer. Genentech initially partnered with the diagnostics Þrm Dako to manufacture this dual product. A signiÞcant advantage will belong to companies with dual drug and diagnostics units (such as Roche and Abbott) and especially to companies with the further beneÞt of a presence in devices (e.g., Johnson & Johnson). Roche Diagnostics has formed large teams to develop molecular markers. Johnson & Johnson’s advanced diagnostics group focuses on molecular oncology, a Þeld which its pharmaceuticals unit is addressing simultaneously. Johnson & Johnson’s drug-eluting stent research aims to leverage the company’s device and its drug capabilities. The convergence of therapeutics and diagnostics will require a restructuring of two different business models. Discovery timelines, approval processes, distribution channels, customers, and margins are all different in the two sectors. Dual salesforces will have to collaborate to reach labs as well as physicians. Additionally, consumer programs will also be needed to ensure proper diagnosis and treatment compliance. From Experience-Based to Evidence-Based Marketing As regulators and payers tightly link premium prices with superior clinical performance and Web-enabled consumers directly access clinical data, the tendency to market me-too drugs will become unsustainable, and a shift will occur from experience-based marketing (focusing on emotional perceptions) to evidencebased marketing (highlighting a product’s clear differential advantage based on research data and deÞnitive clinical end points). As clinical practice is now led

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by evidence-based medicine, biobrands will increasingly rely on evidence-based marketing. Some primary care therapeutic areas will continue to follow the experience-based marketing model. However, the higher return on investment will belong to breakthrough targeted therapies, marketed globally but to particular niches. Such targeted therapies can reach blockbuster status because of their high margins and much lower marketing costs. Even within the primary care category, several factors are already forcing a shift from experience-based marketing to evidence-based marketing: • • • •

Regulators have less tolerance for me-too products. Payers are demanding pharmacoeconomic studies of drugs. Physicians are sensitized to drug recalls and want hard data on efÞcacy and safety. Web-enabled consumers have direct access to scientiÞc information.

Given the wide scope of biobrands, experience-based marketing will continue to coexist with evidence-based marketing. For instance, therapies to treat common allergies will likely be well served by the experience-based approach, whereas therapies for critical areas such as oncology will require evidence-based marketing. Consumer trends support a dual approach to marketing that depends on the target indication and audience. Across major markets, consumers are driven toward hard evidence by several factors: brand proliferation and confusion, Internet access, negative publicity surrounding drug recalls, and an interest in personalized medicine. A Harris Interactive survey (2002) of more than 1300 consumers found that, in health-related Internet searches, medical journals or research institutions were the Þrst or second information sources for consumers in the United States, France, Germany, and Japan. Pharma Web sites came fourth in the United States, seventh in Germany, second to last in France, and last in Japan, after news media, government sites, patient groups, hospitals, and doctors. Targeting New Segments Information technology and scientiÞc advances are transforming segmentation bases; this has already occurred for targeted therapies and is likely to spread to mass-market products. The starting segment for a targeted drug is a genotype. In addition, technographics, which determines how technologies are used by consumers and physicians, provide an inßuential base. In the HIV category, Web-enabled activists are instrumental in accelerating product approvals. A targeted drug for a life-threatening disease often attracts, very early in its development, a patient segment that is Internet savvy; in the case of Novartis’s leukemia drug imatinib (Gleevec), these patients were attracted as soon as Phase I results were announced. Among these patients, a small but vocal group of activist leaders emerged. Because patients with critical illnesses tend to form close communities that share information and advocate new treatments, these

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activists are a powerful complement to the critical inßuence played by opinion leaders in the medical community. Physician specialties relevant to targeted therapies also have tight global networks and are highly inßuenced by trial investigators and opinion leaders. These are the best awareness-builders for the drug, together with physicians participating in early-access programs. Although mass-market drugs are not developed to target a speciÞc gene mutation, patients’ responses to them are affected by genetic polymorphisms, thus leading to new segments. Genes coding for drug-metabolizing enzymes (such as cytochrome P4502D6) have polymorphisms that produce the phenotypes of poor metabolizers or ultrafast metabolizers. Each segment accounts for a sizable proportion of the population and for an equally signiÞcant share of adverse reactions, drug recalls, and high-cost litigation. For instance, some antidepressants are toxic in poor metabolizers and ineffective in ultrafast metabolizers of CYP2D6. These segments correlate with ethnicity in a “gene geography,” as drug response varies across ethnic groups (Meyer U, 2000; Roses A, 2000). For mass-market products such as antihypertensives or antiulcerants, some segments may be derived from the use of information technology: Web networkers (the most active and inßuential users) and relatively passive information seekers. These segments have counterparts among physicians, who diverge in terms of education, Web use, and attitudes; technology-inclined physicians are more receptive to e-detailing and Web conferencing. These segmentation bases entail speciÞc planning steps. When marketing targeted therapies, the following steps apply: • • • • •

Promote diagnosis of disease-related genotypes. Target medical opinion leaders via clinical trials and publications. Collaborate closely with patient activists and advocacy groups. Develop prelaunch awareness among physicians and patients. After launch, continue science-driven marketing with Phase IV clinical trials.

For mass-marketed drugs, key steps include the following: • • • •

If genotyping is not part of trial protocols, bank patient samples for retrospective testing in case of adverse reactions. Target medical opinion leaders via trials and publications. Segment patients by demographics, attitudes, and technographics. Develop awareness by collaborating with patient activists/advocacy groups.

IMPLEMENTING A DUAL BRANDING MODEL Among mass-market drugs, current blockbusters may eventually lead to “blockbuster families” targeting genotype-speciÞc patient groups, but manufacturers will try to retain their mass-market approach as long as possible, because subdividing a drug into its molecular subsets fragments the market. This evolution

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could be driven by mandated microsegmentation; regulators may demand in the future that genotyping be included in most clinical trials. Until then, the two branding models will continue to coexist. Targeted drugs have a higher return on marketing investment than mass-market products, since the focused model is much more cost-effective: •

• • •

The physician audience for a targeted brand may be as small as 5000 specialists worldwide who are reachable with a small, highly trained salesforce—for example, 100 representatives in the United States. By contrast, primary care brands require at least a 3000-person salesforce for U.S. sales. Patient pools tend to be small, highly motivated, and more effectively reached via the Internet than expensive mass media. Targeted therapies are often supplied directly to physicians or hospitals, thereby eliminating the wholesaler share of proÞts. Because customized drugs are largely breakthrough products, they can command high prices. Furthermore, they have no generic or lower-priced competition and do not face great reimbursement barriers, although most companies developing such drugs provide free access programs for uninsured patients.

DUAL BRANDING: SUCCESS STORIES The two models were successfully applied by Novartis and PÞzer in a way that effectively balanced evidence- and experience-based marketing. As the Þrst rationally designed small molecule, Novartis’s Gleevec represents a new strategy for pharmaceutical companies. As the Þfth statin to be launched, PÞzer’s Lipitor could have stagnated in a crowded market, but instead it has become the world’s best-selling brand, thanks to its clinical advantage and science-driven marketing. Gleevec: A Targeted Oncology Breakthrough Novartis’s Gleevec spent decades in development, but it later broke several records. Its clinical trials, which began in August 1998, lasted only three years—half as long as the standard trial duration. Gleevec broke regulatory records, too, with the fastest-ever FDA approval time (72 days) in the United States, where it was approved in May 2001, and speedy reviews in the European Union and Japan, both of which approved the drug in November 2001. It has gone on to enjoy market success and demonstrates the success of evidence-based branding of a targeted therapy. Gleevec created a new class in oncology, the signal transduction inhibitors. Its initial indication was chronic myeloid leukemia (CML). In 95% of CML patients, an abnormal Philadelphia chromosome was found; this led to increased activity of the enzyme tyrosine kinase, which stimulated the growth of leukemia cells. Gleevec is a potent inhibitor of tyrosine kinase. CML was a small market with high unmet need. CML accounted for 20% of all leukemias, and the physician market, comprising specialists in hematology and oncology, was also very small—just 5000 worldwide.

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Clear Differential Advantage. The only curative therapy for CML was bone marrow transplant, but its use was limited to 15–25% of patients because age was a contraindication for some patients, and donor availability was limited. Furthermore, this therapy carried very high costs and mortality rates. The gold standard for CML in the chronic phase was interferon alpha, marketed by Roche and Schering-Plough, but it did not cure CML, and it had severe side effects. Unlike interferon and older cytotoxics, Gleevec had only mild side effects and a very high efÞcacy. In the Phase I trial, an unprecedented 100% of patients had a complete hematologic response (drop in leukemia cells in blood), and one-third had a complete cytogenetic response. Data-Driven Positioning. These Þndings fueled Novartis’s evidence-based marketing approach. Gleevec’s positioning stressed its unique targeted nature and its outstanding clinical data. The message was “precise targeting of the molecular abnormality that leads to CML,” and key points were “outstanding hematological response,” “unprecedented cytogenetic response,” and “well tolerated.” Novartis easily reached top opinion leaders since there are just 15 worldwide and many were trial investigators. A prelaunch expanded access programs and also reached 7000 patients worldwide. Patient-Driven Demand. Gleevec was driven by patient demand from its earliest development phase. As soon as interim data were presented at the American Society of Hematology in December 1999, patient activists lobbied for early access and fast-track approval. Patients also appreciated Gleevec’s innovative pricing approach. The worldwide price was set at $2200 per month, and treatment might continue indeÞnitely for patients who responded. A global assistance program extended exceptionally broad discounting to uninsured patients. While most patient assistance programs address only the lowest-income group, Novartis offered the following terms in the United States: • •

Drug free of charge to anyone earning less than $43,000 per year. Drug cost capped at 20% of income for those earning between $43,000 and $100,000 per year.

This program reinforced the high share of voice gained from the product’s innovativeness and earned Novartis broad and favorable press coverage. As could be expected, Gleevec had a rapid sales uptake, generating $615 million in 2002, with a peak-year sales potential of more than $1 billion. Novartis rapidly won approval for a second indication, for gastrointestinal stromal tumors (GIST), and the goal was to expand usage to other solid tumors. Trials were under way to document GIST survival rates and long-term end points. Physician Education. Many specialists still did not know that therapy success should be measured by cytogenetic testing (rather than only hematologic results), and dosing was sometimes suboptimal. Novartis was addressing this issue with online and ofßine communications.

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263

Lipitor: Winning in a Crowded Mass Market Launched in the United States and the United Kingdom in February 1997, atorvastatin (PÞzer’s Lipitor) was a latecomer to the cholesterol market; four statins preceded it. Nevertheless, it went on to become the world’s top brand, bringing PÞzer nearly $8 billion in 2002 global sales. It owes this status to its superior clinical performance and to evidence-based marketing. The statins were already a ten-year-old class when Lipitor was launched and the marketplace was crowded. Given this competitive situation, PÞzer designed Lipitor’s clinical trials as head-to-head comparisons with the other statins and also obtained fast-track status with the FDA based on a small South African trial that revealed Lipitor’s unique efÞcacy in a genetic disorder impairing cholesterol clearance. Trials showed that Lipitor had higher efÞcacy than the other statins at lower doses in reduction of low-density lipoprotein (LDL) and, unlike other statins, reduced triglyceride levels. However, earlier statins such as simvastatin (Merck’s Zocor) had demonstrated a reduction of morbidity and mortality among high-risk patients, and Lipitor had no such data—although it beneÞted from a positive class effect. Evidence-Based Positioning. The positioning of Lipitor was data-driven and stressed the statin’s high potency in LDL and triglyceride reduction. It was also priced lower than Zocor—but price alone would not have sufÞced. A key part of Lipitor’s success was its powerful premarketing. Thanks to a combined PÞzer/Warner-Lambert salesforce of over 2000 representatives, 86,000 U.S. physicians were visited before launch. A collaboration with the American Heart Association to promote national guidelines and educate consumers about the risks of hyperlipidemia also led to increased credibility. Teaching the science behind the brand was accomplished via a major medical education program. The following factors were key to Lipitor’s success: • • • • • •

Correct starting dose (10 mg) and comparative trial data. Correct pricing and managed care strategy. Evidence-based positioning. High prelaunch investments. Solid manufacturing capability. Decentralization, allowing quick decisions and implementation.

Lessons Learned Several lessons can be derived from these two cases: •

•

Evidence-based marketing applies across various therapeutic categories, because consumers play a greater role in product diffusion—consumers are increasingly knowledgeable and respond to data-driven differentiation. In critical therapeutic areas, consumers get involved and drive demand at the earliest development stage, as the Gleevec case shows; it is crucial to leverage the power of patient advocates together with that of medical opinion leaders.

264 •

GLOBAL BRANDING STRATEGY: FROM THE BLOCKBUSTER TO THE TARGETED MODEL

Some mass-market success factors that contribute to the success of traditional products may apply to targeted products as they expand into primary care categories. Lipitor is a good example, as it is differentiated by its superior performance, and PÞzer communicated this message in a well-timed sequence of prelaunch medical education and disease awareness campaigns.

BUILDING GLOBAL BIOBRANDS Both targeted and mass-market therapies need global scale action to recover research costs. Key success factors for successful global branding may be categorized as follows: • • •

Winning over time—managing beyond the life cycle. Winning over space—speedy, near-simultaneous launches in major markets. Balancing global planning and local adaptation.

Recent launches such as PÞzer’s sildenaÞl (Viagra) and AstraZeneca’s esomeprazole (Nexium) have shown that reaching a $1 billion per year sales level in less than two years after launch depends on a very early interface of marketing and R&D, as well as rapid entry into key markets. Global efÞciencies must also be balanced against local responsiveness. Global branding is easier for targeted therapies than for mass-market products because the specialist audiences for targeted therapies are tightly linked worldwide and treatment protocols tend to be standardized. Targeted therapies also tend to treat diseases with Web-enabled global patient communities. Pricing varies less for these therapies since their use is mandatory in life-threatening diseases and they have no generic competition. By contrast, mass-market drugs face several globalization barriers. Government payers are restricting reimbursement on me-too drugs. In addition, categories such as allergy or menopause vary widely in terms of treatment protocols and consumer attitudes. For both therapy types, the key beneÞt of global branding is scientiÞc consistency linked to an effective capture of opinion leaders in major markets. These components matter far more than the frequently used concept of economies of scale in marketing. Globally standardized campaigns are not necessarily more efÞcient than regional marketing because the media mix is not uniform (direct-toconsumer (DTC) advertising is restricted to the United States and New Zealand), and pricing, brand names, and channels vary across countries because of reimbursement and legal issues. Winning Over Time: Early Marketing/R&D Interface Current megabrands achieve peak sales three to Þve times faster than blockbusters of the previous generation did. Indeed, celecoxib (PÞzer’s Celebrex) reached a record $1.5 billion in its Þrst year on the market.

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265

In addition to the need to recover escalating R&D costs through global sales, companies face the added pressure from shorter time windows between competing products. The hypertension drug propanolol (AstraZeneca’s Inderal), launched in 1968, had a full decade on the market before a follower drug, metaprolol (Novartis’s Lopressor), Þrst appeared. By contrast, the two leaders of the COX2 inhibitor class, celecoxib (PÞzer’s Celebrex) and rofecoxib (Merck’s Vioxx), were launched within a few months of each other in 1999. Global speed to market is crucially important, given Þnite patent lives, and leading brands have demonstrated that a $1 billion sales level can be reached within one year, if global penetration is achieved quickly. SildenaÞl (PÞzer’s Viagra) reached more than 40 countries in its Þrst year on the market, but ßuoxetine (Eli Lilly’s Prozac) had taken six years to achieve the same global presence. To increase the speed of their global rollouts, biotechs have so far relied on partnerships with Big Pharma; now, however, top-tier biotechs such as Amgen are expanding their own global capabilities. Speed to market depends on an extensive premarketing process, starting as early as Þve years before launch. When Celebrex was launched, nearly 90% of U.S. primary-care practitioners were aware of it. Prelaunch planning is also critical to extend a product lifecycle: clinical trials for multiple indications are the most solid way to expand the market continuously. Medical Leadership Strategy. The key to a seamless lab-to-market process is an alignment of marketing outreach and development efÞciency. For both targeted and mass-market drugs, the crucial players are opinion leaders. Currently, 20% of the marketing budget is spent, on average, before launch, and up to half is spent on development of opinion leaders; this percentage is likely to increase given that these experts have a unique ability to create medical brand equity for a new therapy. Clinical trials are the most credible and powerful form of marketing in the prelaunch period. As investigators publicize Þndings, they help establish global treatment protocols and shape a drug’s identity. Large-scale Phase III trials also create a sizable body of early adopters. Opinion leaders also drive publications—another crucial premarketing component. Category leaders such as PÞzer’s antibiotic azithromycin (Zithromax) and Bristol-Myers Squibb’s anticancer drug paclitaxel (Taxol) were the subject of signiÞcantly more published articles than their competitors. Physician and Consumer Outreach. Another premarketing feature of blockbusters is their early share of physician detailing. Lipitor promotion to U.S. primary care practitioners started six months before launch and reached a 30% share of voice at launch. Targeted therapies have an advantage at this stage, because the salesforces they require are vastly smaller. While 3000 representatives are the minimum for a mass-market drug, a specialty drug targeting transplantation surgeons can be effectively launched with 15 salespeople in the United States.

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Key steps to winning over time include the following: • • • • • •

Accelerate speed to market with e-trial management and parallel development tasks. Plan early and across functions. Link medical and market leadership. Deliver data-driven positioning. Develop a global infrastructure. While consumer education is key to disease awareness and diagnosis, it is clear that DTC advertising is not necessary for the early success of targeted products. For example, Lipitor enjoyed a fast sales uptake because of its clinical advantage and extensive medical marketing; DTC did not begin until 18 months postlaunch.

Winning Over Space: Global Planning Versus Local Execution Across industry sectors, the challenges of global marketing are the trade-offs between central efÞciencies and local responsiveness and between scale economies and market focus. Companies tend to reconcile these trade-offs by focusing resources on key markets and by ensuring coordination through global brand teams. While biotechs and pharmas share a need for global scale, their geographic expansion strategies differ in part because of their respective resource levels and in part because of the nature of their products. While targeted therapies have fairly uniform target audiences worldwide, massmarket products face widespread cultural variances. The main trade-off for them is between global integration (which ensures branding consistency, speed to market, and the cross-border transfer of best practices) and local responsiveness (which reßects regulation variance and differing customer tastes and also supports the entrepreneurial drive of country managers). A consensus in the industry has emerged on the marketing activities that should be centrally planned and those that can be locally adapted. Brand positioning (including clinical performance, major differentiators, and key messages) should be centralized, but on the basis of a clinical proÞle that was developed with early input from the main country markets worldwide. While packaging varies, components that support the brand identity (such as symbol, logo, and trademark) should be standardized; however, uncontrollable factors often arise regarding the trade name. While companies begin a search for a trade name as early as Phase I or II, it is increasingly difÞcult to Þnd candidates that are not high-risk or already taken in at least one major market. Another difÞculty is that the FDA does not approve names until late in the development process, a practice that results in lastminute changes. Astra had to change the trade name of its antiulcerant omeprazole from Losec to Prilosec to avoid confusion with furosemide (Lasix). Other aspects of global marketing that vary based on market conditions are pricing, distribution channels, and media mix (e.g., DTC advertising through

REFERENCES

267

mass media in the United States and New Zealand, but only disease awareness via the Internet in other countries). SUCCESS FACTORS Key success factors in building global brands are as follows: • • • • • • •

Rely on evidence-based marketing for targeted therapies. Balance evidence- and experience-based marketing for drugs targeting highprevalence disease areas. Include genotypes and technographics in segmentation bases. Adopt a targeted branding model for biologics and specialty drugs. Accelerate time to peak sales with early marketing input into the development process. Increase speed to global markets with simultaneous registrations and multicountry launches. Balance centralized branding components (strategic positioning, key message) and localized components (advertising execution, packaging, channels).

REFERENCES Bates D. Drugs and adverse reactions: how worried should we be? Journal of the American Medical Association. 1998;279:1216–1217. Dvorin J. Large molecules: too late for big pharma? In Vivo. 2001;11:53–57. Fremont-Smith K, et al. Adverse drug reactions in hospitalized patients [letter]. Journal of the American Medical Association. 1998;280:1741. Harris Interactive survey. Harris Interactive Health Care News. 2002; 2(11). Lazarou JB, et al. Incidence of adverse drug reactions in hospitalized patients: a metaanalysis of prospective studies. Journal of the American Medical Association. 1998; 279:1200–1205. Meyer U. Pharmacogenetics and adverse drug reactions. The Lancet. 2000;356:1667–1671. Morris L. A terrible thing to waste. Pharmaceutical Executive. June 2001:146. Roses A. Pharmacogenetics and the practice of medicine. Nature. 2000;405:857–865. Simon F, Kotler P. Building Global Biobrands: Taking Biotechnology to Market. New York: Free Press; 2003.

Prospects for Genomics-Derived Drugs: New Approaches and Impact on Pharmaceutical Pipelines

SUMMARY Has genomics failed the pharmaceutical industry, or is it simply maturing? Most of the Þrst genomic drug candidates failed, often for lack of efÞcacy. But as existing genomics tools mature and additional tools emerge, researchers are gaining experience, and genomics research is moving much faster. Many new genomics tool companies have emerged, and many existing companies have revamped their business strategies. We believe that the second wave of genomics targets will be far better characterized than the Þrst wave, and we anticipate that several genomics-based drugs will emerge in the next four to six years. In this article, we examine the current state of genomics-based research, analyze companies that are focusing on genomics, and describe particular drugs and diagnostics that have emerged from this sector. We also forecast progress in the Þeld and identify remaining unmet technological needs.

BUSINESS IMPLICATIONS •

After initially generating a large wave of poorly characterized targets that the pharmaceutical industry could not efÞciently validate, genomics is beginning to deliver results at multiple points in the pharmaceutical research and development process.

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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272 •

•

•

•

PROSPECTS FOR GENOMICS-DERIVED DRUGS

Most companies that are still betting heavily on genomics-based targets have had to Þnd new strategies to compensate for their lack of products. In an effort to obtain more-immediate revenue, many companies have either inlicensed nongenomic drugs or are developing such drugs on their own. Because genomics has become widely integrated throughout pharmaceutical research and development, it has become more difÞcult to estimate its contribution to many current projects. In addition to its important role in target identiÞcation, genomics is now a major tool for target validation and clinical development. More than a dozen genomics-based drugs are in clinical trials. Most are aimed at low-prevalence diseases, but a few potential blockbusters could emerge, most notably GlaxoSmithKline’s Lp-PLA2 inhibitor for cardiovascular disease. We anticipate that several genomics-based drugs will emerge in the next four to six years. The emergence of new genomics tools is helping to speed advances in this Þeld. In particular, high-throughput genomics is becoming more reliable, and RNA interference (RNAi) techniques are being adopted rapidly to complement genomics approaches.

INTRODUCTION No scientiÞc Þeld has created so many expectations and disappointments as genomics. Hailed as a vital source of new therapies, genomics initially proved to be largely a source of frustration. Rather than generating a ßood of new drugs, it helped to push the major bottlenecks in drug discovery further downstream, away from target discovery and into target validation, where sufÞcient means to validate novel targets were lacking. Most of the Þrst genomic drug candidates failed, often for lack of efÞcacy. So, has genomics failed the pharmaceutical industry, or is it simply maturing? As an oft-quoted Lehman Brothers and McKinsey report pointed out in 2001, the targets that sprang from the Þrst wave of genomics technologies were many times less well understood than traditional targets. As a result of this greater complexity, fewer drugs are coming out of the pipeline despite the increased number of targets. Although many genomics technologies existed to help elucidate gene function, some of these technologies, such as systematic animal knockout platforms, were too slow to keep up with the ßood of new targets that sprang from genomics analyses. Certain tools, particularly DNA microarrays, proteomics, and genotyping, have needed time to mature. Early on, these tools generated a great deal of data, much of which proved unreliable because of poor experimental design, insufÞcient experience in understanding ideal biological conditions for such studies, lack of quality control, and immature analytical platforms. Nonetheless, genomics research has had many beneÞcial effects: it has generated a swell of potential new drug targets, many of them novel, and it has helped investigators to address major bottlenecks in drug discovery and development, such as the need to identify optimal animal models during target validation.

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273

Now, as existing genomics tools mature and additional tools emerge, researchers are gaining experience, and genomics research is moving much faster. In particular, high-throughput genomics is becoming more reliable, and the new technique of RNA interference (RNAi; used to speciÞcally block gene function) allows researchers to rapidly and efÞciently determine the importance of an interesting gene in a particular pathway and the likely results of inhibiting that gene. In addition, automation and integration of genomics tools with other key aspects of the drug discovery and development process are helping researchers to obtain more and better results. In this article, we examine the current state of genomics-based research, analyze companies that are focusing on genomics, and describe particular drugs and diagnostics that have emerged from this sector. We also forecast progress in the Þeld and identify remaining unmet technological needs. EARLY GROWING PAINS FOR GENOMICS The Human Genome Project excited much interest among investors and drug developers as a possible means of accelerating drug discovery and vastly broadening the range of drug targets explored. Scientists anticipated that genomic information would become extremely valuable when used in concert with new, much higher throughput tools for measuring gene expression, protein expression, genetic variation, and techniques for creating cell and animal models. The scientiÞc community believed that a huge “deluge” of data would spring from genomics and that a new generation of bioinformatics tools—from companies such as Celera, Incyte, GeneLogic, Doubletwist, and LION Bioscience—would emerge to Þlter targets from this ßood of data. But disappointment came swiftly and dramatically. Bioinformatics was probably the Þrst sector to be regarded as widely overhyped. Because of problems with data standards and quality, data Þltering was extremely difÞcult. In addition, many researchers started to realize that they needed more data than the current generation of tools could generate at a reasonable cost. Finding good targets depends largely on having a clear deÞnition of the disease being investigated; even a disease such as breast cancer can have many manifestations and affect many different kinds of patients. A target identiÞed in a postmenopausal woman, for example, might have no relevance in a premenopausal woman. Issues such as gender, age, environmental exposure, previous therapy, and natural genetic variation further complicate efforts to identify good targets. Investigators therefore need hundreds of well-prepared and well-chosen samples; Þnding enough such samples has been one of the most limiting factors in target identiÞcation. Consequently, the genomics industry has experienced a remarkable roller coaster ride. During 2000, 2001, 2002, and 2003, the numbers of announced genomics-related deals were 319, 381, 204, and 173, respectively. The number of high-value (more than $10 million) genomics deals has also drastically declined (Table 1). The terms of such deals are very different now because upfront payments are lower and the genomics Þrms are assuming more risk.

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TABLE 1. Select Top Genomics/Pharmaceutical Deals Deal Makers Vertex/Novartis

Millennium/Aventis

CuraGen/Bayer Millennium/Abbott

Rosetta Inpharmatics/ Merck Roche/deCode Genetics Wyeth-Ayerst Laboratories (Wyeth)/Genome Therapeutics GlaxoSmithKline/Exelixis deCode/Merck

Genome Therapeutics/ Amgen Sequenom/Procter & Gamble Lexicon/Bristol-Myers Squibb

Deal Focus (Therapeutic Area)

Date

Value ($MM)

Drug discovery, development, and commercialization for protein kinase targeting Drug discovery, development, and commercialization (anti-inflammatory indications) Drug discovery, development, and commercialization Drug discovery, development, and commercialization (obesity and diabetes) Acquisition of Rosetta by Merck

5/9/00

800

6/23/00

450

1/16/01

1340

3/12/01

250

7/20/01

540

7/2/01

300

7/16/01

118

10/28/02

350–569

9/27/02

90

1/02/03

104

12/17/03

30

12/18/03

300–450

Diagnostics discovery and development Extension of discovery agreement (osteoporosis) Drug discovery, development, and commercialization Drug discovery, development, and commercialization (obesity) Drug discovery and development (bone diseases) Drug discovery, development, and commercialization (osteoporosis) Drug discovery, development, and commercialization (central nervous system disorders)

This reversal of fortune is well represented by the stock prices of the ßagship genomics companies. The February 2000 announcement of the “almost complete” sequencing of the human genome brought investor interest in genomics to a peak, netting the sector a signiÞcant portion of the record $36-plus billion pumped into biotechnology by investors that year. Many companies’ stock prices soared: Celera Genomics’ stock reached approximately $250, Incyte Genomics (now Incyte Corporation) nearly $150, and LION Bioscience approximately $140. But the climate quickly soured, and investment in biotech plummeted to $10 billion in 2001. The sudden downturn in deals was not just a result of the high cost of genomics and the perceived low payoff. Other problems, such as the falling stock market and glaring gaps in product pipelines, made pharmaceutical companies “tool shy.” Instead of looking for new tools, companies were on a hunt for late-stage products that could be inlicensed to quickly Þll gaps in the pipeline. There was also widespread agreement that genomics tools “did not deliver” and that they required

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275

much more investment, in terms of development and training, than companies had expected. The steep degree of the downturn brought the very value of genomics into question. Many experts began to wonder openly whether genomics really would make a substantive impact on drug development more quickly than any previous technology—or whether it would, like other technologies, require the typical twenty years to produce major rewards. This speculation cast doubt not only on the genomics tool sellers but on companies such as Human Genome Sciences (HGS), Millennium Pharmaceuticals, and others whose platforms were considered genomics-centric. It is now clear that although genomics is helping to speed the discovery and development of many drugs, real “genomics-derived” drugs (i.e., drugs for novel targets discovered mainly by genomics methods) will probably take as long to develop as any other class of compounds. Because the targets are novel, more time and effort are required to validate them, and they are less predictable in the clinic. CURRENT STATUS OF GENOMICS TECHNOLOGIES Many researchers believe that they have only now begun to understand the best way to use genomics tools, both individually and in concert with other techniques. Huge advances have been made in experiment design, standard operating procedures, and data analysis methods, but much work still needs to be done. In this section, we review major areas of genomics research and assess their progress. Genomics has dramatically increased the number of available drug targets. Traditionally, drug discovery and development focused on approximately 500 well-established drug targets. Today, thanks to advanced genomics technologies, the entire genome of approximately 40,000 genes (and the estimated hundreds of thousands of proteins they may encode) is available for exploration. As a result of these advances, most drug discovery and development enterprises are investigating several times more targets at one time than ever before. Although major drawbacks still exist—for example, lack of efÞcient processes for distinguishing major players from minor players in a particular pathway—development of tools and methods for target validation has expanded greatly. Another major development is broader recognition of the need for strict quality control in highthroughput genomics studies in order to produce more reliable results; automation is making a signiÞcant difference in this respect. Finally, the genomics industry has experienced a major shift away from using gene expression, protein expression, and genetic variation to Þnd disease targets; instead, these tools are being used much more widely in the drug development phase for biomarker discovery and patient selection. Major unmet needs in genomic drug discovery and development include the following: •

Analytical software for evaluating images generated by high content analysis tools such as ultrafast microscopy for cellular assays and other sophisticated image capture devices.

276 • • • •

PROSPECTS FOR GENOMICS-DERIVED DRUGS

Laboratory automation solutions for microarrays, proteomics, high content analysis, genotyping, and high-throughput screening. Lower-cost, higher-throughput DNA microarrays, protein expression analysis platforms, and, in particular, genotyping technologies. Tools for data integration, including text mining, pathway mapping, and knowledge management. Better toxicogenomics tools.

Toxicogenomics tools are probably the area of greatest interest right now because these tools are so challenging and pose so many problems for the industry. Ideally, companies would like to have gene-expression signatures that signal the potential toxicity of a compound very early in the process. Several companies—including CuraGen, Iconix, and GeneLogic—offer tools and services in this area. Many other companies, including Abbott Laboratories and PÞzer, are working on such tools in-house. In silico (computer-based) modeling is another area of great interest, but it is in very early stages of development and is not generating much commercial activity. CHANGING GENOMICS STRATEGIES The genomics industry has changed drastically over the last few years and now includes tool companies as well as drug discovery and development companies. Because of the downturn in the genomics tool and database markets, many new genomics tool companies have emerged, and many existing companies have had to change their business strategies. Leading database providers Celera Genomics and Incyte Genomics (now Incyte Corporation) had to add entirely new drug discovery components to their businesses to convince investors that they could remain viable. Other companies, such as Sequenom and deCode Genetics, have had to dramatically accelerate their drug discovery agendas. Pioneering genomicsbased drug developer Millennium Pharmaceuticals has had no success with products developed in-house and has focused instead on developing nongenomic, inlicensed products, including its breakthrough drug for cancer, bortezomib (Velcade). Genome Therapeutics likewise, despite its success in Þnding intriguing new drug targets, has been stymied in its efforts to bring a genomics-based drug to development. HGS has moved the most genomic drugs to trials, by far, but it has also suffered numerous setbacks. Table 2 presents an overview of the pipelines of select genomics-based drug developers; Table 3 presents Þnancial proÞles of these players. SELECT GENOMICS LEADERS Celera Genomics and Celera Diagnostics Celera Genomics was founded in 1998 by Applera. Having sequenced the entire human genome by itself, while racing the international Human Genome Project to a draw in 2000, Celera has the advantage of having stood consistently at the

SELECT GENOMICS LEADERS

277

TABLE 2. Pipelines of Select Genomics-Based Drug Developers Company

Product

Indication

Status

CuraGen deCode Genetics Exelixis

CG-53135 DG-031

Oral mucositis Inflammation

Phase I Phase II (launched in 2004)

XL-119 XL-784

ExonHIT Therapeutics

EH-201

Bile duct tumors Cancer, renal, and/or cardiovascular disease Amyotrophic lateral sclerosis (ALS) Neurodegeneration Anthrax prevention

Phase III Phase I completed; Phase II planned Phase II

EH-202 Human Genome ABthrax Sciences Albuferon Albuleukin Albutropin LymphoStat-B LymphoRad 131

TRAIL-R1 MAb (HGS-ETR1) TRAIL-R2 MAb (HGS-ETR2) Rigel Pharmaceuticals ZymoGenetics

R-112 R-803 IL-21 rFactor XIII

rhThrombin TACI-Ig

Phase II Phase I

Hepatitis C Cancer Growth hormone deficiency Rheumatoid arthritis B-cell cancers, including multiple myeloma and non-Hodgkin’s lymphoma Solid tumors and hematological malignancies Solid tumors and hematological malignancies Asthma; allergies

Phase I/II Phase I Phase I completed; Phase II planned Phase II Phase I

Hepatitis C Cancer Congenital and acquired factor XIII deficiency; cardiac surgery Topical hemostat (i.e., blood coagulant) Systemic lupus erythematosus

Phase I Phase I Three Phase I studies (conducted in 2003)

Phase II

Phase I (in both United States and United Kingdom) Phase I

Phase I Phase I/II (in collaboration with Serono)

Note: Celera Genomics is not included in this table because its drug candidates are in preclinical development.

front line of the genomics data boom. However, human genome data alone were not a sufÞcient basis for a sustainable bioinformatics business, and Celera was forced to move in a new direction two years ago. Celera Genomics and its sister companies, Applied Biosystems and Celera Diagnostics, are all part of the parent Applera. To expand the value of its proprietary data, the three Applera Corporation companies launched the Applera Genomics Initiative (AGI) in summer 2001. The primary goal of this $100 million project was to discover single nucleotide polymorphisms (SNPs) in genes and regulatory regions by resequencing (i.e.,

278

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TABLE 3. Financial Profiles of Select Genomics Companies

Company Celera Genomics CuraGen deCode Genetics Exelixis ExonHIT Therapeutics Human Genome Sciences Rigel Pharmaceuticals ZymoGenetics

Year Founded

2002 Revenues ($MM)

2002 Cash Burn ($MM)

Market Capitalizationa ($MM)

Available Casha ($MM)

1998

88.3

80

1051

790

1991 1996

18.2 41.0

88 20

406 455

358 72

1994 1997

44.3 3.3

39 11 (2001)

526 NAb

200 NA

1992

3.6

197

1829

1049

1996

15.8

NA

308

57

1981

52.8

50

899

235

a Financial data accurate as of January 2004. b NA, not available.

sequencing again, to verify) approximately 25,000 genes and regulatory regions of 39 humans and 1 chimpanzee. More than 4 million SNPs are in the public domain; however, researchers have learned that all SNPs are not equally valuable as markers, and many occur only in certain populations. The AGI was completed in 2003. The researchers found 294,000 SNPs in genes and reported that approximately 75% of these SNPs were new ones that had not been previously described. Celera Genomics has conducted a variety of informatics- and laboratory-based studies and estimates that more than 45,000 of these SNPs are functional in that they affect the amount or type of protein expressed. The company’s participation in the AGI could help give it an advantage in the race to apply genetic variation to development of drugs and diagnostics for complex diseases. Celera Diagnostics, founded in 2000 as a joint venture between Celera Genomics and Applied Biosystems, is applying these SNP data to medical disease association studies aimed at developing novel in vitrodiagnostic products based on genetic markers. The company already has major deals with Abbott and Merck that put it in an excellent position to become a competitive diagnostics Þrm. Celera Diagnostics also has a product on the market that is designed to detect mutations in the HIV-1 that are associated with drug resistance; this product is not related to the company’s human SNP validation work. However, part of Celera’s goal is to get some diagnostics into the marketplace quickly to help fund its groundbreaking work in human genetic variation. Instead of studying families, Celera Diagnostics conducts its human genetic variation studies in large sets of patients that represent equal numbers of cases and controls. Recently, the company identiÞed genetic variations that appear to be predictive of increased risk for myocardial infarction. One variation is believed to be involved in inßammation; another is correlated with increased severity of

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cardiovascular disease in people younger than age 60. The study, which included more than 1000 patients, also conÞrmed the association between seven previously identiÞed markers and coronary heart disease. Data from the study were presented in September 2003 at the 13th Annual Symposium on Atherosclerosis in Kyoto, Japan. The company intends to complete further statistical analysis of the study data in order to evaluate the diagnostic and therapeutic potential of these markers. Celera Genomics is using the same proprietary data to generate drug targets, initially in inßammation, coagulation, and oncology. The company currently aims to develop its most promising small-molecule programs itself, but it has some partners and is seeking more. Celera Genomics also aggressively pursues proteomics-based target discovery, focusing on proteins expressed on the cell surface. Based on this work, the company has three therapeutic antibody development programs: pancreatic cancer, non-small-cell lung cancer, and colon cancer. All the company’s projects are in preclinical development. CuraGen CuraGen has one of the most extensive genomics platforms, which links multiple technologies through a bioinformatics core. Using this platform, the company has identiÞed 8200 “tractable” genomic drug targets—genes that appear to have characteristics that make them potentially suitable drug candidates. The company has patented more than 500 of these targets. As a result, the company is target rich—but most of the drugs it is developing for these targets are in preclinical development. In 2003, CuraGen began Phase I patient dosing trials for its protein therapeutic, CG-53135, which is being investigated as a possible treatment for oral mucositis, a side effect of chemotherapy and radiation treatments in cancer patients. The company also plans to test the drug as a treatment for inßammatory bowel disease (IBD). The company is developing 15 of its own novel protein therapeutics and collaborating with Abgenix on 15 fully human monoclonal antibody (MAb)-based drugs. Fully human MAbs are derived from human sources rather than animals such as mice. CuraGen also expanded its preclinical pipeline by exercising a license option on four fully human MAbs from its Abgenix collaboration. CuraGen’s 2001 $1.34 million deal with Bayer was one of the richest in biotechnology history and helped to ensure the company’s short-term survival. In collaboration with Bayer, CuraGen is developing six small-molecule drugs for treating obesity and diabetes. CuraGen and its collaborators need to start seeing more tangible results from the company’s genomic drug targets research, and more drugs against these targets must begin moving into clinical trials if investor conÞdence is to be maintained. DeCode Genetics Like Celera and GlaxoSmithKline (GSK), deCode Genetics has been focusing on using information about genetic variations to identify optimal disease gene

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PROSPECTS FOR GENOMICS-DERIVED DRUGS

targets. deCode has a unique approach that is starting to gain recognition. The company uses a combination of family linkage studies and case/control association studies. By studying families Þrst, deCode claims, the company can identify a much smaller initial region of the genome that has a high likelihood of harboring the gene of interest. This region is then studied using both SNPs and the more traditional microsatellite markers. The company’s key deÞning factor is its unique genealogy database of Icelanders, who have kept extensive genealogy records for more than 1000 years. In 2002, deCode published a high-resolution map of key markers in the human genome (Kong A, 2002). This “genetic” map was lauded for providing “substantially improved resolution compared to the best previous maps” (Weber JL, 2002). Such maps are useful because these markers are used to Þnd the locations of genes. If many disease-affected people share a certain marker, the area near and around that marker is a prospective site for the disease gene. Researchers have been debating the comparative value of using sibling pairs (one with the disease, one without), families, and unrelated case controls. Each approach has its drawbacks. It is not easy to get a large number of families or sibling pairs to do such studies; however, if unrelated cases and controls are used, the number needed is much larger because unrelated people may have completely different combinations of genes involved in the same illness. Through its combination approach, deCode reports, it has mapped more than 25 genes and identiÞed 15 that are associated with increased susceptibility to common diseases such as stroke, peripheral arterial occlusive disease, and schizophrenia. Most of these discoveries must be considered preliminary Þndings, which some critics suggest could apply only to the Icelandic population. However, deCode claims that it has found critical genes that are involved in large proportions of cases of these three diseases. Large studies by deCode and scientists at the Aberdeen Royal InÞrmary have veriÞed the association between schizophrenia and neuregulin 1, a link Þrst discovered by deCode. An initial study used samples from more than 800 Icelandic patients and unaffected relatives (Stefansson H, 2002). A subsequent study included samples from more than 600 Scottish patients and a similar number of unaffected people (Stefansson H, 2003). The deCode researchers reported that a haplotype deÞned by Þve SNPs and two microsatellite markers confers an almost twofold increased risk of schizophrenia. The company reports that mice with impaired neuregulin 1 have behaviors and neurotransmission disruptions that resemble those observed in human patients with the disease. deCode anticipates that this pathway will be a good source of markers and drug targets. In November 2003, deCode acquired an exclusive worldwide license from Bayer to develop and commercialize a small molecule (DG031), which the company says addresses an inßammatory pathway-related protein whose gene was isolated at deCode. Furthermore, the company reports that this gene predisposes to myocardial infarction (MI). Earlier clinical work was carried out on DG-031 for a different indication; those studies indicate that the drug is well tolerated and has no major safety issues. deCode is planning to launch a Phase II clinical trial

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of the drug early in 2004. In a reversal of the typical pharmaceutical and biotech roles, Bayer will receive royalties on milestones as well as sales if the drug is approved. deCode also has a longstanding and apparently productive deal with Roche for drug discovery and development as well as diagnostics. More recently, in February 2004, the company struck a high-proÞle deal with Merck; the sevenyear alliance will engage deCode’s pharmacogenomics expertise in conducting trials on Merck compounds. Having one of the largest sets of genomic targets, deCode is establishing a leading position in genomic drug discovery and development. Early-stage targets are currently not highly valued, but if the company can continue to validate and further develop these targets, it could be one of the Þrst to capitalize on the Þrst real wave of genomics-derived drugs, anticipated in the next Þve to ten years. Exelixis Like CuraGen, Exelixis has emphasized integration of technologies. The company also has particular expertise in the area of genomics and animal models. The lead Exelixis compound—and until recently, its only drug in development—is a potential treatment for cancer (XL-119) that was licensed from Bristol-Myers Squibb (BMS) in 2001. Exelixis began a Phase III trial for this compound in 2004 to examine its effect as a potential treatment for bile duct tumors. In 2003, the company launched its Þrst clinical trial of a “home-grown,” proprietary, small-molecule anticancer compound, XL-784, based on its genomics research. The drug targets a cell surface protease that is involved in cleaving cell growth factors. That protease was discovered in the Exelixis anti-angiogenesis research program. The compound shows both anti-angiogenic and antiproliferative effects. Concurrent with the cancer trial, Exelixis is exploring the compound’s effects in various animal models of other diseases, including renal and cardiovascular disease. The company’s 2002 alliance with GSK, whose stated goal was to discover and develop novel therapeutics, was one of the year’s biggest deals. If all goes well, Exelixis could receive up to $350 million in milestone payments. It also has the advantage of having at least one compound in Phase III trials. But the little company is, like all its peers, hard-pressed to start delivering more potential drugs to the clinic. Exelixis recently announced that several preclinical projects would move forward. The company submitted investigational new drug applications (INDs) for two receptor tyrosine kinase (RTK) inhibitors (XL-647 and XL-999) in 2004 and a CHk protein kinase inhibitor (XL-844) in 2005. Exelixis lists the following targets as key areas for its preclinical research: •

KIT, an RTK that is mutated in a number of human cancers, including gastrointestinal stromal tumors, and is expressed at higher than normal levels in cancers such as small-cell lung cancer and ovarian carcinoma. EXEL9820 is the company’s lead compound active against this target.

282 •

•

•

PROSPECTS FOR GENOMICS-DERIVED DRUGS

MET, an RTK that is overexpressed in the majority of human tumors, including all the major solid tumor classes, and contributes to the growth, survival, and invasive properties of tumor cells. EXEL-2880 is the company’s lead compound active against this target. ALK, an RTK normally expressed in the developing nervous system that becomes inappropriately activated via chromosomal translocations in a subset of non-Hodgkin’s lymphoma patients. EXEL-6309 is the company’s lead compound active against this target. p70S6K, a serine-threonine kinase that controls cell growth and is at the end of a pathway that is frequently activated through mutation or gene ampliÞcation in many human tumors. EXEL-2942 is the company’s lead compound active against this target.

ExonHIT Therapeutics ExonHIT uses its DATAS (Differential Analysis of Transcripts with Alternative Splicing) gene-proÞling technology to distinguish alternative splice variants in diseased and normal tissues. Alternative splicing is the process that allows multiple proteins to be expressed by a single gene: the intermediate messages between the gene and the different proteins (i.e., the RNA) are pieced together differently. Alternative splicing has proved to be one of the major complicating factors in genomics. The old rule of “one gene, one protein” has been changed; it is now clear that one gene can code for many proteins, and most genes code for more than one. One of ExonHIT’s goals is to generate “spliceozomes”—the full repertoire of RNA-splicing isoforms—for particular central nervous system diseases and cancers. ExonHIT has two drugs in clinical trials, and several are in preclinical stages. The company is focusing on neurodegenerative diseases and cancer. ExonHIT’s Þrst clinical candidate is an established drug (EHT-201) that is being tested against a new target thought to play a role in amyotrophic lateral sclerosis (ALS). The target was identiÞed through the company’s DATAS genomics platform. The drug is being tested in a Phase II trial in Europe that includes 400 patients and 12 medical centers in four countries. In December 2003, ExonHIT launched a Phase I clinical trial with another potential antineurodegenerative, EHT-202. The company says that in pharmacological models of neuronal cell death, EHT-202 appears to play a role in protecting neurons. ExonHIT associated RNA isoforms produced by alternative splice variants with particular roles in neurodegenerative disease models. Like many of its peers, ExonHIT faces a cash crunch because R&D expenses are climbing as its compounds advance through the development pipeline. The company is trying to generate revenue from multiple avenues; for example, it markets a predictive toxicology array called Safe-Hit. ExonHIT is engaged also in several R&D collaborations with Allergan, BioM´erieux, Idec Pharmaceuticals, and Roche. The success of EHT-201 and EHT-202 is essential to the company’s continued existence.

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GlaxoSmithKline (GSK) All the major pharmaceutical companies have genomics programs and use genomics in multiple ways, but GSK has made genomic drug discovery a priority and has been a notable pioneer in this area. In 1993, what was then SmithKline Beecham signed an exclusive $120 million agreement with HGS to form the groundbreaking Human Gene Therapeutic Consortium, with the aim of identifying novel gene targets using Craig Venter’s expressed sequence tag methodology and other genomic tools. A few years later, that agreement was amended to include several other companies, but SmithKline Beecham remained the biggest pharmaceutical company in the consortium. Another genomics project was inaugurated in 1999, when GlaxoWellcome spearheaded the SNP Consortium, whose aim is to map important markers of genetic variation. Glaxo’s Allen Roses, formerly a noted academic researcher in the Þeld of genetics, initiated a pioneering effort in large-scale genotyping with the aim of Þnding the key gene targets in the genome by studying human genetic variation. Subsequently, the formation of GlaxoSmithKline by SmithKline Beecham and GlaxoWellcome brought together two organizations that were already committed to genomics. Most of GSK’s initial results in this Þeld have related to drug development. For example, the company has identiÞed genetic variants that appear to be associated with adverse reactions to its HIV drug abacavir (Ziagen) and is seeking to validate these variants. Genotyping studies are also under way to Þnd a genetic explanation for adverse reactions to alosetron hydrochloride (Lotrenex); this GSK drug for IBD was voluntarily withdrawn in November 2000 because of serious and life-threatening cases of ischemic colitis and constipation that were linked to use of the drug. In June 2002, the FDA approved a supplemental new drug application (NDA) for alosetron under a risk management program and for a narrower indication: women with severe, diarrhea-predominant IBD who have failed to respond to conventional therapy. GSK has the advantage of being able to do very large scale studies and has pioneered the whole genome scan approach, using 100,000 to 300,000 SNPs across the genome. The company also has access to well-phenotyped patient samples and has been collecting such samples regularly for genotyping. GSK has deals with several companies that offer genotyping platforms and services, including Illumina, Perlegen, and Sequenom. GSK aims to complete scans in the following disease areas: asthma, chronic obstructive pulmonary disease, rheumatoid arthritis, Alzheimer’s disease, epilepsy, osteoarthritis, schizophrenia, unipolar and bipolar depression, type II diabetes, obesity, hypertension, acute coronary syndrome, multiple sclerosis, and IBD. The drugs that result from these scans will take at least Þve to ten years to reach the market. However, the company’s approach is highly regarded in the scientiÞc community, and it is very likely that GSK will produce a large proportion of the top genomic targets in many of these disease areas. In the shorter term, GSK is pursuing one leading genomics-based target, which derived from the original HGS/SmithKline Beecham deal. In late 2003,

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GSK announced that 480848, a small-molecule inhibitor of lipoprotein-associated phospholipase A2 (Lp-PLA2 ), would be advanced to Phase III trials in 2004. LpPLA2 provides a novel means of addressing atherosclerosis (Packard CJ, 2000). The drug is one of 20 in development that GSK claims could have more than $1 billion market potential. GSK’s chances for approval of the drug may be enhanced by recent approval of a new diagnostic test that could enable better patient selection. Earlier this year, the biotechnology company diaDexus announced that the FDA has cleared its PLAC test, which measures blood levels of Lp-PLA2 , for marketing as a diagnostic aid that can help physicians gauge patients’ risk for coronary heart disease. Currently, GSK has two drugs in trials against two other HGS-generated targets. One candidate, 659032, another Lp-PLA2 inhibitor, is in Phase I trials for cardiovascular disease. Another drug, 462795, is a cathepsin K inhibitor in Phase I trials for treatment of osteoporosis and osteoarthritis. Human Genome Sciences One of the Þrst genomics companies—and by far the most aggressive—is Human Genome Sciences (HGS), which has experienced serious setbacks since its inception in 1992. The company has Þled an estimated 7500 gene-based discovery patents and approximately a dozen INDs in the last six years, but several drugs have been dropped from development. Notably, the problem with the compounds whose development was discontinued appears to be efÞcacy, not safety. The company still has many novel genomic targets and several drugs in its pipeline that target such genes—for example, LymphoStat-B is a MAb to Blymphocyte stimulator (BLyS), and LymphoRad 131 is a radioiodinated form of BLyS. HGS discovered this novel protein in 1999 and reports that it stimulates the B cells of the immune system to mature into plasma B cells, which can generate antibodies and thus constitute a critical part of the body’s defense against infections and cancer. Another novel HGS target is the TRAIL (tumor necrosis factor-related apoptosis-inducing ligand) receptor-1 (R-1), which is expressed on a number of solid tumors and tumors of hematopoietic origin. The company’s studies suggest that many tumors expressing TRAIL R-1 can be killed by apoptosis (programmed cell death) induced by binding to the TRAIL R-1 MAb. The HGS TRAIL R-1 MAb acts as an agonist to mimic the activity of native TRAIL. The company’s TRAIL R-2 agonistic antibody also induces apoptosis by inhibiting an HGS-identiÞed target. Besides searching for antibodies against its proprietary targets, the company acquired a novel albumin fusion technology that allows it to make new, potentially better versions of established protein drugs. HGS’s Albuferon is a new form of recombinant interferon-alpha, Albuleukin is a new form of interleukin-2, and Albutropin is a new form of human growth hormone. ABthrax (human MAb to Bacillus anthracis protective antigen) was developed last year by identifying a human MAb that speciÞcally recognizes and neutralizes the lethal toxins produced by B. anthracis. Unlike the preventive anthrax

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vaccine, ABthrax works immediately. It may also prevent and treat infections by antibiotic-resistant strains of anthrax. The company is planning to pursue this vaccine with U.S. government support. It has received fast-track designation and would therefore follow an expedited regulatory pathway. Over the last few years, HGS has reported two major clinical trial disappointments. In 2003, the company announced that a Phase II clinical trial of topically administered Repifermin (keratinocyte growth factor-2 (KGF-2)) in adults with chronic venous ulcers did not meet its primary end point. There were no significant differences between Repifermin and placebo on time to complete wound closure or complete wound closure by 16 or 26 weeks of treatment. In 2002, HGS announced it would discontinue development of Mirostipen (myeloid progenitor inhibitory factor (MPIF)), which was then its most advanced genomics-based drug. Mirostipen simply did not show sufÞcient efÞcacy in Phase IIa trials for chemotherapy-induced neutropenia. HGS announced in 2004 that its goal was to advance clinical trials in two main areas: immunology/infectious disease and oncology. The company has preclinical candidates in several therapeutic areas, including oncology, immunology, endocrinology/metabolism, and infectious disease. Through its various agreements, HGS has also distributed hundreds of novel drug targets to partners; the company may have to wait a while before it sees royalties from many of these alliances. Currently, GSK’s Lp-PLA2 inhibitor is the most advanced drug against an HGS-generated target; if it succeeds, this drug will launch in 2008. Scientists and pharmaceutical executives share widespread respect for the HGS approach. However, it is clear that the company’s Þrst set of targets was not ideal. Like everyone else, HGS has stepped up its functional genomics effort and the second round of targets could be substantially better. Still, the company will probably have to outlicense some later-stage projects to achieve its goal of being the Þrst company to bring an array of genomics-based drugs to market. Rigel Pharmaceuticals Rigel Pharmaceuticals was formed in 1996 around an integrated functional genomics platform that includes expertise in protein pathway elucidation via yeast and mammalian two-hybrid and proteomics technologies. The company claims that its unique functional screening approach allows it to validate targets earlier in the disease process than traditional genomic approaches. In its Þrst four years of existence, Rigel identiÞed 23 new drug targets in ten therapeutic areas. Rigel now has three drugs in trials. The company’s Þrst trial was launched in 2003 and involves a small-molecule inhibitor of mast cells (R112); mast cells produce immunoglobulin E, a protein associated with allergies and asthma. In the fall of 2003, Rigel announced it had initiated a Phase I trial of R803, an experimental drug to treat the hepatitis C virus (HCV). Rigel has stayed the course and is delivering products that keep investors interested. In 2003, the company even raised $46 million in a private placement led by MPM Capital. This Þnancing was very helpful, but the company is now

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PROSPECTS FOR GENOMICS-DERIVED DRUGS

in full-ßedged development phase and will need additional infusions of capital as well as continued progress with compounds in clinical trials. ZymoGenetics ZymoGenetics stands out among the small genomics Þrms because it has actually developed marketed products. This experience has provided ZymoGenetics both valuable expertise and cash. None of the products are genomics-based, and most were developed for collaborators before the current company was formed. Founded as Zymos in 1981, the company collaborated with Novo Nordisk to develop recombinant human insulin in 1982 and then cloned human factor VII. Renamed ZymoGenetics in 1983, it was acquired by Novo in 1988 and continued to develop novel protein therapeutics and manufacturing methods for several major protein drugs, including Novo’s factor VIIa (NovoSeven), Johnson & Johnson’s PDGF-BB (Regranex), and Eisai’s TPA analogue (Cleactor). In 1994, the company became the Þrst to clone thrombopoietin (TPO). That same year, ZymoGenetics established a bioinformatics-driven, genomics discovery research effort. In 2000, ZymoGenetics reestablished itself as an independent company at the peak of the genomics boom. ZymoGenetics’ most advanced bioinformatics-derived product candidates are TACI-Ig and IL-21 (interleukin-21). The Þrst of these is a soluble receptor with potential applications for treatment of autoimmune diseases, including systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA). TACI-IG is in joint development by ZymoGenetics and Serono and entered Phase I trials in 2003. IL21 is a novel protein that regulates natural killer cells and cytotoxic T cells; it has potential applications for treatment of cancer. IL-21 is licensed to Novo Nordisk for commercialization outside North America; ZymoGenetics retains commercialization rights within North America. In 2003, ZymoGenetics also launched a Phase I clinical trial for one of its recombinant protein drugs, rFactor XIII. OUTLOOK FOR GENOMICS-BASED THERAPEUTICS Four major questions remain about genomics in drug discovery and development: • • • •

Overall, will genomics-derived drugs move through discovery and development more quickly than traditionally derived drugs? Will any of the Þrst genomics-derived drugs succeed? Will the study of genetic variation produce a new wave of drugs? If so, when will that happen?

While most drugs based on novel genomics-derived targets are moving very slowly, often showing insufÞcient efÞcacy to justify continued development, it will soon be clear whether genomics can actually speed discovery. As pointed out earlier, HGS has Þled about a dozen INDs in just a few years, most of them for genomics-based drugs. Rigel, with just 135 employees, is on track to Þle two INDs per year, and Exelixis appears to be following suit. If these three companies

REFERENCES

287

can move that many drugs to market, genomics will indeed have speeded up the process. It is also clear that genomics holds great promise of accelerating many individual aspects of drug development. Genome-wide RNAi libraries are becoming widespread, providing invaluable tools to probe gene function and improve understanding of drug modes of action. Gene expression and protein expression signatures are increasingly used as valuable biomarkers for population selection and drug efÞcacy evaluation. Genetic variation is being used to better select patients for trials. Companies such as Bristol-Myers Squibb, Millennium Pharmaceuticals, PÞzer, Novartis, and Wyeth are using genomics strategically to accelerate development of some of their drugs. The Þrst wave of genomic targets was probably far less well characterized than the second wave will be. The improvement will be owed to developments such as RNAi and better tools for text mining (e.g., to collect literature references on targets). Therefore, we anticipate that several genomics-based drugs will emerge in the next four to six years. The number of genomics-based diagnostics and prognostic tests will increase rapidly and steadily, which will further advance the development of genomics-based drugs. But the major potential wave of genomics products will be generated by the wide-ranging genetic variation studies of companies such as Celera Genomics, DeCode, Galileo Genomics, GSK, Perlegen, and Sequenom. GSK and deCode are the most advanced in this race; both are likely to produce a signiÞcant number of major new drug targets. It will take at least eight to Þfteen years for the ensuing wave of products to reach the market, and substantial research remains to be done. The most important factor that could accelerate progress in this research would be the introduction of less expensive genotyping tools. However, good progress has been made in terms of bringing tools to maturation, Þlling gaps in technology, and advancing some reasonable drug targets. Evidence of this progress is likely to be seen within the next Þve years as drugs against genomics-derived targets begin to make their way through company pipelines. REFERENCES Kong A, et al. A high-resolution recombination map of the human genome. Nature Genetics. 2002;31:241–247. Packard CJ, et al. Lipoprotein-associated phospholipase A2 as an independent predictor of coronary heart disease. New England Journal of Medicine. 2000;343:1148–1155. Stefansson H, et al. Neuregulin 1 and susceptibility to schizophrenia. American Journal of Human Genetics. 2002;71:877–892. Stefansson H, et al. Association of neuregulin 1 with schizophrenia conÞrmed in a Scottish population. American Journal of Human Genetics. 2003;72:83–87. Weber JL. The Iceland Map. Nature Genetics. 2002;31:1–2.

Prying Open the Pipeline: Drug Discovery and Development in the Postgenomics Era

BUSINESS IMPLICATIONS •

•

•

•

In the postgenomics era, the most successful biotechnology companies will combine expertise in traditional pharmaceutical company drug development strategies with the unique capabilities that biotech companies possess in the areas of genomics, proteomics, bioinformatics, and metabonomics/metabolomics. This systems biology approach is essential for companies seeking to evolve into fully integrated Þrms that embrace a business model based on product development as opposed to platform technology alone. Large pharmaceutical companies and venture capitalists (VCs) are focusing their attention on biotech companies that have in-house development programs. Many key biotech Þrms are aware of this focus and have begun to tailor their businesses to attract these players. With the expanding application of RNA interference (RNAi) technology to drug development, we anticipate that biotech companies that already have strong intellectual property positions in this area will become increasingly attractive to pharmaceutical players. As a result, we expect increased alliance and collaborative activity. This activity will continue between U.S. and Canadian companies as well as between those in Europe and in Australia through 2006, the end of our forecast period. Government funding is increasing to enhance genomics and proteomics knowledge, with a push toward bringing diagnostics and therapeutics to

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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market. Authorities are encouraging the creation of small- and medium-size enterprises as well as initiatives supporting collaboration among industry players on innovative research projects. Bringing these companies together could lead to increased cross-border alliances and acquisition activity. INTRODUCTION In the late 1990s, as the Human Genome Project neared completion, biotechnology companies appeared to represent a new era centered on genomics, with promising research, enthusiastic management, and ambitious business plans. Industry experts believed that genomics would Þll major gaps in pharmaceutical company pipelines. However, on the whole, investors in these companies have been left short-changed, with pipelines almost universally failing to materialize as anticipated. This disappointment resulted from the industry’s failure to quickly recognize the immediate challenge of the postgenomics era of assigning biological function to all of the proteins encoded by the genome. In this environment, a crisis of conÞdence among pharmaceutical company shareholders has remained a constant. In addition, exacerbating this problem are proÞt warnings that have become the norm in successive quarters as well as the failure of new products to launch in sufÞcient numbers or to generate revenues close to expectations. Negative shareholder sentiment extends from the largest pharmaceutical companies to small- and mid-size pharmaceutical and biotech Þrms that are attempting to redeÞne their strategies. As a result, investors are now looking for biotech companies that are already successful and appear to be poised for worldwide success. In this article, we contend that the most successful biotech companies operating in the postgenomics era are likely to be those that acquire integrated systems biology capabilities and demonstrate that their businesses are modeled on product development as opposed to a technology platform. In addition, we assert the notion that application of advances in genomics and proteomics to drug development, in Europe and other regions outside the United States, will allow for increased cross-border partnering as well as merger and acquisition activity on a global basis. Table 1 shows select platform technology companies seeking collaborations or consolidations with other companies in the biotech/pharmaceutical industry. STATE OF THE PHARMACEUTICAL INDUSTRY: WHY MUST PHARMA LOOK TO BIOTECH? According to a study published by CMR International in 2000, average total development time for new compounds, from identiÞcation to launch, increased from just under eleven years in 1996 to thirteen years in 1999. With ailing pipelines and the loss of patent exclusivity looming for many agents, pharmaceutical companies are anxious to Þnd methods of launching new drugs quickly, especially drugs based on the several obvious and easy targets that have already been investigated.

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TABLE 1. Select Platform Technology Companies Seeking Consolidation and Collaborations Company

Status

Autogen (Australia)

Public (ASX listed)

Bionomics Ltd (Australia)

Public (ASX listed:BNO)

Cadus Pharmaceutical (United States)

Public (NASDAQ: KDUS)

Caprion Pharmaceuticals (Canada)

Private

CompuGen (United States)

Public (NASDAQ: CGEN)

Curagen (United States)

Public (NASDAQ: CRGN)

Partnerships • Merck-Lipha • Kyokuto • Sequenom

• Johnson & Johnson • Genmab • Hybrigen • Ozgene • Bristol-Myers Squibb • Lederle • Solvay Duphar • Trega Biosciences • IDEC Pharmaceuticals • Ortho-Clinical Diagnostics • IDEXX Laboratories

• Aventis

• • • •

Bayer Biogen Genentech GlaxoSmithKline

• Roche

Comments Targeting obesity and type II diabetes, four products in preclinical development. Ongoing programs also in identifying targets for depression and anxiety. Targeting breast cancer, epilepsy, angiogenesis; three products in preclinical development. Drug discovery utilizing yeast cells as a platform exploiting the similarities between the yeast and the human genome. Application of subcellular proteomics platform to development of diagnostics and drug targets. Caprion’s programs include diagnostic products for mad cow disease and human variant Creutzfeldt–Jacob disease (vCJD), early-stage programs in amyotrophic lateral sclerosis (ALS), and an oncology-focused collaboration with IDEC Pharmaceuticals. Software for the analysis of genomics and proteomics data; DNA chip design. Drug discovery in obesity and diabetes, cancer, and autoimmune and inflammatory diseases through a functional genomic information technology platform.

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TABLE 1. (continued) Company

Status

Deltagen (United States)

Public (NASDAQ: DGEN)

Devgen (Belgium)

Private

DeveloGen Private (Germany)

ExonHit (France)

Private

Partnerships • Merck • Pfizer • Roche • Schering-Plough

Comments

Development of new technologies for elucidating gene function. Validated targets: DT011M for treatment of diabetes and obesity; DT021I for inflammatory disease; DT031I for Crohn’s disease; CD123, a licensed marker distinguishing normal from abnormal cells in acute myelogenous leukemia (AML). • Janssen Pharmaceutica Drug and drug target discovery • Genentech through functional genomics. Proprietary technology platform based on the Caenorhabditis elegans roundworm to rapidly discover and validate specific drug targets. Programs targeting metabolic and central nervous system disorders. None known Discovery of genes as therapeutic targets in obesity, diabetes, and liver disease. Technology platform built on two areas of expertise: stem cells and phenotype-first genomics. Has a range of gene delivery technologies of both viral and nonviral origin from its merger with Hepavec (a German gene delivery company) in October 2000. • Roche Applied Science Focus on development of • Allergan therapies for cancer and • IDEC neurodegenerative diseases. • BioMerieux-Pierre Fabre Four compounds under development: EHT-0201 and EHT-0202 for ALS, and EHT-0101 and EHT-0102 for cancer. EHT-0201 in Phase II clinical trials. Entered into collaboration with Roche to develop a diagnostic test that can identify bovine spongiform encephalopathy in living animals. Also developing a series of molecular cancer diagnostic tools.

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TABLE 1. (continued) Company

Status

Genaissance Public Pharmaceu(NASDAQ: ticals GNSC) (United States)

Genfit (France) Private

Partnerships • Janssen Pharmaceutica

• • • • • • • •

GenOdyssee (France)

Private

Genome Ther- Public apeutics (NASDAQ: (United GENE) States)

Comments

Genomics and informatics to seek pharmaceutically relevant genes. Identified candidates for development in its own pipeline of products—HAP-Clozapine and HAP-Statin—utilizing its proprietary genetic markers. Also developing its own internal pipeline of products using its proprietary genetic markers. Merck-Lipha Targeting diabetes, obesity, Aventis vascular inflammation, Sanofi-Synthelabo thrombosis, lipid disorders, inflammation, asthma, and UCB Pharma CNS disorders with Biomerieux-Pierre Fabre products in varying stages Fournier of development. Has a Kowa Company bioinformatics subsidiary IBM called IT.Omics.

• Urogene

• • • •

Functional genomics company targeting development of therapeutic proteins and diagnostics. Has functional proteomics and bioinformatics expertise in-house. AstraZeneca Identification and BioMerieux characterization of human Schering-Plough and bacterial genes for the American Home Products development of pharmaceutical, vaccine, and diagnostic products. Ramoplanin is in Phase III development for the prevention of bloodstream infections caused by vancomycin-resistant enterococci (VRE). Helicobacter pylori, Staphylococcus aureus, and fungal infection products in development through alliances with major pharmaceutical companies.

STATE OF THE PHARMACEUTICAL INDUSTRY: WHY MUST PHARMA LOOK TO BIOTECH?

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TABLE 1. (continued) Company

Status

Genset (France)

Public (Nouveau Marche: GNSET; NASDAQ: GENXY)

Hybrigenics Private (France)

Partnerships

Comments

• Abbott Location and identification of genes • American Home Prod- and polymorphisms associated with diseases and drug responses. ucts Preparing Famoxin, a therapeutic • Janssen protein, for clinical trials. Target • Pharmacia indications in obesity and type II • Sanofi-Synthelabo diabetes. Serono increased its • • • • •

stake in Genset in September 2002 to close to 86%. Incyte Genomics Targeting viral diseases, cancer, and Lynx Therapeutics metabolic disorders; building a Merck Sharpe & Dohme pipeline of proprietary biomarkers, small molecules, and antibody drug Mindsense products. Oxford Glycosciences

• Servier • XTL Biopharmaceuticals Keryx Biopharmaceuticals (United States) Lexicon Genetics (United States)

Public (AIM: KRX;)

Public (NASDAQ: LEXG)

Locus Private Discovery Inc. (United States) Lynx Thera- Public peutics (NAS(United DAQ: States) LYNX) Myriad Public Genetics (NAS(United DAQ: States) MYGN)

• Novo Nordisk • Osteotech

Application of bioinformatics to protein kinase sequences to identify therapeutic peptides.

• Abgenix Knockout mouse technology to define • American Home Prod- gene function and discover ucts pharmaceutical products to treat • Boehringer Ingelheim human disease. Lead discovery/optimization stage • Bristol-Myers Squibb pipeline targeting cardiology, • Pharmacia immunology, neurology, and oncology. • Aventis Protein modeling of receptors, enzymes, and other proteins for drug discovery • BASF • DuPont • Aventis • Bayer • Eli Lilly • Pharmacia • Novartis • Roche • Schering-Plough

Discovery of gene expression patterns and genomic variations through the application of a proprietary cloning technology. Identification of disease-causing genes and development of targets through application of proprietary genetic tests for colon, endometrial, breast, and ovarian cancers, and cardiovascular disease. Drug discovery efforts target cancer, rheumatoid arthritis, acute thrombosis, Alzheimer’s disease, HIV/AIDS, and other viral diseases.

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TABLE 1. (continued) Company

Status

Proteome Sciences (United Kingdom)

Public (AIM: PRM)

Quark Biotech (United States)

Private

Syrrx (United States)

Private

Partnerships • Academic Institutes

• Fujisawa • Perkin-Elmer • Sankyo • Shionogi • Taisho • Sankyo • Hoffman-La Roche Pharmacia • Celera Genomics • Cubist Pharmaceuticals

Comments Technologies for the discovery of relevant protein markers of disease with potential as diagnostic markers and novel therapeutic targets. Functional genomics and bioinformatics applied to the identification of genes suitable as targets for drug development. High-throughput structural proteomics.

We believe that all future drug development will eventually be grounded in the biotech industry. Large pharmaceutical companies that will win in this landscape will shrewdly align themselves with biotechs that have technologies that are complementary to their own. Essentially, the major effort for pharmaceutical companies will be the inlicensing of the most promising products emerging from companies in the genomics and proteomics Þelds. In some cases, pharmaceutical companies will acquire biotech companies. In addition to plans to work with or acquire biotech companies, some pharmaceutical companies have restructured themselves internally to foster a greater level of innovation. Nonetheless, these efforts have not paid off so far. For example, GlaxoSmithKline (GSK), which has more than 100,000 employees worldwide, is now considered so large that some analysts believe that innovation and creative thinking often are crushed by bureaucracy. Like many of its peers, GSK seems to have no objective way of assessing its return on capital from research. In addition, such companies often pay large sums of money to law Þrms defending patents on old drugs; in the third quarter of 2002, GSK announced an approximately $230 million allocation for 2002 drug patent defense. In 1999, GlaxoWellcome (now GSK) had the idea to create smaller research teams modeled on biotech companies that would nurture and promote innovative research ideas. GlaxoWellcome separated its research department into six semiautonomous units with their own budgets to try to mimic the biotech “atmosphere” within its very large R&D division. However, GSK has yet to realize success in this effort, and analysts do not believe that the R&D department at GSK is

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any nearer to producing the billion-dollar blockbuster drugs that the market now demands. Note that GSK is just one of the pharmaceutical companies facing major drug development challenges today. ROLE OF VENTURE CAPITALISTS Venture capital Þrms have gained hard-earned wisdom from their own negative experiences with biotech company overvaluation. Today, venture capitalists (VCs) are looking for technology, good science, and highly experienced management. They seek products emerging from the biotech companies in their portfolios that will progress from identiÞcation and Phase I/II clinical development to licensing by pharmaceutical companies with the sales and marketing infrastructures to make the product succeed. VCs are no longer willing, and are not in a position, to offer capital based on an intriguing business plan or hot technology platform. VC investment strategies are moving toward maintenance schemes in an attempt to allow biotech companies to mature. A maintenance scheme selectively feeds funds into a biotech company, while the management gains valuable experience and the products are moved closer to market. In addition, this strategy slows down the company’s move to an initial public offering (IPO) listing until the market becomes more favorable. For the ßedgling biotech, a slower path to public Þnancing also allows the company to build solid partnerships and to mature into a Þrm that can negotiate on more conÞdent footing with potential partners. VCs in Europe found that the valuations for portfolio companies that are geared toward IPOs were not favorable through 2003; thus, portfolio management may need to tap private equity as one means of keeping promising biotech companies in business. An extension of this maintenance technique that is Þnding favor in Europe is the strategy of combining a private company with limited cash and a pipeline of products with a struggling publicly traded company that has signiÞcant cash reserves. In Germany, many of these struggling public companies have been Þnanced with grants from federal and state agencies to stimulate the biotech industry. They are now considered ripe targets for the aforementioned strategy. In the absence of a market for biotech IPOs, Þnding such companies is one of the key goals of life sciences VC teams that believe that some of these companies are not sustainable as stand-alone biotech Þrms; experts expect that such consolidation will increase. INTERDISCIPLINARY COLLABORATIONS: FINDING A FORMULA FOR SUCCESS Advances made in the genomics and proteomics Þelds have unearthed a wealth of data that are increasingly difÞcult to interpret when applied to the drug development process. A primary reason for this dilemma is that identiÞcation of lead compounds based purely on genetic data does not provide all of the information needed to conduct successful target validation studies. Namely, in

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human systems, most biological functions arise from interactions among several biomolecular components. Furthermore, data on metabolism need to be added to the expanding information base so that researchers can understand the effect that lead compounds may have on selected biological systems, both in terms of efÞcacy and toxicity. The inability of genetics and proteomics to provide a full picture of a drug candidate has limited the translation of the heavy investments that the pharmaceutical industry has made in these areas into the blockbuster drugs it had anticipated. In fact, genomics and proteomics platform technologies that are in the form of database products are now seen as low-proÞt commodities. Therefore, the highly attractive investment climate for genomics and proteomics companies has slowed, forcing companies that had once enjoyed high valuations based purely on their platform technologies to rethink their business strategies. In this postgenomics era, the most pressing issue confronting platform-technology-based biotech companies is the need for a tangible therapeutic or diagnostic product that can be tied to advances in proteomics technology. Because the failure of drugs during development is the major contributing factor to the escalating costs of bringing drugs to market, pharmaceutical companies now are working to terminate unsuccessful compounds even more quickly. An example of such a failure occurred at PÞzer in 2000 when one of the company’s promising drug candidates showed positive results in rats but was terminated because of a lack of efÞcacy during limited clinical trials. Although this termination came after an investment of $71 million, the decision to terminate saved millions of dollars in expenditure on clinical studies. Large pharmaceutical companies are now purely interested in cooperating with biotech companies possessing technologies that demonstrate target identiÞcation and validation capabilities and, better still, companies that have proved they are able to generate leads in preclinical and clinical development. As a result, biotech companies are shifting their strategies to encompass the concept of systems biology. SYSTEMS BIOLOGY The concept of systems biology involves the integration of genomics, transcriptomics, proteomics, metabonomics/metabolomics, and bioinformatics. The aim of systems biology is to act as an interface between biology and the disciplines of chemistry, information technology (IT), and medicine to allow identiÞcation of relevant targets at the earliest possible stages of the drug development process. The higher quality of targets identiÞed using this approach should allow for a smaller number of drugs reaching clinical trials, increased success in clinical trials with these drugs, and thus reduced overall costs. As a result of this expanded approach, the structure of the drug development process is changing. For example, biotech companies that have focused on genomics or proteomics platform technologies in the past will increasingly need to incorporate the other aforementioned disciplines and thus broaden their

EXPANDING BIOINFORMATION TECHNOLOGY COLLABORATIONS

297

technological basis. This development will add to the already growing number of collaborations and acquisitions between biotechs and pharmaceutical companies. With the continued expansion of data will come the need for deployment of more sophisticated hardware and software that will require a greater interplay between life sciences companies and large IT Þrms. In this context, the life sciences industry is focusing more and more on bioinformatics. As mentioned earlier, one of the key growth areas of systems biology is metabolism. Pharmaceutical companies recognize the Þelds of metabonomics and metabolomics as independent and useful techniques for evaluating the toxicity of drug candidate compounds. Metabonomics is the quantitative measurement of timed multiparametric metabolic responses of multicellular systems to pathophysiological stimuli or genetic modiÞcations. Metabolomics is the systematic, comprehensive analysis of the full complement of nonpeptidic, organic, lowmolecular-weight molecules in a particular cell, tissue, biological sample, or organism. When these techniques are applied to the drug development process, researchers can attempt to identify the target organ of any observed toxicity as well as derive the biochemical mechanism of the toxicity. They also may be able to derive new biochemically based assays for disease diagnosis. Adding metabolism data to the equation also increases researchers’ understanding of the effect of target compounds on biological and cellular pathway events, thus completing the systems biology approach to target screening. Experts now consider systems biology to be perhaps the key integrated approach to studying biological systems. EXPANDING BIOINFORMATION TECHNOLOGY COLLABORATIONS For biotech companies to shift to the systems biology paradigm, they need substantial informatics support. Each aspect of systems biology requires the use of informatics if a company intends to competitively achieve deals with pharmaceutical investors and VCs. Major IT companies such as IBM, Oracle, Hewlett-Packard, and Sun Microsystems have long recognized the substantial potential business opportunities available through supplying solutions to companies active in the life sciences sector. With the growth of the genomics, proteomics, and bioinformatics sectors as well as a move among life sciences Þrms to integrate these capabilities in-house, an even more lucrative revenue-generation opportunity has emerged. Note that the inßux of IT money into the life sciences arena has coincided with genomics businesses beginning to move away from basing their business plans on marketing subscriptions to their databases toward developing in-house drug development programs. In 2000, IBM initiated a program aimed at increasing its links with life sciences companies. With an allocated $100 million, IBM has forged partnerships and made equity investments in genomics companies that it has identiÞed through rigorous analyses as having technologies complementary to its own. In this way, it aims to demonstrate the capabilities of its hardware and storage solutions in

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the drug discovery area. Some of the companies that IBM has partnered with are MDS Proteomics, Lion Bioscience, Proteome Systems, Structural BioInformatics, and TurboWorx. An important driver of the IBM strategy is to break the monopoly that Oracle has on the life sciences informatics market. We estimate that Oracle controls approximately 75% of this market. The size of the life sciences informatics market is difÞcult to pinpoint because of the different deÞnitions of informatics that various companies use in their analyses. However, data from consulting and market intelligence Þrms suggest that the market for life sciences informatics is set to grow from approximately $10 billion in 2001 to $38 billion in 2006. The efforts of companies such as IBM to develop the bio-IT market may begin to open new opportunities for biotech companies needing to tap Þnancing. IBM has taken on the role of VC through its equity investments. However, it now seems that the company’s plan is to limit its equity investment program; its focus will be on gaining more leverage and broader coverage in the industry by working with other VCs to identify the systems biology companies that are likely to succeed. This move may reignite VC and investor conÞdence in biotech Þnancing in the genomics and proteomics Þelds. Myriad Proteomics, a forerunner in the postgenomics Þeld, demonstrates another possible emerging bio-IT trend. This privately held company was formed in March 2001 as a subsidiary of Myriad Genetics through a $185 million joint venture with Hitachi and Oracle. Under the terms of that deal, Myriad Genetics contributed technology valued at $82 million, while Hitachi and Oracle—along with Swiss investor Peter Friedli, who heads Friedli Corporate Finance in Zurich— contributed a combined $85 million in cash as well as $18 million in data storage hardware and database software products. A primary objective of the joint venture is for Myriad Proteomics to use the technologies in the mapping of the human proteome. The venture is gathering information on proteins and their interactions within human cells and compiling these data into a proprietary database (completed in 2004). Subsequently, Myriad Proteomics will license the database to biotech and pharmaceutical companies that can mine the information in efforts to develop therapeutics. This type of joint venture may be a one-time only transaction for companies such as Hitachi and Oracle; nevertheless, it challenges other hardware and software companies to follow suit. In addition, it reinforces the notion that the most successful biotech companies will adapt their technology platforms to not only attract hi-tech partnerships but also to align their businesses toward product identiÞcation and development. Another example of this bio-IT trend is the announcement in September 2002 of an agreement between IBM and IT.Omics, the bioinformatics subsidiary of French biotech company GenÞt. Under the terms of this agreement, IT.Omics will implement IBM’s DiscoveryLink data integration software as part of its life sciences platform and become the Þrst European DiscoveryLink expertise center to showcase the technology.

COLLABORATIONS AS A STRATEGY FOR STAYING ONE STEP AHEAD OF THE COMPETITION

299

RNA-INTERFERENCE-BASED THERAPEUTICS TO LEAD TO ADDITIONAL COLLABORATIONS Although collaborations occurring at a later stage foster the systems biology concept, as discussed in the previous section, early-stage technologies with a strong promise in delivering therapeutics will receive signiÞcant investment. Therapeutics based on RNA interference (RNAi) is such an area and will likely foster collaborations between biotech and pharmaceutical companies. Expectations that the RNAi Þeld will expand successfully and rapidly and that it will have utility in several therapeutic areas are widespread. Most notably, it is proving to be a possibly revolutionary development in the anticancer and antiviral Þelds. Indications are growing that VCs are increasingly investing in biotech companies possessing this expertise and that RNAi technology may hold substantial rewards for pharmaceutical company investors that wish to develop RNAi-based therapeutics. We believe that pharmaceutical company interest will increase in collaborations with biotech companies with strong intellectual property and established Caenorhabditis elegans genomewide RNAi feeding libraries that can be used as tools for high-throughput screening (HTS) and validation of drug targets. Table 2 lists the current key players in RNAi technology. Fundamental to certain functional-genomics-based HTS methods is the nematode C. elegans,and this model is the basis for current RNAi technology. Functional genomics is one of the HTS strategies that has been a focus at biotech companies, and it is now critical to the advancement of drug development programs. This technology is used to address high-throughput approaches to wholegenome or systemwide molecular genetic studies, and pharmaceutical company investment in this area is increasing. The C. elegans genome sequence was the Þrst animal genome to be sequenced in its entirety, and—in terms of the drug development process—the validity of this technology has been veriÞed through the positive testing of established drugs in C. elegans disease models. The unique properties of C. elegans facilitate the construction of disease-speciÞc in vivomodel systems that are adjusted to perform HTS on complex chemical compound libraries—therefore ideally lending itself to the development of RNAi technology. Biotech companies with this technology stand on the verge of the potentially lucrative position of serving both the needs of the pharmaceutical and agrochemical industries in terms of their R&D processes. COLLABORATIONS AS A STRATEGY FOR STAYING ONE STEP AHEAD OF THE COMPETITION Through 2006, the biotech sector will experience increased consolidation on a global basis, with greater transatlantic activity through collaborative biotech efforts between U.S. and European companies in both the biotech and pharmaceutical industries. In addition, companies in other countries that are seeking enhanced international exposure and access to major drug markets may become involved in collaborations with U.S. and European companies. Prominent among

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TABLE 2. Key Players in RNA Interference (RNAi) Therapeutics Company

Status

Comments

Alnylam Pharmaceuticals (United States)

Private

Benitec (Australia)

Private

Cenix Biosciences (Germany)

Private

Devgen (Belgium)

Private

Intradigm (United States)

Private

Alnylam Pharmaceuticals (Cambridge, Massachusetts) is an emerging leader in RNAi-based therapeutics and has raised $17 million in series A and B private equity financing. The Cambridge, Massachusetts-based company was founded in 2002 by Polaris Venture Partners, Cardinal Partners, and an international group of scientists that discovered key aspects of the novel biological phenomenon of RNAi. Australian firm Benitec has patented a method that uses DNA constructs to induce RNAi. Benitec claims that its technology platform can be used for high-throughput target validation, functional analysis of anonymous genes, and development of RNAi-based therapeutics. Cenix BioScience is a pioneer in RNAi-based technologies and therapeutics. In September 2002, it announced that it had raised $4.9 million in an international financing round to advance its novel RNAi drug development programs. Cenix’s main offerings for collaborative research projects include rapid genomewide RNAi screens for drug target discovery and high throughput RNAi-based target validation. Cenix is expanding its in-house product development programs for RNAi-based therapeutics, addressing major disorders such as cancer, infectious diseases, and inflammation. Devgen is developing its own in-house drug development programs. Devgen entered into a collaboration with Genentech to validate the function of novel drug targets. Devgen will use its proprietary C. elegans in vivobased RNAi technology. Intradigm presented scientific results at the American Chemical Society meeting in Boston in August 2002 that claimed success in internal programs demonstrating RNAi’s ability to silence endogenous genes, downregulate encoded proteins, and inhibit tumor growth in tumor-bearing animal models. Intradigm was created in June 2001, with Novartis Venture Fund as one of the initial investors.

these may be Australian biotech companies that have cutting-edge technologies nurtured by a range of government-led initiatives and a highly skilled workforce. Amrad, a biotech company based in Melbourne, Australia, is one such company. Amrad’s core business is the creation of innovative drugs to treat chronic severe pain, infertility, cardiovascular disease, neuromuscular disease, and stroke.

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The company reports that it has Þve projects in various stages of preclinical and clinical development. Amrad’s business strategy is to move the products of its R&D efforts through clinical trials and to the market using its own resources as well as collaborations with international pharmaceutical companies. At this time, Amrad has alliances with Serono, Gencell, Edwards LifeSciences, Cambridge Antibody Technology, and Chugai Pharmaceuticals. In June 2002, the company reportedly had $34 million in cash and investments, which will allow it to fund two to three more years of R&D activity, according to its managing director, Dr. Sandra N. Webb. U.K. company Shire Pharmaceuticals is an example of a company that has already found success in biotech cross-border collaborations. Founded in 1986, Shire has grown into a leading specialty pharmaceutical company, increasing its value from about $24.5 million in 1986 to $8.2 billion today. The company’s core strategy is built on R&D combined with strategic inlicensing of biopharmaceuticals. Management focuses on what it considers to be the eight key pharmaceutical markets: the United States, Canada, the United Kingdom, Italy, France, Germany, Spain, and Japan. Shire is now the third largest pharmaceutical company in the United Kingdom, and its drug for treating attention deÞcit and hyperactivity disorder (mixed salts of a single entity amphetamine product; Adderall XR) is a market leader in the United States. Rolf Stahel, the architect of Shire’s growth since 1994, oversaw the evolution of the company through shrewd merger and acquisition activity. The Þrst acquisition was of Imperial Pharmaceutical Services in 1995, and the last under Stahel’s stewardship was the 2001 merger of Shire and the Canadian pharmaceutical company Biochem Pharma. Shire’s strategy in merging with Biochem Pharma was to further its vision to be a leading specialty pharmaceutical Þrm by adding that company’s biotech expertise and diverse product portfolio to its business. This portfolio included the HIV/AIDS treatment 3TC (lamivudine, abacavir, tenofovir). The merger also allowed Shire to enter the growing market of vaccines and biologics. In November 2002, Shire announced that Stahel would leave by mutual consent so that the company could bring in a new CEO “better prepared to take the reins and grow the company to enter the top tier” in the increasingly competitive biotech and biopharmaceutical sector. This move was aggressive at a company that posted sales in the third quarter of 2002 of $754 million, reßecting impressive growth over the third quarter of 2001 of 23%. In March, Shire announced that Matthew Emmens, who formally worked for Merck, would succeed Stahel. ESCALATING ACTIVITY IN THE EUROPEAN BIOTECH INDUSTRY In light of the trend toward revised business models within genomics and proteomics companies, we believe that interest and activity in these areas are likely to be heightened in Europe because the number of European companies entering these Þelds will continue to expand through 2006. Driving this notion is the commencement of the European Union’s 6th Framework Program, which holds

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as one of its key objectives the enhancement of competitiveness in the European biotech sector. This program clearly emphasizes granting funds to projects that use the knowledge gained in genomics and proteomics research and that apply this knowledge to the development of drug or diagnostic products. Additionally, the European Union (EU) budget of $2.4 billion for life sciences, genomics, and biotech is meant to nurture small- and mid-size enterprises in the region and to establish biotech centers of excellence across the EU. As a result of this funding, through 2006, an increased number of proteomics companies will emerge across Europe. This funding will also lead to speciÞc research projects in the EU that focus on small molecular entities (SMEs). Such projects may take the form of either cooperative research projects at companies examining several SMEs in therapeutic areas of common interest or collective research projects carried out for industrial associations or industry sectors where SME is prominent. In addition, individual member countries and their speciÞc programs are contributing to the pan-European effort that the EU has initiated. For example, in France, support from national and regional authorities has allowed the creation of eight “Genopoles” created to nurture technology transfer between academic centers and companies focusing solely on the Þeld of genomics and postgenomics. These Genopoles are situated in Evry-Ile de France (Paris region), Lille (Nord Pas de Calais), Strasbourg (BioValley), Lyon and Grenoble (Rh oˆ ne-Alpes), Marseille, Montpellier, Toulouse, and Bordeaux. Also, in 2001, the French Ministry for Research launched the GenHomme Programme, which further seeks to nurture the collaboration between public institutions and private companies and to support ongoing genomics and post-genomics research with an allocated $270 million through 2006. Additionally, the Ministry for Research and the Ministry for the Economy, Finance and Industry allocated $26.5 million in 2001 for Þve priority research themes: bioinformatics, nanobiotechnologies, technologies for advancing postgenomics R&D, cancer, and gene and cell therapies. OUTLOOK Increased government funding is enhancing the Þelds of genomics and proteomics knowledge, and thus contributing to the advancement of new diagnostics and therapeutics toward successful marketing. Authorities are encouraging the creation of small- and mid-size enterprises as well as initiatives supporting collaborations among industry players on innovative research projects. Bringing these companies together will lead to increased cross-border alliances and acquisition activity. Genomics/proteomics companies that have developed integrated technology capabilities and target development programs are attractive merger and acquisition targets. Table 1 is not a complete list of such companies but shows some of the leading integrated platform companies that are likely to consolidate or collaborate with one another in the next few years. In the longterm, we also expect to see a greater number of pharma/biotech collaborations. The biotech companies that are likely to be winners in terms of product development will best combine the molecular diagnostic and drug therapy applications

OUTLOOK

303

of their businesses. The development of molecular diagnostics is an attractive option for these companies because it allows for more rapid generation of products than the generation of therapeutic products, bringing in much needed revenues that beneÞt the bottom line and can be used to Þnance early-stage clinical programs.

Glycomics: The Impact of Carbohydrates on Biological Function and the Implications for Drug Discovery

SUMMARY Glycomics, the study of carbohydrate biology, has largely been neglected as a source of new drugs, even though carbohydrates play important roles in a wide range of physiological processes and diseases. Carbohydrates are extremely difÞcult to analyze and synthesize; lacking a technology up to the task, few companies have made signiÞcant efforts and many have abandoned them. Now, however, new technologies have emerged that simplify and accelerate carbohydrate analysis and synthesis. The result is renewed interest in carbohydrate-based drug discovery and development in pharmaceutical and biotechnology companies. The potential market for glycomics-derived drugs is large, and several companies already have drugs in development for various carbohydrate-related disorders. In this article, we provide an overview of glycomics, describe its applications in drug discovery and therapy, and highlight the activities of companies active in the Þeld. BUSINESS IMPLICATIONS •

Glycomics, the study of carbohydrate biology (glycobiology) and its application to drug discovery and development, is an emerging discipline that is garnering increasing interest. The potential market for glycomics-derived drugs is large: four core applications—atherosclerosis, inßammation/infection, cancer, and coagulation—generate billions of dollars annually in current

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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INTRODUCTION

•

•

•

305

drug sales, and carbohydrate-based therapies have applications in most other therapeutic markets as well. Drug companies have mostly overlooked glycobiology-based drug discovery, and the market remains untapped. Many glycomics start-ups have failed, and few pharmaceutical companies devote signiÞcant resources to the Þeld. Now, recent advances in technologies that simplify and speed carbohydrate synthesis and analysis are fueling new interest in glycobiology, but additional investment and technological breakthroughs are necessary to realize the potential of glycomics. Many biotechnology Þrms have active glycomics programs. Several companies have various drugs in development for treating a wide range of carbohydrate-related diseases and disorders. Some Þrms, including Momenta Pharmaceuticals and Glycominds, are developing novel technology platforms that can be used in glycomics research and drug discovery. Companies that have ßexible and proprietary technologies, particularly for rapid analysis and synthesis, will initially take the lead in glycomics, reaching their zenith by 2006–2007. They will yield the lead to drug developers with specialized platform technologies, including tool companies reinvented as generators of drug leads. In the second half of the decade, large pharmaceutical companies will team with or acquire these small innovators, and strategic partnering will ßourish.

INTRODUCTION Carbohydrates (sugar molecules) play important roles in a wide variety of physiological processes, many of them central to a spectrum of diseases. Carbohydrates can combine with proteins and lipids to form glycoconjugate molecules—glycoproteins and glycolipids, respectively—that modulate cellular structure and function and mediate intercellular communication and cell signaling. These diverse molecules are implicated in many medical conditions, including inßammation, infection, cardiovascular disease, cancer, and lysosomal storage disorders. Because of their central roles in processes related to disease, carbohydratebased molecules and their interactions are potential targets for therapeutic intervention. Several carbohydrate-based therapies have been commercialized. The anticoagulant heparin (and its derivatives) is an example of a very successful carbohydrate-based therapy that has been on the market for decades. Another example is the Hib vaccine, a glycoprotein that induces an immune response against the meningitis-causing pathogen Haemophilus inßuenzae type b. Despite the potential of carbohydrate-based therapeutics, however, drug companies have mostly overlooked glycobiology (the study of carbohydrate biology) and its application to drug discovery. Many biotechnology companies that have attempted to harness this technology have failed, and few pharmaceutical companies devote signiÞcant resources to concentrated glycobiology programs. An important reason for lack of activity in this Þeld is that carbohydrates are extremely difÞcult to synthesize and characterize. Until recently, few technologies were available

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for easy generation or modiÞcation of glycoconjugate structures. Therefore, the carbohydrate-based drug market remains relatively untapped—but this situation may not last much longer. Recent advances in technologies that simplify and speed carbohydrate synthesis and analysis are fueling new interest in glycobiology and drug discovery. Many scientists in academia and industry are stepping up efforts to unravel the complexities of carbohydrates and their applications in drug discovery and design. In this article, we provide an overview of glycobiology, also called “glycomics,” describe its applications in drug discovery and therapy, and highlight the activities of companies active in the Þeld. THE GENOME, THE PROTEOME, AND THE GLYCOME Now that the human genome has been sequenced, scientists are working hard to understand the functions of individual DNA sequences. Of primary importance to researchers are the genes, which encode proteins, and regulatory sequences, which modulate and control gene expression. Research efforts are also increasingly focused on deciphering the proteome, the entire set of proteins expressed by the genome. Proteins, not genes, are the actual mediators of biological function; therefore, these molecules are more immediately relevant to basic biological studies and to drug discovery. The proteome is much larger and far more diverse than the genome. Several factors contribute to the proteome’s staggering complexity. A plethora of conditions (e.g., cell or tissue type, age, disease, nutritional and metabolic states, environmental stimuli) can inßuence the level of gene expression—and subsequently protein levels—in an organism or cell by turning genes on or off. In addition, many single genes can express multiple proteins of differing length and composition; in some cases, these proteins are completely unrelated to one another. Proteins are also more structurally complex than genes: genes are linear structures that consist of four building blocks (nucleotides); proteins have three-dimensional structures and consist of 20 building blocks (amino acids). In higher organisms, the proteome has yet another level of complexity, conferred by post-translational modiÞcations of newly synthesized proteins. One type of modiÞcation is the enzyme-catalyzed attachment of carbohydrates—a process called glycosylation—to nascent or just-completed proteins in the endoplasmic reticulum (a compartment within the cell). Carbohydrates have more than 30 building blocks (monosaccharides; see Figure 1). The nine monosaccharides most commonly found in humans could theoretically be assembled into more than 15 million possible tetrasaccharides (carbohydrates built from four sugar units)—and carbohydrates can form chains containing many more than four units. These carbohydrate chains may be branched, a feature that adds even more to the huge variety of “ready-to-use” proteins. Furthermore, carbohydrate chains may be attached at many points on a protein and may have signiÞcant structural diversity. Researchers have recently coined the term glycome to describe the totality of all glycoconjugates and pure carbohydrates with structural or storage functions. It

GLYCOSYLATION

307

FIGURE 1. Examples of carbohydrate structures.

is this enormously complex and diverse glycome, not just the underlying genome or proteome, that truly deÞnes an organism. Therefore, comprehensive studies of the glycome will provide researchers with new insight into the physiological processes that lead to disease and may ultimately yield new therapeutic approaches and drugs. GLYCOSYLATION Carbohydrates do not modify protein chains at random; proteins have speciÞc glycosylation sites to which the correct carbohydrate chains are attached. Various glycosylation enzymes carry out these attachments in the endoplasmic reticulum and the Golgi apparatus (both are specialized subcellular compartments). Linkage points consist of either a nitrogen atom in an asparagine residue (N-glycosylation) or an oxygen in a serine or threonine residue (O-glycosylation). Some proteins are also glycosylated at their carboxy-terminal end (terminal glycosylation). In most mammals, including humans, carbohydrate side chains of biologically active glycosylated proteins usually carry a “terminal cap“ of sialic acid, a speciÞc carbohydrate that is essential for several glycoprotein functions.

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A protein may have multiple glycosylation sites. In principle, researchers can predict the location of these sites from a gene’s nucleotide sequence. However, the nature and length of the carbohydrate chain can be unraveled only by using direct experimental methods. As mentioned previously, scientists have only recently developed rapid, efÞcient methods to perform these analyses (see “The Tools of Glycomics”). Glycosylation bestows various functions on a protein. Carbohydrates serve as tags that help guide proteins to their cellular destinations. In the Golgi apparatus, glycoproteins are sorted according to their attached carbohydrate labels and transported to various locations in the cell. Carbohydrate chains also modulate the folding of the protein and its assembly into three-dimensional tertiary and quaternary structures. Glycoconjugates that reside on the cell surface mediate many important molecular interactions (such as the interaction depicted in Figure 2). Thus, glycosylation can enable a protein or cell to interact with other proteins or other intra- or extracellular targets; aberrant glycosylation can disrupt these interactions. Because incorrectly glycosylated proteins may be unable to perform their biological functions, disruptions in the normal glycosylation processes may alter or subvert the functions of many proteins and, potentially, lead to disease. Correct glycosylation is also an important factor in the manufacture of therapeutic glycoproteins. Many recombinant human proteins currently marketed or in

FIGURE 2. Schematic of a glycoconjugate-mediated cell-surface interaction.

APPLICATIONS OF GLYCOMICS IN DRUG DISCOVERY

309

development require correct glycosylation for full biological function. Incorrectly or incompletely glycosylated therapeutic proteins have a much shorter plasma half-life and tend to be immunogenic; both factors greatly reduce the drug’s bioavailability at the intended site of action. Recombinant protein molecules produced in bacterial expression systems (which are unable to perform any type of glycosylation) or in yeast expression systems (which perform glycosylation differently than human cells) may have little or no biological activity. To increase the activity of recombinant therapeutic glycoproteins, such as erythropoietin, biopharmaceutical companies produce these molecules in mammalian cells (such as Chinese hamster ovary (CHO) cells) that can carry out glycosylation. These expression systems are very expensive, frequently inefÞcient, and difÞcult to control. Less expensive and more efÞcient means of ensuring appropriate glycosylation of therapeutic proteins would help lower the cost of many biopharmaceuticals and help manufacturers increase their production capacities. Improved production methods for protein therapeutics provide companies with commercial opportunities. GlycoFi (Lebanon, New Hampshire) is a process development and contract service company that specializes in optimizing the industrial production of glycosylated proteins with biological activity. APPLICATIONS OF GLYCOMICS IN DRUG DISCOVERY The role of carbohydrates in health and disease is both ubiquitous and complicated. Glycomics researchers are only beginning to understand the implications and the mechanisms of the glycoconjugates. A particularly fascinating focus of this research is carbohydrate-bearing adhesion molecules, which are responsible for crucial recognition and interaction events between cells as well as between the extracellular matrix and the cytoplasmic compartment. In the following sections, we brießy describe the Þelds in which glycomics is most likely to identify, deÞne, and exploit new drug targets and therapies (see Table 1). Cancer Malignancy of a solid tumor is determined by three major factors: loss of inhibition of cell division on contact with surrounding tissue, ability to invade surrounding tissue in bulk and/or by segregation of cells that can seed metastases, and ability to support its own rapidly dividing and metabolically highly active tissue by neovascularization (angiogenesis). All three factors depend on the functions of certain transmembrane glycoproteins, such as the selectins and integrins, which are involved in the initial stages of cell adhesion and recruitment during cancer metastasis; inhibition of these cell-surface molecules could potentially prevent tumor metastasis. The search for selectin inhibitors is therefore a very active area of glycomics-based cancer research. Companies are also using glycobiology to develop carbohydrate-based cancer vaccines. These drugs work by stimulating the immune system to recognize certain glycoconjugates displayed on the surfaces of tumor cells. Examples of this

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type of vaccine include Optimer Pharmaceuticals’ Globo-H development candidate (see “Corporate Highlights”) and Biomira’s (Edmonton, Alberta) Theratope vaccine, now in Phase III clinical trials for metastatic breast cancer and Phase II trials for colorectal cancer. Inflammation A key event in the complex inßammatory response is the inÞltration of white blood cells into the affected tissue at the site of an injury. Like the metastatic processes described previously, the inßammatory response is mediated by transmembrane selectins. Certain selectins expressed on the endothelial cells that line blood vessels adhere to sialyl Lewis x (sLex), a carbohydrate structure displayed on the surface of circulating leukocytes (white blood cells). The captured leukocytes slow down, may roll along the endothelial wall, and then enter the affected tissue. Compounds that interfere with the interaction between the sLex structure and speciÞc selectins (particularly the P-selectin) may be useful as anti-inßammatory agents. Potential applications for glycomics-driven drug development in this area include rheumatoid arthritis, inßammatory bowel conditions (such as Crohn’s disease), and vascular reperfusion injury after cardiac or cerebral ischemia. A good example of the anti-inßammatory potential of carbohydrate drugs is deligoparin (OP-2000), a subcutaneously administered ultra-low-molecularweight heparin. Utilizing Elan’s Medipad drug delivery system, Incara Pharmaceuticals (Research Triangle Park, North Carolina) developed deligoparin for the treatment of inßammatory bowel disease. In January 2001, Elan and Incara commenced enrollment for a six-week, pivotal Phase II/III study of deligoparin in patients who had developed symptoms of active ulcerative colitis and were

TABLE 1. Selected Glycomics-Based Therapeutic Areas and Applications Therapeutic Area Cancer Inflammation Infection

Coagulation Aging and diabetes Congenital disorders of glycosylation

Transplantation Fertility

Applications/Approaches Glycoprotein inhibitors, selectin inhibitors Vaccines (e.g., Globo-H, Theratope) Glycoprotein inhibitors, selectin inhibitors Vaccines, carbohydrate decoys Glycoprotein-processing enzyme inhibitors (e.g., iminosugars) Glycoprotein inhibitors, heparin mimic Inhibitors of advanced end-product formation Enzyme replacement therapies (e.g., glucocerebrosidase, agalsidase alpha, agalsidase beta) Carbohydrate-processing inhibitors (e.g., miglustat) Inhibitors of glycoprotein antigen recognition interactions Modulators of glycoprotein interactions

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311

receiving standard treatment with aminosalicylates. In September 2002, Incara announced that it would discontinue development of deligoparin because the trial, which had enrolled 138 patients, had failed to meet its predeÞned end points; however, the clinical end points had called for an extensive partial or even complete remission of ulcerative colitis symptoms—a criterion that no agent for this condition has met so far. If less ambitious—but clinically relevant—end points had been chosen, deligoparin might now be regarded as a good candidate for the treatment of colitis symptoms. Infection Glycoprotein cellular adhesion factors trigger and control the docking of pathogens to target cells and their subsequent invasion of those cells. Many pathogens, including both bacteria and viruses, recognize speciÞc carbohydrate structures displayed on their target cells prior to infection: for instance, bacterial infection requires binding to carbohydrates on lectins on mucosal surfaces, and the inßuenza virus recognizes sialic acid on cell surfaces. Researchers may be able to use carbohydrate decoys that inhibit this adhesion as the basis for developing novel, potent anti-infective therapies. Researchers are also experimenting with a new class of drugs called “iminosugars” to develop drugs effective against hepatitis B and C infections. Iminosugars are synthetic, small-molecule variants of naturally occurring sugars. United Therapeutics (Silver Spring, Maryland) is developing a compound, UT231-B, that can interfere with glycoprotein-processing enzymes needed for the assembly of hepatitis C virus particles in the host cell. Such interference may frustrate the virus’s ability to infect other cells. UT-231-B is in Phase I clinical trials for the potential treatment of hepatitis C infections. Coagulation Researchers have long applied carbohydrate-based research to the development of drugs that prevent thrombosis (blood clotting) and platelet adhesion to vessel walls. Thrombosis occurs when thrombocytes (platelets) aggregate through the action of their activated surface receptors (glycoprotein IIb/IIIa). Various processed and fractionated versions of heparin (a natural sulfated glycoprotein) prevent this aggregation: heparin constituents with more than 18 saccharide units can inhibit thrombin, an enzyme that cleaves Þbrinogen and leads to coagulation; smaller heparin fragments, with fewer saccharides, can catalyze the inhibition of activated factor X (a key protein in the blood-clotting process). Many companies are applying glycobiology to the discovery of novel anticoagulants and drugs that can compete with low-molecular-weight heparins. Aging and Diabetes Glycation is the nonenzymatic glycosylation of proteins, lipids, and nucleotides. The effects of glycation, which is a normal process, build up over time. As

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an organism ages, glycated proteins in blood vessel walls become increasingly cross-linked by advanced glycation end products (AGEs). These cross-linkages result in a thickening of the intima (the inner coating of blood vessels), a higher susceptibility to atherosclerotic changes, and a diminished vascular responsiveness to blood pressure changes. In diabetics, similar events take place—but much faster—when uncontrolled diabetes chronically elevates blood glucose levels. When the basal membranes of the kidney’s glomeruli (the Þltrating cells) are thickened by AGE formation, the Þltration process is impaired, leading to proteinuria and eventually end-stage renal disease. As the overall population ages and as the prevalence of diabetes increases, the inhibition or even partial reversal of AGE formation becomes an increasingly interesting objective. The receptor for AGE (RAGE) might have broader importance. In addition to binding AGEs, this cell surface receptor binds diverse other natural ligands, including amyloid Þbrils (implicated in Alzheimer’s disease) and mediators of inßammation. RAGE might therefore represent a therapeutic target for a group of seemingly unrelated disorders. Congenital Disorders of Glycosylation Congenital disorders of glycosylation affect relatively small numbers of patients; these patients exhibit variable psychomotor symptoms, mental retardation, and metabolic irregularities. Most of these disease indications represent an unmet medical need. Therapeutics developed for these diseases would likely qualify for orphan drug status in the United States and Europe. A prominent example of such a disorder is Gaucher’s disease, an inherited lysosomal storage disorder that results from the accumulation of incompletely degraded glucosylceramide, a type of fat. People with Gaucher’s disease have one or more mutations in the gene that encodes glucocerebrosidase, the enzyme that breaks down glucosylceramide. Prevalence of the disorder in the general population is approximately 1 per 100,000; prevalence is much higher in Ashkenazi Jews. Another example is Fabry’s disease, which afßicts approximately 5000 people worldwide; this disease is caused by a lack of alpha galactosidase A (also known as ceramidetrihexosidase) that results in a buildup of the fatty substance globotriasylceramide in the body’s blood vessels. Despite the tiny treatment populations, congenital disorders of glycosylation are a potentially lucrative market, partly because treatments are very expensive. Genzyme claims that approximately 3400 Gaucher patients worldwide are treated with Cerezyme (imiglucerase for injection), the company’s recombinant glucocerebrosidase enzyme product; Genzyme reported $619 million in Cerezyme sales for 2002. The market for Fabry’s disease could also be signiÞcant: the cost of treatment per patient per year is approximately $160,000–165,000. Two companies already have products that address the Fabry’s disease market: Genzyme’s agalsidase beta (Fabrazyme) and Transkaryotic Therapies’ alsidase alpha (Replagal). Both drugs are approved in Europe; Transkaryotic Therapies’ product claims about 60% of this market. In April 2003, Genzyme received marketing approval for its drug in the United States.

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Transplantation Rejection The histocompatibility antigens that allow the mammalian body to discriminate between “self” and “nonself” are membrane-bound glycoproteins that extend carbohydrate chains to the outside of tissue cells. The ABO blood group antigens, which determine the compatibility of transfused blood, belong to this category of glycoproteins. Recognition interactions between these glycoprotein antigens and certain immune cells (T cells and natural killer cells) lead to the cellular immunological response that initiates the body’s rejection of transplanted organs and tissues. These interactions are potential targets for the development of drugs aimed at suppressing transplant rejection. Fertility Glycoprotein interactions are involved in the processes of sperm cell maturation and survival, egg fertilization, migration of the zygote through the fallopian tubes, and embryo implantation and maintenance. A better understanding of these interactions could result in completely new approaches to treating certain forms of infertility.

THE TOOLS OF GLYCOMICS Drug developers have historically neglected glycobiology in their quest to discover novel therapeutics because they lacked effective tools to decipher, synthesize, and modify highly complex carbohydrates. Compared with the speed and convenience that characterize modern high-throughput technologies for sequencing or synthesizing nucleic acids or peptides, the analogous carbohydrate technologies are slow and tedious to use. Only recently have scientists begun to develop technologies for rapid sequencing and synthesis of carbohydrates. In the 1990s, Dr. Ram Sasisekharan and his colleagues at the Massachusetts Institute of Technology (MIT) developed the Þrst practical methods for rapid sequencing of complex carbohydrates. In 2001, Sasisekharan—with MIT colleagues Ganesh Venkataraman and Robert Lugar and Polaris Venture Partners—founded Momenta Pharmaceuticals (Cambridge, Massachusetts) to further develop and commercialize their new high-throughput analytical technology. In February 2001, Peter H. Seeberger’s team at MIT demonstrated the Þrst automated equipment for solid-phase carbohydrate synthesis. Optimer Pharmaceuticals (San Diego, California) has licensed a competing glycan synthesis method, Optimer Programmed One-Pot Synthesis (OPopS), from the Scripps Research Institute (see “Corporate Highlights”). These emerging technologies are generating enthusiasm for carbohydrate-based drug discovery. However, scientists still lack automated tools for determining interactions between carbohydrates and other types of biological targets. In the following sections, we describe some of the technologies and tools that researchers use for glycomics studies.

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Analysis and Modeling To analyze a glycoprotein’s carbohydrate attachments, researchers must Þrst isolate the glycoprotein. Lectins can be used to isolate glycoproteins because these molecules bind selectively to certain classes of carbohydrates; lectins that are selective for a particular class of carbohydrates can be used to isolate glycoproteins that contain members of that class. For instance, galectins are speciÞc for galactose-containing carbohydrates and may be used to isolate glycoproteins that contain galactose structures. Lectins can be coupled to solid supports and used in various protein separation and analytic methods, including afÞnity chromatography, enzyme immunoassays, and surface plasmon resonance. Once a glycoprotein is isolated, determination of its glycosylation structures can be broken down into two sequential tasks: determining the sites of carbohydrate attachment to the protein backbone and determining the composition and branched structure of the attached carbohydrate chains. The latter task is particularly challenging because determining a linear primary sequence is not enough: every branch of the forked, tree-like structure must be investigated. The technique developed by Sasisekharan and colleagues for determining glycoprotein structure uses an iterative approach. Once the size and gross sugar composition of a carbohydrate structure has been determined, a computer algorithm generates a structural “master space” consisting of all chemically possible sequences and branch structures. To narrow the number of possible sequences, researchers factor in experimental evidence obtained through a combination of chemical and enzymatic degradation and mass spectrometry. The cycle is repeated until the possibilities have been narrowed to a single unique sequence (Venkataraman G, 1999). With respect to automation and speed in glycoprotein structure determination, combinations of capillary electrophoresis and nuclear magnetic resonance (NMR) spectroscopy may offer the best prospects (Guerrini M, 2002). Researchers also use various types of mass spectrometry to characterize these structures (Dell A and Morris HR, 2001). Synthesis and Design Most automated synthesis methods currently used for oligosaccharides resemble the solid-phase synthesis methods that researchers have long used for peptides. At present, oligosaccharide synthesis methods are limited to producing carbohydrates that contain approximately 12 sugar units, a process that requires 12–18 hours. These short sugar chains can be combined by sequential use of annealing and branching enzymes. Clearly, the next step in development of higher-throughput synthesis methods is combinatorial carbohydrate synthesis, using methods similar to those used to make peptides and organic small molecules. The rational design of complex carbohydrates that have biological functions of therapeutic interest is now becoming possible. In January 2003, Sasisekharan and colleagues reported that they had designed and synthesized low-molecularweight heparins that show increased anticoagulant activity and several other

PUBLIC INITIATIVES AND INTERNET RESOURCES

315

improvements (Sundaram M, 2003). Hopefully, additional research in this area will accelerate the ability of researchers to improve existing therapies and design novel therapeutics.

PUBLIC INITIATIVES AND INTERNET RESOURCES The Consortium for Functional Glycomics (CFG) is a research project that aims to elucidate the interactions and mechanisms underlying carbohydrate-mediated cellular communication (see Table 2 for a list of public glycomics initiatives and resources). The consortium includes scientists working in the Þeld of glycobiology and is funded by the National Institute of General Medical Sciences (part of the National Institutes of Health (NIH), U.S. Department of Health and Human Services). The bioinformatics element of the CFG is developing several complex relational databases that will eventually become useful tools for glycobiology research. Most chemical and biochemical academic societies around the world are putting more emphasis on glycobiology, historically a sideline research area. Several academic organizations have special interest groups for scientists involved in carbohydrate research—for example, both the American Chemical Society and

TABLE 2. Select Public Glycomics Initiatives and Resources Initiative/Resource

Sponsoring Organization

Web Site

American Chemical Society (ACS), Division of Carbohydrate Chemistry Center of Biological Sequence Analysis (N-linked glycosylation) Center of Biological Sequence Analysis (O-linked glycosylation) Consortium for Functional Glycomics Royal Society of Chemistry (RSC), Carbohydrate Group United Kingdom Carbohydrate Chemistry Network

ACS

membership.acs.org/C/CARB/

Technical University of Denmark

www.cbs.dtu.dk/services/NetNGlyc

Technical University of Denmark

www.cbs.dtu.dk/services/NetOGlyc

National Institute of General Medical Sciences RSC

http://web.mit.edu/glycomics/consortium

Engineering and Physical Sciences Research Council

www.uea.ac.uk/che/UKCCN/welcome.htm

www.rsc.org/lap/rsccom/dab/perk002.htm

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the Royal Society of Chemistry have divisions dedicated to glycomics researchers, and the United Kingdom Carbohydrate Chemistry Network connects scientists in various disciplines who have an interest in glycobiology. Two Web-based services of the Center of Biological Sequence Analysis (CBS) at the Technical University of Denmark provide software tools that produce neural-network-generated predictions of mucin-type O-linked and N-linked glycosylation sites in mammalian proteins. These services are accessible over the Internet without charge. CORPORATE HIGHLIGHTS Many companies, particularly biotechnology Þrms, have active glycomics programs, and several companies have carbohydrate-based drugs in development for a range of diseases and disorders. Other Þrms offer various tools and technologies for glycomics research. In this section, we proÞle select companies that engage in glycomics-based drug discovery and development or provide research tools for the industry (Table 3). BioMarin Pharmaceutical BioMarin Pharmaceutical (Novato, California), a privately held company, is developing enzyme replacement therapies for several diseases, particularly lysosomal storage disorders. The company’s lead compound is laronidase (recombinant alpha-L-iduronidase; Aldurazyme), an enzyme replacement therapy for the treatment of mucopolysaccharidosis I (MPS I). This lysosomal storage disorder, known as Hurler’s syndrome, causes an array of progressive and debilitating symptoms that lead to premature death. BioMarin is developing its product in a 50/50 joint venture with Genzyme General. On April 30, 2003, the FDA granted marketing approval to laronidase for treating patients with the Hurler and Hurler–Scheie forms of MPS 1 and for treating Scheie patients with moderate to severe symptoms. The therapy has been granted orphan drug status in the United States. In June 2003, laronidase was approved for treating MPS I in 15 countries of the European Union. BioMarin is also developing recombinant human arylsulfatase B (Aryplase), an enzyme replacement for mucopolysaccharidosis VI (Maroteaux–Lamy syndrome). This candidate product has gained fast-track and orphan drug status from the FDA and the European Agency for the Evaluation of Medicinal Products (EMEA). BioMarin began evaluating the drug in Phase III clinical trials in the third quarter of 2003. In October 2001, BioMarin acquired IBEX Technologies’ pharmaceutical assets, including that company’s lead product, heparinase I (Neutralase). This compound cleaves the natural anticoagulant heparin. In September 2002, BioMarin initiated Phase III trials for the drug’s use in coronary bypass surgery. The company has contracted with Diosynth RTP, a subsidiary of Akzo Nobel, to manufacture the product.

CORPORATE HIGHLIGHTS

317

GlycoDesign GlycoDesign (Toronto, Ontario) is one of many highly specialized biopharmaceutical companies in Canada. Like so many other innovative pharmaceutical start-ups, the company restructured in 2002 by adopting budgetary restraints and downsizing its workforce to reduce its cash burn rate. In April 2003, the company announced its pending acquisition by Inßazyme Pharmaceuticals (Vancouver, British Columbia) in an equity transaction valued at approximately $12.8 million. Shortly after, GlycoDesign announced additional reductions in head count and research expenditures in anticipation of the merger. Shareholders approved the merger in May 2003. GlycoDesign focuses on the discovery of what the company calls “carbohydrate-processing inhibitors,” small-molecule compounds that inhibit

TABLE 3. Profiled Companies Active in Glycomics Company

Web Site

BioMarin Pharmaceutical GlycoDesign/Inflazyme

www.biomarinpharm.com

GlycoGenesys

www.glycogenesys.com

Glycominds

www.glycominds.com

Momenta Pharmaceuticals

www.momentapharma.com

Neose Technologies

www.neose.com

Optimer Pharmaceuticals

www.optimerpharma.com

Oxford GlycoSciences/Celltech Group

www.ogs.com/

www.glycodesign.com/ www.inflazyme.com

www.celltechgroup.com

Technology/Therapeutic Focus Enzyme replacement therapy Carbohydrateprocessing inhibitors; Core-2 discovery platform Lectin inhibitors; cancer Glycochip carbohydrate array; Glycomics Database; diagnostics Rapid carbohydratesequencing technology Carbohydrate synthesis and manufacture; technologies for modifying glycosylation Optimer Programmed One-Pot Synthesis (OPopS) for rapid carbohydrate synthesis; cancer vaccine Carbohydrateprocessing enzymes; miglustat

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the activity of enzymes involved in glycoprotein synthesis. The company’s technology platform includes genomics, bioinformatics, combinatorial chemistry, high-throughput screening, and computational and medicinal chemistry. The platform employs GlycoDesign’s Core-2 technology, a solid-phase glycosyltransferase high-throughput robotic screen, for discovery of small-molecule inhibitors of the core-2 glycosyltransferase enzyme (Donovan RS, 1999). The Core 2 technology was developed in an R&D collaboration with Seikagaku (Tokyo, Japan). When the partners restructured their agreement in January 2003, GlycoDesign regained rights to all intellectual property and commercialization rights arising from the collaboration; Seikagaku retains exclusive Japanese rights to the technology for internal R&D and use with third-party collaborators and will pay royalties to GlycoDesign on products discovered using the Core-2 assay system. Glycosyltransferases are particularly interesting targets for drug discovery because they can modulate glycosylation patterns of therapeutically relevant molecules in a relatively straightforward fashion. GlycoDesign’s glycosyltransferase program is aimed at discovery of small-molecule compounds that inhibit the activity of the Core-2 GlcNAc-T enzyme, a glycosyltransferase that catalyzes synthesis of the sLex carbohydrate structure. As previously explained (see “Inßammation”), this structure resides on the surface of circulating leukocytes and binds to selectins, an action that can trigger inßammation. Compounds that interfere with sLex synthesis could therefore be useful as anti-inßammatory therapeutics. GlycoDesign is also developing compounds for treating cardiovascular diseases and disorders. The company’s most advanced project is GH-9001, a glycosaminoglycan-based Phase I candidate drug intended for the treatment of cardiovascular disease and deep vein thrombosis. This project is attempting to create a small-molecule drug that can compete in the low-molecular-weight heparin market. The compound was originally developed at Vascular Therapeutics, a company that GlycoDesign acquired in August 1999. GlycoDesign is developing GH-9001 in a strategic alliance with the Danish company Leo Pharma (formerly Leo Pharmaceutical Products). The companies will share development costs until Phase II. Leo Pharma has an exclusive option to commercialize the compound in Europe and Canada and will pay royalties and milestones to GlycoDesign. In October 2002, the company reported positive results from a Phase I trial that assessed the safety and pharmacodynamics of GH- 9001. GlycoDesign and Leo Pharma are also developing another antithrombotic compound, GD-4040, which has demonstrated more efÞcacy than standard heparin and low-molecular-weight heparin in preventing arterial thrombosis in rabbits. The drug also prevented deep vein thrombosis in a rabbit model but was not superior to low-molecular-weight heparin. GlycoDesign is seeking development partners for two additional projects: a therapeutic for the prevention of neurocognitive deÞcits following cardiopulmonary bypass surgery and an antithrombotic coating for implantable medical devices.

CORPORATE HIGHLIGHTS

319

GlycoGenesys GlycoGenesys (Boston, Massachusetts), a biotechnology Þrm, is using glycobiology to develop novel drug candidates. The company is focusing its efforts on the development of GCS-100 (formerly GBC-590) as a potential treatment for multiple forms of cancer. The compound is an intravenous formulation of a modiÞed citrus pectin that inhibits galectin 3, a cell-surface lectin that is involved in cancer angiogenesis, metastasis, and malignant cell apoptosis. GCS-100 has demonstrated enhanced anticancer activity in chemoresistant cell lines that overexpress the Bcl-2 protein, which suppresses apoptosis and is overexpressed in many tumors. GlycoGenesys has evaluated GCS-100 in a Phase IIa clinical trial for refractory or relapsed pancreatic adenocarcinoma and for late-stage refractory or relapsing colorectal cancer. The drug was well tolerated and showed early signs of clinical activity. In late 2002, the company completed a Phase I doseescalation trial. GlycoGenesys is seeking strategic partners to help fund further clinical development of GCS-100 for pancreatic, colorectal, and other cancers. Glycominds Glycominds (Lod, Israel) is developing the GlycoChip, the Þrst commercially available, standardized, high-throughput screening array for analyzing protein–carbohydrate interactions. Similar in concept to arrays that researchers use in genomics and proteomics research, this product consists of a collection of molecules arrayed on a solid surface. The GlycoChip displays complex carbohydrates that are covalently linked to the chip. When exposed to a test solution, the carbohydrates on the chip can bind to and capture proteins and other molecules in the solution. Glycominds is collaborating with Schott Glass Technologies (SGT; Duryea, Pennsylvania), a U.S. subsidiary of the Schott Group, to design an improved version of the current glass substrate GlycoChip. Glycominds is using the GlycoChip as a diagnostic tool for identifying biomarkers of disease. These markers could be used to detect phenotypic variations between patients or variations caused by a particular patient’s changing metabolic or disease state. The company focuses on Þnding biomarkers associated with autoimmune and inßammatory diseases that can be developed into diagnostic tools for predicting and monitoring disease progression and drug response. In a recent proof-of-principal achievement, Glycominds was able to use the GlycoChip to differentiate between multiple sclerosis patients in different disease states (e.g., exacerbations, relapses); the company believes it has discovered a surrogate test that may be clinically useful for distinguishing between such patients. Glycominds seeks partnerships with pharmaceutical and diagnostic companies for collaborations in biomarker discovery and development in the areas of multiple sclerosis, rheumatoid arthritis, osteoarthritis, atherosclerosis, psoriasis, and Crohn’s disease. Like many genomics companies, Glycominds has also secured a foothold in the database business. The current release of its Glycomics Database (www.glycomics.com) includes more than 35,000 glycans from various sources and contains

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information on structures, functions, and interactions. The Glycomics Database uses LinearCode, a standardized form of notation for carbohydrate structures that was developed by Glycominds. LinearCode presents complex information about carbohydrates and glycoconjugates (including stereospeciÞcity, ring and bond conformation, chemical modiÞcations, and branch patterns) in a relatively simple format. Many researchers in the glycomics community have accepted LinearCode as a standard notation. Momenta Pharmaceuticals As mentioned earlier, Momenta Pharmaceuticals was founded in 2001 to commercialize technologies developed at MIT (see “The Tools of Glycomics”). The company is using its technology, which enables rapid, precise characterization and engineering of complex carbohydrates and glycoslyated molecules, to develop a more complete understanding of the roles that carbohydrates play in cellular function, disease, and drug action. Momenta intends to apply these insights to the development of improved versions of existing therapies and, eventually, the creation of novel therapies. In May 2003, Momenta raised $19 million in private equity Þnancing. The company is seeking corporate partners for a variety of development programs. Neose Technologies Neose Technologies (Horsham, Pennsylvania) develops technologies for the synthesis and manufacture of complex carbohydrates. The company has a suite of technologies for enhancing the safety and efÞcacy of currently marketed therapeutic proteins. GlycoAdvance is a set of products and services for correcting incomplete or incorrect glycosylation of industrially produced recombinant glycoproteins. The company’s GlycoPEGylation technology allows the targeted addition of polyethylene glycol (PEG) to carbohydrate chains of glycoproteins; adding PEG to these drugs helps preserve their bioactivity and improve their pharmacokinetic proÞles. GlycoConjugation, an extension of GlycoPEGylation, is a tool for improving antibodies and glycoproteins by attaching new bioactive or functional carbohydrate moieties. Neose plans to develop its Þrst proprietary products in hematology. In January 2003, the company selected erythropoietin as a target for improvement using the GlycoPEGylation technology. The company intends to apply a similar strategy to developing proprietary versions of other currently marketed drugs. In February 2003, Neose announced the private placement of $17 million of common stock to Þnance its internal development programs. Most Neose efforts to date have focused on collaborations with large pharmaceutical companies that apply Neose platform technologies. In 2002, Neose entered into two R&D agreements with Novo Nordisk to apply GlycoAdvance and GlycoPEGylation technologies to improve therapeutic proteins. Neose also has an agreement with Monsanto to use GlycoAdvance to enhance monoclonal antibody therapeutics produced in plants. According to an agreement

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with Novazyme (purchased by Genzyme in March 2001 for $225 million) that was restructured in March 2001, Neose could collect royalties on sales of lysosomal storage disease products made using Neose technology. In an ongoing R&D collaboration (begun in 2000) with Neuronyx (Malvern, Pennsylvania), which specializes in central nervous system glycobiology, Neose will use its technology to synthesize and produce modiÞed glycolipids for the treatment of Parkinson’s disease. In December 2001, Neose entered into an R&D and licensing agreement with Wyeth Pharmaceuticals to apply GlycoAdvance technology to improve Wyeth’s recombinant P-selectin glycoprotein ligand (rPSGL-Ig). The compound was in Phase II trials for myocardial infarction; Neose technology was to be applied to Phase III development and commercial manufacturing. However, in May 2002, due to disappointing Phase II results (which, Neose states, were unrelated to its technology), Wyeth decided to cancel clinical development of the compounds and terminated the collaboration in September 2002. Optimer Pharmaceuticals Optimer Pharmaceuticals (San Diego, California), a biotechnology company that focuses on carbohydrate science, was founded in 1998 on technology licensed from the Scripps Research Institute, including Optimer Programmed One-Pot Synthesis (OPopS), which enables rapid synthesis of oligosaccharides. The company claims that OPopS systems can reduce the time required for synthesis from months (using conventional methods) to hours. The technology can be used to modify or attach carbohydrate groups to existing compounds or to generate carbohydrate arrays for screening. Optimer is using OPopS for medicinal chemistry in the company’s drug discovery programs. Optimer’s lead drug candidate, OPT-22, is a cancer vaccine that consists of the Globo-H hexasaccharide conjugated to keyhole limpet hemocyanin, a highly immunogenic protein. Optimer licensed the drug from Memorial Sloan-Kettering Cancer Center in September 2002; the drug is in Phase I clinical trials for prostate and breast cancer. The company is developing another compound, OPT-80, for treating Clostridium difÞcile associated disease (CDAD) and was testing the drug in Phase I clinical trials in late 2003. Another discovery-stage compound is OPT-88, which Optimer is evaluating as an intra-articular injection therapy for osteoarthritis. The company also has drug discovery programs in the areas of infectious disease and HIV. Oxford GlycoSciences/Celltech Group Oxford GlycoSciences (OGS; Abingdon, United Kingdom) is a biopharmaceutical company that specializes in the development and application of proteomics technology. OGS’s goal is to discover proteins that are important in diagnosing, treating, or monitoring disease. The company is pursuing development efforts in the areas of antineoplastic drugs, antifungals, and glycosphingolipidosis drugs.

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Most OGS glycobiology-based drug development efforts have centered on compounds that target carbohydrate-processing enzymes. The company’s lead compound is miglustat (Zavesca; OGT-918), an orally bioavailable, small-molecule, glucosylceramide glucosyltransferase inhibitor (chemically, an iminosugar) that the company licensed from G.D. Searle in 1997. In November 2002, the European Commission granted marketing authorization for miglustat for the treatment of type I Gaucher’s disease in patients for whom enzyme replacement therapy is unsuitable; the commission requested provision of follow-up safety data based on postmarketing surveillance. The European Commission’s approval of miglustat surprised many analysts because in June 2002, the FDA sent OGS a not approvable letter and requested additional clinical data to support the drug’s safety and efÞcacy. In March 2003, OGS submitted an amendment to its new drug application (NDA) to address the FDA’s concerns. In March 2003, Actelion (Allschwil, Switzerland), OGS’s marketing partner, launched miglustat in Europe. Excepting Israel, Actelion has worldwide marketing rights to miglustat; in an agreement signed in November 2001, OGS granted exclusive marketing rights in Israel to Teva Pharmaceuticals. In June 2003, the Israeli Ministry of Health approved miglustat for treating type I Gaucher’s disease in patients for whom enzyme replacement therapy is unsuitable. Analysts predict that annual sales of miglustat could exceed $20 million in Europe and Israel and reach approximately $20 million in the United States. OGS is also testing OGT-923, an analogue of miglustat, which is in Phase I clinical trials begun November 2002. The compound is a potential treatment for glycosphingolipid storage disorders with nervous system involvement, such as Sandhoff’s disease and Tay-Sachs disease. In January 2003, OGS and Cambridge Antibody Technology Group (CAT) announced their intention to merge under an all-share acquisition by CAT worth £110 million. At the same time, Celltech Group (Slough, United Kingdom) was preparing an unsolicited cash offer of £101.4 million. OGS denounced the offer as inadequate and a “spoiling tactic” to “acquire OGS on the cheap.” However, a steep drop in CAT’s share price between January and April caused the value of that company’s original offer to plummet by approximately 35%. By mid-April 2003—after OGS had considered other options and continued to burn through its cash reserves—Celltech had acquired signiÞcant blocks of OGS stock and effectively taken over the company. Celltech has indicated its interest in retaining the miglustat franchise, oncology assets, and other OGS programs. Glycomics Activities of Large Pharmaceutical Companies Many major pharmaceutical companies have implemented some form of carbohydrate research in their R&D departments; however, most of these activities are not concentrated glycomics programs. Some Þrms are collaborating with glycomics or glycobiology specialty companies for speciÞc research projects; we expect this partnering trend to continue. Japanese companies are presently at the forefront of biochemical research activities that focus on carbohydrates. Eisai’s

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Tsukuba Research Laboratories (Tsukuba, Ibariki, Japan) Þgures prominently, in part through a development program for modiÞed tissue plasminogen activators. Sankyo and Meiji Seika Keisha (both in Tokyo) also dedicate signiÞcant resources to carbohydrate chemistry, especially projects concerned with inhibitors of carbohydrate processing. Providers of Specialty Tools and Technologies for Glycomics Dozens of small companies offer various research tools, reagents, specialized technologies, and analytical services that can be useful in glycomics research and drug development. Among them are Dextra Laboratories (Reading, United Kingdom), Ludger Glycotechnology (Oxford, United Kingdom), Iduron (Manchester, United Kingdom; a division of Matrix Therapeutics), and Glygen (Columbia, Maryland). Proteome Systems (North Ryde, Australia, and Woburn, Massachusetts) offers GlycoSuiteDB, a relational database of glycoprotein-derived glycan structures and related information (Cooper CA, 2001). OUTLOOK From a commercial perspective, the emerging Þeld of glycomics resembles the related Þelds of genomics and proteomics. All three disciplines involve highthroughput analysis and the exploitation of large data sets that describe fundamental aspects of human biology, and all three have signiÞcant implications for understanding disease and designing new medicines. The success of companies operating in these areas depends on the development and utilization of technologies that enable researchers to conduct direct experiments and analyze results on a very large scale. The experiences of companies in the genomics and proteomics sectors may provide a model for what we can expect in the glycomics sector. These experiences suggest that well-positioned “tool” companies—those with ßexible and proprietary technologies, particularly for rapid carbohydrate analysis and synthesis—will initially lead the Þeld. The tools segment of the glycomics business will reach its zenith by 2006–2007. This Þrst generation of companies will be followed by a proliferation of drug developers with specialized platform technologies and tool companies that have reinvented themselves as generators of drug leads. Momenta Pharmaceuticals is an example of a company that is already positioning itself within this business model. Large pharmaceutical companies will team with these small innovators in the early stages; many big pharmas will eventually decide to integrate glycomics platforms into their internal R&D programs. We expect strategic partnering and technological integration through merger and acquisition to be widespread during the second half of the decade. After a period of growth, the industry will consolidate, and only a few of the early specialty companies will survive as independent entities. Mergers and acquisitions will also root out many companies that have a sound technical basis but insufÞcient funding or poor management.

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Glycomics companies are targeting markets that generate billions of dollars a year in drug sales. The four core applications of atherosclerosis, inßammation/infection, cancer, and coagulation would alone provide an enormous target market. Because glycosylation plays a central role in nearly every therapeutic area, the potential for its application is extensive. Even the markets for enzyme substitution therapies that treat the symptoms of congenital glycosylation disorders, such as Gaucher’s and Fabry’s diseases—which affect only a tiny percentage of the human population—could ultimately generate billions of dollars in revenue. Nevertheless, even though researchers have long recognized glycobiology as the “unexplored third land” in the realm of biology (the Þrst two being genomics and proteomics), applied carbohydrate research has so far remained on the sidelines of drug discovery and development. Some experts have noted that the emerging area of glycomics is at least ten times as complicated as conventional proteomics. Combined with a paucity of high-throughput technologies, this complexity makes glycomics a difÞcult Þeld to exploit. Quantifying the commercial scope of glycomics at this early stage is therefore difÞcult. We believe that the potential of glycomics-based drug discovery is huge, but realizing this potential will require an enormous investment in technological development and signiÞcant breakthroughs in analytical and synthetic capabilities.

REFERENCES Cooper CA, et al. FlycoSuiteDB: a new curated relational database of glycoprotein glycan structures and their biological sources. Nucleic Acid Research. 2001;29(1):332–335. Dell A, Morris HR. Glycoprotein structure determination by mass spectrometry. Science. 2001;291:2351–2356. Donovan RS, et al. A solid-phase glycosyltransferase assay for high-throughput screening in drug discovery research. Glycoconjugate Journal. 1999;16(10):607–615. Guerrini M, et al. A novel computational approach to integrate NMR spectroscopy and capillary electrophoresis for structure assignment of heparin and heparin sulfate oligosaccharides. Glycobiology. 2002;12(11):713–719. Sundaram M, et al. Rational design of low-molecular-weight heparins with improved in vivo activity. Proceedings of the National Academy of Sciences, USA. 2003;21; 100(2):651–656. Venkataraman G, et al. Sequencing complex polysaccharides. Science. 1999;286:537–542.

High-Content Screening for Drug Discovery and Target Validation

SUMMARY High-content screening (HCS) is an emerging technology that allows researchers to visualize individual cells and measure multiple cellular processes over time. HCS provides information on functional changes occurring at the cellular level, which is potentially more predictive of clinical outcomes than biochemical screening data. A powerful feature of HCS is that potency, selectivity, and cytotoxicity can all be determined in a single assay. However, efÞcient methods for extracting and integrating the image data generated by HCS assays have yet to be fully developed. Therefore, many companies formerly focused on high-throughput screening (HTS) are now developing instrumentation, software, and reagents for HCS. In this article, we review the current state of HCS technology, the results of our survey of HTS lab directors, and opportunities for HCS technology development, and we proÞle several companies that are leading the way in the development of innovative tools and software for HCS. BUSINESS IMPLICATIONS •

High-content screening (HCS) is a method for tracking inter- and intracellular activity by using probes and various imaging technologies. It is regarded as one of the most promising recent advances in drug discovery. Among the numerous advantages offered by HCS is the ability to observe the effect of a potential drug on various pathways within an individual cell over an extended period. HCS permits researchers to perform multiple, simultaneous assays in a single cell—that is, multiplexing.

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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HCS is much more challenging than high-throughput biochemical assays or cell-based assays that do not rely on imaging technology. Despite rapid changes in imaging technologies, efÞcient methods for extracting and integrating the image data generated by these assays have yet to be fully developed. Furthermore, HCS is most clinically relevant when researchers use untransformed human or animal cells (“primary cells”), but these cells are relatively expensive and cannot be maintained for long periods in vitro. New probes and labels currently in development will provide improved spectral properties, less disruption of cell components, and the ability to detect small molecules within cells. Many companies formerly focused on highthroughput screening (HTS) are now developing instrumentation, software, and reagents for HCS. Interviews with HTS laboratory directors at major pharmaceutical companies reveal that HCS has been embraced as an important and promising technology in drug discovery and development. Wider adoption is expected as further advances are made in supporting technologies that are essential to performing HCS more rapidly and at lower cost.

INTRODUCTION High-content screening (HCS) is an emerging technology that allows researchers to visualize individual cells and measure multiple cellular processes over time. HCS provides information on functional changes occurring at the cellular level, data that are potentially more predictive of clinical outcomes than biochemical screening data. HCS grew out of traditional cell microscopy and is being enabled by developments in probes, imaging, liquid- and cell-handling instrumentation, and image analysis software. Most HTS directors use charge-coupled device (CCD) imaging for HCS because of their experience with the speed and quality of this technology; others have begun to use laser scanning to achieve higher throughput. New imaging and detection technologies in development will facilitate higher throughput with a high level of quality and resolution. Higher throughput and better software are the two main areas where improved imaging platforms are being sought for HCS. The term high-content screening was coined by the founders of Cellomics, the Þrst company to pursue this technology. HCS is distinguished from microscopy by its use of automation and multiplexing—that is, simultaneous assays that can be conducted in the same well. HCS differs from high-throughput, cell-based assays in its use of imaging that allows visualization of individual cells. Whereas cell-based screening measures speciÞc parameters of cell populations, HCS can be used to study the processes occurring within individual cells. HCS differs from ßuorescent-activated cell sorting (FACS) in its ability to facilitate the study of tissues, adherent cells, and cell–cell interactions. A powerful feature of HCS is that potency, selectivity, and cytotoxicity can all be determined in the same assay. This feature offers the potential to speed lead optimization and improve

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the quality of new molecules. At present, the chief limitations of HCS are its high cost and its relatively low throughput; these factors have led most researchers to limit their use of HCS to secondary screening applications. To evaluate the pace at which HCS is being adopted, we conducted an interview survey during the fourth quarter of 2003 of directors of high-throughput screening (HTS) laboratories in the United States and Western Europe. Participants were employed at major pharmaceutical and biotechnology companies and at several academic and government-funded laboratories. Results of the survey (discussed in detail in the following section) show that HCS is being used in primary and secondary screening applications by approximately onefourth of the pharmaceutical and drug discovery companies included in the survey. We expect HCS to continue to make inroads as a key tool for drug discovery. Technological advances that will drive increased use of HCS are higher throughput, improved multiplexing, and better data analysis tools for linking data output with biological function. MARKET POTENTIAL OF HIGH-CONTENT SCREENING HCS is useful in drug screening because it provides information about the actual functional changes that occur in a disease target rather than simply whether or not a compound binds to a receptor. It also results in fewer false hits, thereby saving time and money. HCS has proved useful in target validation, assay development, and prioritization of lead compounds, and in elucidating the drug mechanism of action within a cell. The major hurdles that remain to be overcome in HCS include the achievement of a high-throughput platform with high-quality resolution and better analytical tools. Survey of HTS Laboratory Directors As part of a larger study of high-throughput screening (HTS), we conducted an interview survey of 51 directors of HTS laboratories, focusing on their use of and views of HCS. (Because one director contributed data from three laboratories, data were collected on a total of 53 laboratories.) Interviews were conducted using a structured questionnaire consisting of approximately 60 questions, with additional questions asked depending on the responses provided. The geographic distribution of respondents was diverse: 62% of the laboratories are located in the United States, 11% in the United Kingdom, and the remaining 27% in Austria, Denmark, Germany, Spain, Sweden, and Switzerland. The survey included nine of the top ten pharmaceutical companies (ranked by 2003 sales), as well as many of the larger biotechnology companies. Most HTS directors agreed that HCS is the most promising new technology in drug discovery, but opinions varied as to the proper deÞnition of HCS. Some interviewees deÞned HCS as any screen that provides multiple end points from one well, while others deÞned it as automated subcellular imaging.

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FIGURE 1. Stages of drug discovery in which high-content screening users apply the technology.

Of the 53 HTS laboratories represented in this study, 24% are currently using some type of HCS in primary or secondary screening. Among the HTS laboratories using HCS, only 25% apply the technology to primary screening, whereas 75% use HCS in secondary screening. (Secondary screening includes selectivity, potency, toxicity, and cell permeability testing.) Three laboratories in our survey also use HCS for assay design, compound prioritization, or target validation. Figure 1 depicts the applications for which HCS is currently used. We also asked HTS laboratory directors (including those who are not yet using the technology) for their views about the potential utility of HCS in drug discovery. Most lab directors (including current users and nonusers) believe that it is too early to determine whether HCS will prove useful in improving the efÞciency of drug discovery (see Figure 2). One concern expressed by many respondents is the challenge of interpreting the results from HCS studies and applying this information directly to drug discovery. Laboratory directors whose labs now use HCS stated that the technology has proved useful in validating drug leads, determining mechanism of action, prioritizing compounds, or designing assays. Figure 3 shows the percentage of HTS lab directors currently using or planning to use HCS and the percentage of nonusers who believe this technology is promising. Figure 4 shows the applications for which HCS is currently used. Before laboratories can apply HCS, they must have the capability to perform cell-based assays. As HTS laboratories gain experience with cell-based assays, use of HCS will expand in the HTS setting. Our survey revealed that all but three HTS laboratories currently perform some cell-based assays, and two of these three expect to be using cell-based assays by 2005. Figure 5 shows survey respondents’ expectations regarding the use of cell-based assays over the period 2003 to 2005. As a percentage of all assays used by the HTS laboratory directors interviewed, cell-based assays will represent more than 50% of assay throughput by 2005. Expenditures on cell-based assays will account for more than 13% of the overall HTS market. In summary, our survey of HTS laboratory directors suggests that HCS is one of the most promising technologies for numerous aspects of drug discovery,

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FIGURE 2. Usefulness of high-content screening in drug discovery: 51 HTS laboratory directors respond.

FIGURE 3. Current use of high-content screening in high-throughput screening.

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FIGURE 4. Current and planned use of high-content screening.

FIGURE 5. Expected changes in use of cell-based assays, 2003 to 2005: percentage of respondents.

including creation of assays that are more predictive of clinical results, better understanding of drugs’ mechanisms of action, target identiÞcation, and lead optimization. Drivers of High-Content Screening Growth in HCS development is being driven by the expectation that the technology will accelerate drug discovery and improve compound quality. HCS is part of a general trend toward the “industrialization” of biology that enables the identiÞcation of

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more targets and the screening of more compounds in more assays. While uncertainties remain as to whether HCS can meet the high expectation it has engendered, HCS is being adopted despite the technological limitations of this nascent Þeld. Some of the key reasons for this adoption are summarized here: • • • •

HCS allows multiplexing that could reduce the number of assays that have to be run, thereby reducing costs. HCS can speed and improve target validation because the multiple targets within cells can be tested in a single assay. Cell-based assays facilitate assessment of cytotoxicity, cell permeability, and compound potency. When multiple cell types are present in the same assay, compound selectivity, potency, and cytotoxicity can be measured concurrently.

The major driver of HCS use, however, is the ability to examine processes within living cells. Studying the actions of drugs in human cells can provide more-relevant information about biological function than studies conducted in isolated biochemical systems. Certain assays are difÞcult or impossible to reduce to a biochemical system—for example, apoptosis, mitosis, colony formation, neurite outgrowth, or cell motility. In addition, when the target is not known, it simply is not possible to create a biochemical assay. Finally, conducting cell-based assays earlier in the drug discovery process could lower costs by eliminating undesirable compounds early, thus reducing the cost of downstream development.

OPPORTUNITIES FOR TECHNOLOGY DEVELOPMENT Many important advances in HCS have been made during the past Þve years. Nonetheless, signiÞcant challenges remain. For example, existing technologies do not yet permit the imaging of multiple dynamic processes in living cells, processes that cannot be captured by examining isolated biological molecules in vitro or by analyzing indirect biochemical or genetic data. Other challenges include determining the biological relevance of multiparametric data, the integration of data from different platforms, maintenance of relevant cell types, the need for more robust assays, and spectral overlap. The principal advantage of HCS is that it enables researchers to study the movement and interactions of cellular components as they occur; these processes cannot be captured by examining isolated biological molecules in vitro or by analyzing indirect biochemical or genetic data. Biological molecules change their structures, and as cellular processes occur, interactions between individual molecules change. Furthermore, imaging frozen or Þxed cells does not provide information on dynamic cellular processes. HCS therefore provides the best method for understanding how cells work and for identifying opportunities for therapeutic interventions—but opportunities remain for improving HCS assays and tools.

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HCS requires the coordination and integration of many advanced technologies, such as automation, molecular biology, genetic engineering, protein engineering, ßuorescence chemistry, cell biology, imaging tools, terabyte data management, and image processing algorithms. Improvements in each of these areas are necessary if HCS is to be broadly adopted by HTS laboratories. Table 1 summarizes needed improvements in HCS tools noted by participants in our survey of HTS laboratory directors. In particular, researchers using HCS state that they need better reagents and tools for use in dynamic imaging. Probes and Detection Technology The ability of HCS to follow dynamic cellular processes is limited by the chemical and photo properties of existing molecular probes (sometimes called reporters). These probes can be proteins with endogenous ßuorescence or ßuorescent or luminescent chemicals. Typically, signals arising from tens to thousands of probe molecules are required for detection of cellular processes. This requirement can be a limiting factor in HCS because biomolecular targets may be present in cells in very low concentrations (e.g., a few molecules per cell). The limitations of existing probes include their large size relative to the biological molecule being detected (which can sterically interfere with the process being measured), spectral overlap between probes (which can hinder distinguishing between signals), low spectral intensity of the probe, and photobleaching. Other potential problems are related to a probe’s interaction with a cell. For example, it may be difÞcult to engineer coexpression of the probe and the molecule of interest, or to get the probe to enter a particular organelle. Probes. Live-cell microscopy was largely enabled by the application of ßuorescent proteins such as green ßuorescent protein (GFP), found in jellyÞsh. The ßuorescent proteins can be fused to almost any protein of interest in a cell by using genetic TABLE 1. Improvements Needed in High-Content Screening Tools • More-sensitive probes (probes with higher quantum yield) that can detect single molecules. • Generic probes that can be applied to multiple targets. • Solutions to problems of low spectral intensity and photobleaching. • Smaller labels that minimize disruption of molecular processes. • Improved fluorescent proteins found in nature, evolved in vitro, or engineered. • Engineered proteins that bind optically active and membrane-permeable probes. • Multifunctional probes that can be used with more than one detection method (e.g., with optical or electron microscopy). • Improved multiplexing strategies. • Methods to deliver particles such as nanoparticles or quantum dots without disrupting cells. • Improved signal-detection technology.

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engineering. GFP ßuoresces when excited by light, requiring no other enzymes or cofactors. When GFP is fused to a protein of interest, the formation and location of the protein can be followed by microscopy. Types of experiments that use ßuorescent proteins in HCS include monitoring of cell signaling by second messengers, monitoring intracellular chemical environments, protein–protein interactions, and protein translocation. The wild type GFP has been mutated to improve its folding properties, improve stability, increase ßuorescent kinetics, produce brighter molecules, reduce photobleaching, and ßuoresce with different colors (Lippincott-Schwartz J, and Patterson GH, 2003). Blue, cyan, yellow, and red ßuorescent proteins are now commercially available from Promega, Clontech (now a division of BD Biosciences), Evrogen (Moscow, Russia), MBL International (Woburn, Massachusetts), and other companies. Another widely used reporter is luciferase, which does not autoßuoresce and requires the addition of a luciferin label (i.e., a small-molecule ßuorescent reporter group). Luciferase enzymes catalyze chemical reactions that produce light. Luciferin–luciferase systems are found in bacteria, insects such as the Þreßy, and many other organisms. Different luciferases have slightly different photo properties, as do different luciferins. Numerous reagent suppliers market luciferases and luciferins. Other ßuorescent proteins can be used to tag proteins of interest. Promega has developed the HaloTag Fusion Marker, which has an interchangeable ßuorophore (a molecule or protein, such as GFP, that can ßuoresce). HaloTag is an engineered haloalkane dehalogenase enzyme from Rhodococcus rhodochrous. The enzyme is covalently bonded to a ligand such as ßuorophore or biotin, a commonly used afÞnity tag. The advantage of the HaloTag approach is that the cells do not have to be reengineered to produce ßuorophores of different colors, as would be required with constructs engineered to express a particular GFP. Dyes are covalently attached to the HaloTag, and these dyes can be exchanged for other dyes or for afÞnity tags such as biotin. Commercial opportunities exist for engineered ßuorescent proteins with new properties for investigation of cellular processes. As researchers conducting HCS gain more expertise in following multiple processes in cells, the demand for ßuorophores with more colors and different photo properties will expand. SpeciÞc needs are for brighter molecules, more red-ßuorescing proteins, and ßuorophores with narrower emission peaks so that multiple colors do not overlap. Because animal tissues have the greatest transparency in the infrared and far-red regions, new ßuorescent proteins with emission in these regions of the light spectrum could be particularly useful as intracellular sensors. Fluorescent Labels for Other Molecules. GFP has enabled multiple screening applications (especially for monitoring protein expression and protein localization) courtesy of the bright ßuorescence signal it can produce. Similar methods are needed for detection of RNA, small molecules, metabolites, second messengers, and drug candidates.

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Scientists at Columbia University recently reported the development of probes that use RNA aptamers to bind small ßuorescent organic molecules (Stojanovic MN and Kolpashchikov DM, 2004). Aptamers are short RNA sequences that can speciÞcally bind small molecules or proteins. The RNA-based probes have three domains: one that binds a ßuorescent reporter (the ßuorescent molecule that is detected); a detector domain optimized to bind the ßuorescent small molecule of interest; and a connector domain between the reporter and detector domains. Thus far, reporters have been demonstrated for adenosine triphosphate (ATP), ßavin mononucleotide (FMN), and theophylline. Similar RNA aptamer constructs could be used as sensors for intracellular imaging. These aptamers offer several advantages compared with other probes. They allow detection of small molecules and, because they are smaller than either GFP or luciferase, present less chance of steric interference. In addition, they do not need to be covalently linked to a protein molecule. Successful application of aptamers for small-molecule detection in cells would open up the possibility of monitoring cellular molecules over time and would dramatically improve researchers’ ability to monitor cellular processes, drug response, and drug metabolism. Quantum Dots. Quantum dots are semiconductor nanocrystals of 2–10 nm that emit light following excitation with light at a higher frequency. Quantum dots have several attractive properties that could make them an alternative labeling technology to chemical ßuorescent dyes or ßuorescent proteins for some HCS applications. Quantum dots are embedded in an inert plastic matrix and are sufÞciently small (5–50 nm) that many hundreds can Þt inside a cell. Encapsulated particles can be decorated with biological molecules such as oligonucleotides, small molecules, antibodies, or proteins that can enter cells by endocytosis. Because quantum dots are photostable, they can emit light for up to 30 minutes; in contrast, ßuorescent dyes emit light only for seconds. The extended duration of emission would permit more kinetic processes to be followed. The excitation source required for quantum dots can be any wavelength, provided it is shorter than the emission wavelength. In contrast, dyes and ßuorescence proteins often require speciÞc excitation frequencies and expensive light sources. It is possible to create quantum dots that emit nearly any color, and multiple colors can be emitted from a single bead, thereby providing huge numbers of coding possibilities. Companies developing quantum dots for a variety of applications, including HCS, are CrystalPlex, Evident Technologies, and Quantum Dot. Cell Handling Like non-imaging-based cell assay methods, HCS requires successful handling of living cells. Producing, growing, and maintaining these cells can be difÞcult. Handling noncancerous (untransformed) cells that come directly from human or

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animal donors (“primary cells”), which cannot be cultured for long periods, adds an additional measure of difÞculty. Primary cells are more predictive of human response but are more difÞcult to maintain and assay than cancer (transformed) cell lines—but transformed cell lines have undergone many signiÞcant changes that undermine their predictive utility. Better technologies for cell sourcing and handling would help to improve the utility of HCS. Among the new types of instrumentation needed to address these issues are highly automated cell-handling instruments, capable of stringently controlling cell media and environmental conditions, and ßuid dispensers that can handle cells. Procedures and techniques for freezing cells while maintaining cell viability would provide signiÞcant advantages for the logistics of conducting cell-based assays and in reducing interassay variation because cells could be frozen, stored, and subsequently thawed when robotics and other assay reagents are ready to use. Companies developing equipment for automated cell handling include Cytogration, Tecan, RTS LifeSciences, and The Automation Partnership. Engineered Cells. A wide range of opportunities awaits the creation of engineered cells, including cell lines that express particular targets, cells with expression vectors that provide stable expression levels, and cells that require only simple growth conditions. Researchers working in drug development need consistent and reliable assays, and such assays can be achieved with engineered cells already transfected to express reporter genes, drug targets, or drug-metabolizing enzymes. Because live cell lines are expensive to transport, and because drug targets are usually proprietary, researchers typically purchase vectors and transfect their own cells. Commercial opportunities exist for companies that can offer cells that express widely used drug targets with reporter systems already included and cells that are express reporter systems, which can simply be transfected with proprietary drug targets. One vendor of engineered cell lines is BD Biosciences, whose BD Living Colors cell lines are stably transfected to express a variety of reporters (e.g., HEK293 cells expressing GFP coupled to a proteosome-targeting sequence). Stratagene markets ReceptoScreen stable cell lines that express cytokines and chemoattractant receptors. Otsuka Pharmaceutical markets CHO cells that express estrogen or androgen receptors and a luciferase reporter system for use in endocrine disruptor assays. Image Analysis and Data Management Two of the major challenges presented by HCS are the need to extract data from images automatically and the need to manage large volumes of data. HCS images produce terabytes (1012 bytes) of data, thus creating challenges with respect to both storage and analysis. Image analysis programs are needed to quantify and analyze different assays, adding the task of writing image analysis programs to that of assay development. Historically, image analysis has been done visually in microscopy. To achieve automation, however, computer algorithms that analyze images are required.

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Approaches to image analysis include artiÞcial intelligence and pattern recognition, both of which were initially developed for military applications. Each new assay requires development of image analysis protocols. Software companies with image management or analysis expertise that are developing products for HCS include SciMagix, SpotÞre, and DeÞniens. Correlation of image information with biological function is very complex. One of the Þrst hurdles is to deÞne which of the many potential parameters should be measured. For several biological questions, monitoring just one or two parameters yields the necessary data—but identiÞcation of the right parameters to measure is often difÞcult. Machine learning, a type of artiÞcial intelligence, is a method of training computer programs to understand new data, such as cellular images. Two common types of machine learning that are classiÞed by the outcomes they produce are supervised and unsupervised learning: •

•

Supervised Learning. In this type of machine learning, the algorithm is trained on examples of known results. After the training, the program can be used to determine the answers to questions about new data, where the answer is not known. In the context of cell image analysis, an example of supervised learning would be the use of drugs with known cellular effects (e.g., apoptosis) to train the automated image analysis program to recognize an image of an apoptotic cell. Unsupervised Learning. In this type of machine learning, the program is not trained using known results; instead, it is provided with a model that best Þts all the data provided. In the context of cell image analysis, the program may be provided with several images and may identify some characteristic that groups images into cellular responses (e.g., apoptosis or neurite outgrowth).

Reify is using unsupervised learning algorithms to characterize changes in cell morphology. The company recently developed algorithms to measure contraction of cardiomyocytes (Hack AA, 2004). Cytoprint (now part of Atto Biosciences) uses a very different approach, one that derives biological information for image analysis. Algorithms are developed by training the software program to recognize example images of positive and negative controls. Once trained, the program evaluates the effects of a test compound. The program can also be trained to recognize out-of-focus ßuorescence images and nonspeciÞc morphology changes (Elling J, 2004). The German company DeÞniens’ Cellenger system performs semiautomatic, object-based image analysis. The system Þrst recognizes the largest objects and stores such features as shape or color, then links those data to biological entities such as the nucleus or mitochondria. The system then determines more detailed features of these images, analyzing quantitative features such as area, length, width, and intensity (Biberthaler P, 2003). Cellomics provides a portfolio of algorithms called BioApplications that biologists can use to analyze assay data instead of having to write their own scripts or programs. The software collects data relevant to the biological properties of the

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cells being assayed, such as size, shape, amount of ßuorescent label, and pattern of ßuorescence. Data are reported for the whole cell population, for subpopulations, and for individual cells. Cellomics is prominent among companies with expertise in biology that are partnering with or acquiring software and information technology Þrms. GE Healthcare has licensed Cellomics technology to accelerate the drug discovery process, and IBM will integrate the company’s rapid screening technology with its own data and storage management software. Examples of other collaborations include Molecular Devices’ acquisition of Universal Imaging in 2002, which provided the Þrm with image analysis expertise, and Atto Biosciences’ acquisition of image analysis software developer Cytoprint. A few companies, such as Vitra Bioscience, have opted to develop their own image analysis software. BUSINESS MODELS AND STRATEGIES All the companies developing new instrumentation, reagents, or software for HCS are tool providers. Table 2 lists select companies providing HCS-related products and applications. With the exception of Norak Biosciences, which has a separate drug discovery division, none of the companies listed in Table 2 is engaged in developing its own therapeutic products. Among the companies listed, Þve Þrms—Blueshift, Cellomics, Cellumen, Reify, and Vitra—were founded solely to capitalize on HCS, while the others moved into HCS applications from related areas, such as high-throughput screening. Trends in Partnering and M&A The complexity of HCS—a Þeld that requires expertise in automation, imaging, cell biology, chemistry, and image analysis—is driving increased merger and acquisition (M&A) activity between Þrms with complementary technologies. In July 2004, Atto Biosciences was purchased by Becton Dickinson’s BD Biosciences division for $25 million; in the same month, Axon Instruments merged with Molecular Devices in a $200 million deal. As noted previously, HCS companies are also collaborating with software companies to analyze image data. They are also partnering with reagent supply companies, particularly those that supply improved chemical dyes and genetically engineered proteins. PROFILES OF SELECT HCS COMPANIES Atto Bioscience Atto Bioscience (Rockville, Maryland) is developing confocal microscopy hardware, software, and assays for cellular analysis. The company was founded as Atto Instruments in 1985 and was purchased by BD BioSciences in July 2004.

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TABLE 2. Select Companies Offering High-Content Screening Products and Applications Company GE Healthcare (formerly Amersham Biosciences) Amnis

Atto Bioscience (subsidiary of BD Biosciences) Axon Instruments

BD Biosciences BioImage A/S Blueshift Biotechnologies Cellomics

Cellumen

CompuCyte CrystalPlex

Cyntellect

Definiens Evident Technologies

Evotec Technologies Gmbh (subsidiary of EvotechOAI)

Molecular Devices

Norak Biosciences

Web Site

Products and Technologies

www1.amershambiosciences. IN Cell analysis system, confocal com imaging www.amnis.com ImageStream 100 system, flow cytometry and microscopy combination instrument www.atto.com BD Pathway, high-content, automated confocal imaging; CARV, confocal adaptor www.axon.com ImageXpress, automated cellular screening instrumentation platform www.bdbiosciences.com LivingColors, cell lines that express reporter genes www.bioimage.com Redistribution assays, for protein translocation www.blueshiftbiotech.com Dynamic fluorimetry technology; laser scanning fluorimeter www.cellomics.com ArrayScan HCS reader; KineticScan HCS reader for conducting kinetic assays; HCi informatics platform www.cellumen.com Developing new assays to manipulate and measure cell processes www.compucyte.com iCys research imaging cytometer www.crystalplex.com PLxBeads, encapsulated quantum dot beads prepared for bioconjugation www.cyntellect.com LEAP technology, cell manipulations using lasers combined with imaging www.definiens.com Cellenger, software for pattern recognition www.evidenttech.com EviTags, encapsulated quantum dot beads prepared for bioconjugation; EviFluors, quantum dot nanoparticles www.evotecOpera, confocal fluorescence technologies.com microplate imaging reader; Clarina II, workstation with fluorescence plate reader for HCS www.moldev.com Discovery-1, HCS screening system; AquaMax, liquid handling system www.norakbio.com Transfluor technology, translocation assay for analysis of GPCRs

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339

TABLE 2. (continued) Company

Web Site

Reify

www.reifycorp.com

Promega

www.promega.com

Quantum Dot

www.qdots.com

Scimagix

www.scimagix.com

Tecan

www.tecan.com

TTP Lab Tech

www.ttplabtech.com

Vitra Bioscience

www.virtualarrays.com

Products and Technologies Visible Discovery system, instrumentation and analysis software for detection and measurement of dynamic cellular processes using a high-resolution, video microscopy device Chroma-Glo, luciferase assay system; Dual-Luciferase, reporter assay system; Rapid Response, reporter vectors; Chroma-Luc, reporter vectors that encode red and green luciferases; Monster Green, mutant GFP; HaloTag, universal reporter protein; EnduRen, substrate for luciferase assays Qdot nanocrystals; Qdot bioconjugates, encapsulated quantum dot beads prepared for bioconjugation; Qdot microspheres, biologically inert encapsulated quantum dots CellMine HCS Image informatics software Cellerity, automated system for cell growth, passaging, harvesting, plating, and counting Acumen Explorer, nonconfocal fluorescence imager for microplates CellPlex assays; CellCard System for examining multiple cell types in a single well

One of the company’s instrumentation products is the BD Pathway highcontent bioimager. Because it uses a white light confocal optical system, the BD Pathway can be used with many different ßuorescence dyes. Kinetic imaging capabilities allow measurement of intracellular free calcium. An example of this application would be distinguishing immature and mature neurons. Mature neurons display synchronized calcium oscillations, and these oscillations can be disrupted by ion-channel inhibitors. Atto’s assay products include ACT:One, a cell-based assay for G-proteincoupled receptors (GPCRs) that Atto acquired when it purchased Aptus Pharmaceuticals. A subset of GPCRs signal via cyclic AMP (cAMP), especially the

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Gi (cAMP inhibitory) class. ACT:One can directly measure changes in cAMP concentrations inside cells, which makes it particularly useful for investigating compounds that modulate Gi-coupled receptors (Vanek PG, 2004). Atto acquired Cytoprint to gain image analysis technology based on pattern recognition algorithms. The Cytoprint approach, developed initially for military applications, uses techniques that are not biased by the researcher’s assumptions about the biological processes, allowing measurement of changes in cell phenotypes. The software uses statistical methods to compare multiple images. By comparing many images, the system improves pattern recognition and reduces errors. Cytoprint technology can be used for clustering test compounds’ biological responses, grouping together compounds that produce similar phenotypic effects (Elling J, 2004). Blueshift Biotechnologies Blueshift Biotechnologies (Sunnyvale, California) is a start-up company that is developing instrumentation and consumables for cell and bead-based assays. The company’s technology (dynamic ßuorimetry) involves measurement of different ßuorescence properties: ßuorescence lifetime, anisotropy, and ßuorescence resonance energy transfer (FRET). These ßuorescence-based measurements provide information on the local environment and binding state of molecules over time. While most HCS is based on image capture and analysis, Blueshift technology measures particular features of images, which reduces data complexity and diminishes data storage and analysis challenges. This approach to cell image analysis borrows from product inspection techniques used in the semiconductor industry. One of the time-consuming tasks in HCS assay development is the need to develop image-processing algorithms for each assay. Blueshift has identiÞed 20 dimensional parameters that can be extracted from images. Examples of these dimensional parameters include color changes, color ratios, intensity measurements, geometry changes, and brightest pixel to total brightness; a speciÞc example is an increase in the cell perimeter, a parameter used in a neurite outgrowth assay. According to the company, using data reduction to measure a set of key parameters eliminates much of the data storage challenge and speeds assay development. Blueshift’s business model is to sell its laser-scanning-based instrumentation and to continually develop applications. The company plans to offer a platform that will run generic as well as proprietary assays, with beta instruments available by the end of 2004. Blueshift has collaborations with EDC Biosystems, Kendro, Parallel Synthesis Technologies, and Matrical. Cellomics Cellomics (Pittsburgh, Pennsylvania) has developed a cell-based assay platform that offers complete systems for HCS, including HCS instrumentation (Þxed end-point and kinetic systems), informatics, cellular image analysis software, ßuorescent reagents, kits, cell lines, and multiparametric assays. Part of the company’s strategy is to develop an extensive range of partnerships with companies

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341

that provide complementary technologies in order to complete its portfolio of HCS solutions. The company also offers intensive HCS training courses for researchers. The Arrayscan VTI HCS Reader is Cellomics’ Þfth-generation, CCD-based imaging instrument. When used with the Apotome, a grating imaging device from Carl Zeiss for performing optical sectioning, the ArrayScan can perform “confocal-like” optical sectioning to eliminate out-of-plane ßuorescence. Cellomics also sells reagent kits for a variety of assay types, including cell signaling, cell morphology, cell motility, toxicology, and cell cycle assays. Reagent partnerships include Molecular Probes (Alexa dyes), Ambion (RNAi assays), and Norak Biosciences (Transßuor technology). Cellomics has developed several software image analysis applications (BioApplications) for particular types of assays. Among them are the recently introduced Spot Detector BioApplication for analysis of GPCRs and receptor internalization assays and the Morphology Explorer BioApplication, which provides multiple quantitative measurements for assays such as colony formation, cell spreading, and actin and tubulin rearrangement. Cellomics’ High Content Informatics (HCi) suite provides tools for acquiring, analyzing, storing, and visualizing HCS data. Informatics partnerships include SpotFire (DecisionSite visualization software), IBM Healthcare and Life Sciences (to specify informatics hardware [HCi Appliance] for storing and managing HCS image data), and EMC (content-addressed storage). Cellomics has initiated a licensing program to broaden the availability of HCS to other companies. GE Healthcare (formerly Amersham BioSciences) and Becton, Dickinson and Company (through its BD Biosciences unit) have acquired HCS licenses from Cellomics. In addition, Cellomics and GE Healthcare are collaborating to develop an interface for GE’s cellular imaging platform to Cellomics’ HCi software. Cellumen Cellomics’ founders recently established Cellumen (Pittsburgh, Pennsylvania), a Þrm that focuses on advanced reagents for HCS using the Cellomics instrumentation platform. The company is developing novel methods to study protein–protein interactions, new classes of biosensors, and ways to manipulate cell processes using sophisticated reagents (e.g., random siRNAs). The availability of multicolor ßuorescent dyes for protein tagging will allow multiple cellular processes to be followed temporally and spatially. The company is devoting signiÞcant resources to the study of signaling pathways and intercellular cross-talk. Cellumen is also entering into collaborative discovery programs with pharmaceutical companies. Cellumen has developed proprietary technology that enables its system to follow multiple processes using the same frequency. Cyntellect Cyntellect (San Diego, California) is a start-up company that is developing laserbased instrumentation to image and manipulate cells. SpeciÞc applications of

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the company’s Laser Enabled Analysis and Processing (LEAP) technology platform include enrichment of desired cell types, injection of macromolecules such as DNA, and stimulation of reporter groups or endogenous, optically active molecules. Treatment of cells with particular types of lasers can cause cell membranes to become permeable to molecules that are ordinarily unable to penetrate the cell wall. Examples of molecules of interest for insertion are DNA, siRNA, chemical reporter groups, ions, and proteins. Quantum dots can also be inserted with this technology. While other methods to permeabilize cells (e.g., electroporation) already exist, these methods are only modestly effective for some important cell types, including neurons and B cells. The combination of manipulation and imaging allows for some interesting applications of the LEAP platform for drug discovery. For example, using siRNA to knock out certain genes permits observation of the effects on the same cells both before and after knockout. Eli Lilly is collaborating with Cyntellect on opto-transfection applications of the LEAP platform. In addition to using the LEAP platform to introduce particular molecules into cells, the laser can also be used to eliminate certain cell types: lower power permeabilizes cells, but higher power destroys them. This application is useful for collecting particular cell subtypes, such as pure T-cell phenotypes. Cyntellect is collaborating with the University of Texas to use the LEAP platform for analysis of HIV-infected T cells. The LEAP platform also has the potential to improve biomanufacturing efÞciency. Antibody-secreting cells are used to manufacture therapeutic antibodies, and high-producing clones could increase production yields. The LEAP platform allows in situ selection of the most productive clones. Cells are applied to the capture matrix, and antibody secretion is measured using a ßuorescent reagent. The laser enables elimination of low-yielding cells; the remaining cells are retrieved and expanded. Cyntellect currently has beta sites and released a commercial instrument late in the Þrst half of 2005. Molecular Devices Molecular Devices (Sunnyvale, California) develops and markets a variety of instruments, reagents, and assays for bioanalytical research. The company provides individual components as well as complete turnkey systems and kits. Molecular Devices has developed instruments for ßuorimetry, high-throughput screening, and cell-based assays and has expanded its capabilities to HCS. The company developed the Discovery-1 HCS system: a hardware–software platform for conducting a variety of HCS assays. Molecular Devices has also developed software for HCS assays, including GPCR cycling for Norak Transßuor assays, general granularity analysis, nuclei counting, and apoptosis. In July 2004, Molecular Devices acquired Axon Instruments, developer of the ImageXpress system for HCS. ImageXpress supports kinetic HCS experiments and facilitates image management, analysis, and data visualization.

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343

Many researchers conducting HCS were previously involved in traditional microscopy. Molecular Devices based its HCS software on the widely used MetaMorph image analysis program for traditional microscopy. Molecular Devices’ Discovery-1 suite of software tools for HCS allows researchers to write assayspeciÞc image analysis tools. However, the company has developed other turnkey analysis solutions that do not require researchers to write their own image analysis algorithms. Molecular Devices’ business model is to develop and market bioanalytical instruments, reagents, and kits, thus acquiring expertise in key areas. The company acquired LJL Biosystems in 2000, Universal Imaging in 2002, and Axon instruments in 2004. Among the company’s HCS customers are AstraZeneca, Cytokinetics, and Renovis. Vitra Bioscience Vitra Bioscience (Mountain View, California) is a venture-backed company that develops turnkey systems for multiplexing cell-based assays. The company uses multiple cell types in the same assay well, thereby allowing testing of compound potency and selection of drug candidates at the same time. Vitra has developed the CellCard System, which comprises the color-encoded CellCard carriers, dispensing and imaging equipment, and analysis software. CellCard carriers are rectangular particles that have a transparent section for cells and color bar code. A cell line is grown and attached to differently coded CellCard carriers; the CellCard carriers are then mixed and dispensed into microtiter plates. The plates are imaged, using the color coding on the cards to identify the cell type on the card. Vitra has developed CellPlex assays for proliferation, apoptosis, cell cycle, cytotoxicity, kinases, and GPCRs. The CellCard platform allows more controls because within each well, it incorporates both simultaneous interrogation of multiple targets and data normalization. The company is developing ways to reduce the number of cells needed for multiplexed assays by Þvefold so that more extensive compound proÞling could be done at earlier stages of development. A reduction in the number of cells needed per assay would open up the use of primary cells to HCS. Because primary cells do not grow in culture, they are expensive and difÞcult to obtain and are therefore reserved for examining late-stage compounds. One interesting application of the CellCard System is its ability to enable primary cells from many individuals or patients to be examined simultaneously in the same well, potentially offering the dramatic improvement of eliminating drug failure due to individual variation. Vitra has a collaboration with Bristol-Myers Squibb to validate the CellCard System. In a study to examine the effects of test compounds on ten different cancer cell lines, researchers studied membrane integrity, cell-cycle progression, and apoptosis and showed results consistent with running these assays individually, thus reducing assay and reagent costs.

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OUTLOOK HCS is an emerging Þeld with enormous potential to impact the drug discovery paradigm. Our survey found that more than 94% of HTS labs participating in our survey used cell-based assays in 2003, and use of these tools continues to grow. As a percentage of all assays used by our study sample, cell-based assays will represent more than 50% of throughput by 2005, continuing the trend to increasing use of cell-based assays in HTS laboratories. Of the 53 HTS laboratories surveyed, 36% expect to increase their use of cell-based assays between 2003 and 2005, 19% expect to decrease their use, and the remaining 45% anticipate that their use of cell-based assays will remain the same (Figure 5). Our study shows that cell-based assays already used in laboratories on a regular basis include translocation assays, protein trafÞcking, antisense and RNAi knockouts, apoptosis, cellular proliferation, neurite outgrowth, and simultaneous cytotoxicity and efÞcacy assays. Technologies that are driving improvements in cell-based assays and HCS include more sensitive molecular probes, better molecular probe targeting, quantum dots, improved image analysis technologies, and improved data management tools. Trends in HCS use include more kinetic assays, higher levels of multiplexing, multiplexing of different cell types, greater variety of assays, combining manipulation of cells and assays, and identiÞcation of predictive parameters. Because HCS is an emerging technology, scientists are just gaining experience in its use. As they become more experienced, scientists will begin to increase multiplexing, use kinetic assays, identify the best parameters to measure, and expand the repertoire of available assays. Examples of manipulations that can be coupled with HCS are antisense knockouts, RNAi knockouts, and gene transfection. HCS provides information on functional changes occurring at the cellular level, thus potentially providing data that are more predictive of clinical outcomes than biochemical screening. Using HCS, the potency, selectivity, and cytotoxicity of test compounds can be determined in the same assay, thereby speeding lead optimization and providing a better understanding of the many functional changes caused by compounds. HCS has relatively low throughput compared with biochemical assays and requires expensive cell handling and imaging instrumentation. Currently, HCS is reserved mostly for secondary screening applications; nevertheless, a few laboratories have begun to apply HCS to primary screening. REFERENCES Biberthaler P, et al. Evaluation of murine liver transmission electron micrographs by an innovative object-based quantitative image analysis system (Cellenger ®). European Journal of Medical Research.2003;8:275–282. Elling J, et al. Automated HCA image quality assessment. Poster Number P12004, Society for BioMolecular Screening Meeting, 2004. Elling J, et al. HCS assay discovery with novel Image clustering tools. Presentation 1604, Society for BioMolecular Screening Conference & Exhibition. Orlando, Florida, 2004.

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Hack AA, et al. Measurements of electrically paced adult cardiomyocyte populations in tissue culture. Poster number P10191, Society for BioMolecular Screening Meeting, 2004. Lippincott-Schwartz J, Patterson GH. Development and use of ßuorescent protein markers in living cells. Science. 2003;300:87–91. Stojanovic MN, Kolpashchikov DM. Modular aptameric sensors. Journal of the American Chemical Society. 2004;126:9266–9270. Vanek PG, et al. Application of a novel live-cell cAMP biosensor to GPCR drug discovery. Poster Number P04001, Society for BioMolecular Screening Conference & Exhibition, Orlando, Florida, 2004.

High-Throughput ADMET Screening: Improving the EfÞciency of Drug Discovery

SUMMARY To be effective, a drug must reach its intended target and confer a therapeutic beneÞt without causing unacceptable toxicities, so ensuring good absorption, distribution, metabolism, excretion, and toxicity (ADMET) properties is crucial to a given drug’s success. Although these properties are the subject of extensive screening—many drug candidates are eliminated as possible lead candidates during the development process because of poor ADMET properties—nearly two-thirds of drug companies do not test for ADMET characteristics in the early stages of drug discovery and development. Pharmaceutical companies are currently under tremendous pressure to reduce the cost of drug discovery and development, compress development cycle times, and increase R&D efÞciencies. Reducing expensive, late-stage, and clinical failures could have a dramatic and positive impact in all these areas, and predicting ADMET properties earlier in the drug discovery cycle will make the process more efÞcient and less expensive. In this article, we examine the current applications and tools in high-throughput ADMET screening (a process that includes predicting bioavailability, how rapidly a drug is metabolized and eliminated, possible interactions between drugs, and potential toxicities) and how these tools are used in practical ADMET screening. In addition, we present a detailed discussion of the current activities of leading players in the ADMET Þeld and describe current business models, promising opportunities, and potential technological breakthroughs. Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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INTRODUCTION

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BUSINESS IMPLICATIONS •

•

•

•

•

Good absorption, distribution, metabolism, excretion, and toxicity (ADMET) properties are crucial to a drug’s success. More than half the drug candidates that fail during the development process do so because of poor ADMET properties. Using in vitro ADMET screening methods early in discovery could reduce late-stage failures and reduce the cost and increase the efÞciency of drug development. The successful application of in vitro ADMET testing in early-stage drug discovery offers many potential beneÞts. Companies could predict the in vivo effects of drugs in humans, reduce the use of animals for drug testing, and conserve precious test compounds. A long-term goal is the development of models for predicting ADMET properties from chemical structures. Currently, 65% of pharmaceutical and biotechnology companies perform in vitro ADMET studies during or after the lead optimization stage. A critical challenge for drug developers is to reduce costs and increase throughput of in vitro ADMET tests so that they can be used to test larger numbers of compounds at earlier stages. Quantifying the beneÞts of early-stage ADMET screening is necessary to provide evidence that this approach saves development time. However, measuring the productivity of early ADMET testing is challenging. Its success will be demonstrated only by reduced compound attrition in preclinical and clinical studies. Several companies are developing innovative assays and technologies for ADMET testing. Promising technologies include stem cells for producing human hepatocytes and engineered cell lines that express drug-metabolizing enzymes and transporters. Within three to Þve years, stem-cell and cellengineering technologies will probably be mature enough to dramatically improve the predictive ability of in vitro ADMET screening.

INTRODUCTION The clinical success of a pharmaceutical compound depends on several characteristics that are associated with its physiological effects in the human body. To be efÞcacious and safe, a drug must be able to reach its intended biological target and confer a therapeutic beneÞt without causing unacceptable toxicities. To achieve these effects, the drug must be appropriately absorbed and distributed throughout the body, and it cannot be broken down or excreted too quickly or too slowly. Therefore, researchers try to predict a potential drug’s in vivo performance by measuring various properties that describe the compound’s absorption, distribution, metabolism, excretion (ADME), and toxicity (collectively, ADMET) characteristics.

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Good ADMET properties are crucial to a drug’s success. In fact, more than half the drug candidates that fail during the development process do so because they have poor ADMET properties. These failures are most costly when they occur during clinical trials. Traditionally, companies have tested ADMET properties only after a compound has demonstrated pharmacological activity and has been chemically optimized for potency. Now, companies are attempting to reduce the cost of clinical failures and increase the efÞciency of drug development by Þnding less expensive, faster methods of testing and predicting ADMET properties that can be used in the early stages of discovery. In this article, we present an overview of approaches to in vitro ADMET compound screening, discuss potential breakthrough technologies, identify potentially lucrative business opportunities, examine practical business models, and highlight the activities of leading companies in this Þeld. ADMET TERMINOLOGY ADME : Absorption, distribution, metabolism, and excretion. ADMET : ADME plus toxicity. Bioavailability: The ability of a drug to be absorbed from the GI tract, enter general circulation, and become available to the target tissue. Clearance: A measure of the body’s ability to eliminate a drug. Distribution: How a drug that has been absorbed into the bloodstream is distributed throughout various tissues and compartments in the body. Excretion: Also referred to as elimination, the process of removing compounds from the body, usually via the kidneys or the gastrointestinal (GI) tract. Metabolism: Chemical changes of drugs or environmental compounds by enzymes. Metabolism leads to changes to facilitate elimination and is accomplished by Phase I (functionalizing) enzymes and Phase II (conjugating) enzymes. Metabolism typically causes inactivation of pharmacological activity but can also lead to production of pharmacological activity of inactive prodrugs. Oral Absorption: The passage of a compound (drug) into the tissue of the GI tract and its subsequent entry into the portal vein. Pharmacokinetics: The study of the bodily absorption, distribution, metabolism, and excretion (ADME) of drugs over time. Toxicity: Adverse, damaging effects of foreign compounds on the body. Toxicity is frequently organ-speciÞc; the liver, kidney, and central nervous system are common targets.

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CURRENT PRACTICES IN ADMET SCREENING Historically, pharmaceutical companies have selected lead compounds on the basis of their biological activity, potency, selectivity, and synthetic accessibility. ADMET characteristics, particularly bioavailability, toxicity, clearance, and metabolism (see the sidebar “ADMET Terminology”), are typically evaluated later—during the lead optimization stage—and used to guide the selection of clinical candidates. Currently, 65% of pharmaceutical and biotechnology companies conducting drug development perform in vitro ADMET studies during or after the lead optimization stage, while only 35% conduct these screens prior to this stage (see Figure 1).

Cost of Screens One important reason that companies conduct ADMET testing during the lead optimization stage rather than earlier in development is cost. ADMET assays, particularly those that use human cells or tissues, can be expensive to perform—up to $150 per data point. Table 1 summarizes the highest prices that companies are willing to pay for primary high-throughput screening (HTS; assays that identify compounds with biological activity (hits) against a drug target), secondary screens (assays that conÞrm activity by a second method or test for compound selectivity), and in vitro ADMET assays. These data show that the price per data point that companies are willing to pay varies widely. Companies routinely test large compound libraries (hundreds of thousands of compounds) in HTS primary assays and use these results to select much smaller subsets of active compounds for secondary and ADMET screening. In general, researchers prefer to conduct in vitro ADMET screening after primary or secondary screening because fewer compounds need to be examined at these later stages. Companies are also more willing to pay more per data point when the number of compounds to be screened is relatively small.

FIGURE 1. Timing of in vitro ADMET screening

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TABLE 1. Highest Amount High-Throughout Screening (HTS) Directors Are Willing to Pay per Data Pointa Assay Primary screening Median Range Secondary screening Median Range ADMET screening Median Range

Price per Data Point ($US) 0.50 0.05–5.00 1.25 0.15–100.00 1.00 0.50–150.00

a Based on data from 51 directors of HTS laboratories

worldwide.

Throughput and Speed Another reason companies perform ADMET screening later in the discovery process is that most in vitro ADMET screens are relatively low-throughput. When ADMET tests are conducted during or after lead optimization, the number of compounds that need to be screened is low, and achieving high-throughput levels is not critical. However, as companies move ADMET testing to the earlier stages of discovery, many more compounds must be screened. To meet this demand, companies conducting early-stage ADMET screening will need to both convert existing ADMET assays to higher-throughput formats and develop new, faster assays. Increasing the throughput of ADMET assays is one of the premier challenges facing pharmaceutical companies seeking to push ADMET screening upstream in the discovery process. Many companies are using automation to achieve this goal. Still, few companies have succeeded in accelerating ADMET assays to the levels of throughput achieved in primary activity assays. Other Factors Companies have avoided early-stage ADMET screening for other reasons as well. For example, compound libraries change frequently, and keeping up with these changes may not be cost-effective. Also, some groups resist early ADMET screening because poor ADMET properties can be modiÞed later in development using medicinal chemistry. THE CASE FOR EARLY-STAGE, HIGH-THROUGHPUT ADMET SCREENING Pharmaceutical companies are under tremendous pressure to reduce the cost of drug discovery and development, compress development cycle times, and increase

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TABLE 2. Benefits of Conducting In Vitro ADMET Assays in Early-Stage Discovery Benefit Predict drug–drug interactions via induction/inhibition studies. Identify which enzymes are responsible for a drug’s metabolism. Evaluate human pharmacokinetics. Study species-specific metabolism and toxicity. Reduce the use of test animals.

Decrease product development time. Use smaller amounts of test compounds. Develop computational models to predict ADMET properties.

Approach/Technique Determine which enzymes and isoforms are involved in metabolism and measure their expression levels. Perform assays using recombinant drug-metabolizing enzymes. Perform assays using human cells or subcellular fractions. Compare different animal models to human response. Improve the predictive ability of cell and tissue systems and use them as alternatives to animal testing. Identify problematic issues and eliminate poor drug candidates early, thereby reducing need for further medicinal chemistry optimization and formulation. Convert assays to smaller scale with microfluidics or microtiter plates. Create databases that correlate chemical structures, ADMET properties, and outcomes in humans and animal models; develop software tools to predict these outcomes based on chemical structures.

R&D efÞciencies. Reducing expensive, late-stage, and clinical failures could have a dramatic and positive impact in all these areas. Eliminating compounds with poor ADMET properties early in the development cycle can help companies avoid signiÞcant expenditures on compounds that will fail during or after lead optimization. Early-stage ADMET testing offers a variety of other potential beneÞts to drug developers (see Table 2). For example, companies could predict the in vivo effects of drugs in humans, reduce their use of animals for drug testing, and, by using smaller quantities for testing, conserve precious test compounds. A longterm goal of widespread ADMET proÞling is the development of models that can accurately predict particular aspects of ADMET properties, especially absorption, metabolism, and toxicity, from chemical structures alone. Computationally screening an entire compound collection is less expensive than using biochemical or cell-based assays. Despite an incomplete understanding of the rules that predict ADMET properties, some companies are developing and/or conducting computational screens. As mentioned previously, some companies do not perform early-stage ADMET screening because chemists can modify poor ADMET properties later in the drug development process. However, using medicinal chemistry to alter ADMET properties is frequently difÞcult—it is empirical, slow, and expensive. In contrast, with sufÞcient knowledge about a speciÞc drug target, the medicinal chemistry to modify a compound’s potency and selectivity may be easier and more efÞcient

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to optimize than ADMET properties. Therefore, researchers may be able to effectively optimize compounds for potency and selectivity after screening them for ADMET properties. Regulatory agencies are also providing an impetus for implementing in vitro ADMET screening on new chemical entities (NCEs). In a 1997 FDA publication, the agency encouraged routine in vitro evaluation of metabolism and drug–drug interactions whenever feasible (Guidance for Industry, 1997). NO CONSENSUS ON STRATEGIES IN ADMET SCREENING Having recognized the importance of ADMET properties to the success of drug candidates, many pharmaceutical companies are changing their deÞnition of a lead compound from one that is efÞcacious, potent, and selective to one that also has acceptable ADMET properties. Despite this paradigm shift, no true consensus exists among pharmaceutical companies on the best strategy for determining these characteristics. Complicating matters further, different divisions within the same company often have different ADMET testing strategies. A few pharmaceutical companies have conducted librarywide proÞling for certain ADMET properties. Others do not see the value in eliminating compounds with undesirable ADMET characteristics before conducting a primary HTS, mainly because chemists can modify poor ADMET properties during the lead optimization stage. Even companies that do believe that early-stage ADMET screening provides signiÞcant beneÞts are resisting this approach because current techniques are not yet fast enough or cheap enough to justify their widespread implementation. As an alternative to librarywide compound proÞling, some companies opt to study in detail small numbers of compounds that researchers believe to be representative of a chemical class. APPLICATIONS AND TOOLS IN HIGH-THROUGHPUT ADMET SCREENING In this section, we review the primary applications of high-throughput ADMET screening and describe some of the assays and tools commonly used in these screens. Table 3 summarizes these and other tools used to evaluate a wide range of ADMET properties. Predicting Bioavailability Bioavailability and oral absorption are usually among the Þrst parameters to be assessed, both because these parameters are important to a drug’s success and because these screens are relatively simple to conduct. Current screening approaches include in vitro cell culture assays and cell-free methods. These assays typically evaluate a cell or cell membrane’s permeability to speciÞc test compounds and are therefore useful for predicting in vivo absorption and bioavailability.

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TABLE 3. Assays and Tools for Predicting ADMET Properties Property Bioavailability (oral absorption)

Blood–brain barrier permeability Cardiotoxicity Dermal toxicology Drug-drug interactions

Drug resistance to chemotherapy, protease inhibitors, or other drugs Endocrine disruption Excretion (Phase II metabolism enzymes)

Hepatotoxicity Immunotoxicity Metabolism (Phase I metabolism enzymes)

Metabolism of amine drugs Mutagenicity, teratogenicity Neurotoxicity Physicochemical properties (e.g., lipophilicity, solubility, pKa ) Serum protein binding Toxicity

Assays/Tools In vitro cell culture models include CaCo-2 cells, MDCK cells, microvessel endothelial cells; cell-free assays include artificial cell membranes, PAMPA MDCK cells, microvessel endothelial cells, modified PAMPA hERG potassium channelsa by patch clamp or engineered cells with fluorescent reporter Normal human-derived epidermal keratinocytes Assays with isoforms of recombinant drug-metabolizing enzymes, antibodies to isoforms, cell lines engineered with inducers of CYP expression (PXR, CAR), measurement of CYP mRNA in primary hepatocytes Transporter proteins that actively expel drugs from cells

Engineered cells expressing human estrogen receptor UDP glucuronyl transferase (UGT), sulfotransferase (SULT), glutathione S-transferase (GST), N-acetyltransferase, histamine methyltransferase, catechol-O-methyl transferase, thiopurine methyl transferase MTT or ATP assay in hepatocytes Lymphocyte inhibition Microsomes, S9, hepatocytes, HepG2 (hepatoblastoma carcinoma-derived cells), recombinant or purified enzymes (P450, esterases, epoxide hydrolase, alcohol dehydrogenase, aldehyde dehydrogenase, dihydropyrimidine dehydrogenase, NADPH quinone oxidoreductase) Monoamine oxidase Whole embryos Brain slices Log P, log D, pKa , aqueous solubility Human serum albumin binding assays Cytotoxicity analysis in mammalian cells

a A potassium channel blockage of the hERG channel is associated with the prolongation of the QT interval

and cardiac death. Acronyms: ATP, adenosine triphosphate; CAR, constitutive androstane receptor; hERG, human eag-related gene; MDCK, Madin–Darby canine kidney; PAMPA, parallel artificial membrane permeation assay; PXR, pregnane X receptor.

In Vitro Cell Culture Assays. Researchers have developed a variety of in vitro cell culture models to assess a compound’s ability to penetrate cells in the intestine, on the skin, or at the blood–brain barrier. In these assays, speciÞc types of cells are grown on a porous membrane that separates two chambers. Compounds are added to one chamber and incubated with the cells. A compound’s ability to penetrate the cellular barrier is then assessed by various

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analytic techniques, including liquid chromatography (LC), LC/mass spectrometry (LC/MS), or ultraviolet (UV) absorption. CaCo-2 cells are human cells derived from colon adenocarcinoma and are widely used in assays to evaluate oral absorption and bioavailability. Because multiple physiological processes occur in CaCo-2 cells, they permit a more complete biological evaluation of absorption processes than do most other in vitro assays. These cells can be used to study cell permeability, passive diffusion, active drug transport, drug efßux, and drug metabolism. Limitations of CaCo-2 cells include a relatively long culture time (the cells typically require 21 days in culture), lack of mucus (which is normally present in the intestine), transport properties that are representative of the colon rather than the small intestine (where most drug absorption takes place), and the immortalized nature of this cancer cell line. Other useful cell types for evaluating permeability include Madin–Darby canine kidney (MDCK) cells and microvessel endothelial (blood vessel) cells. MDCK cells, which possess permeability to passively absorbed compounds similar to that of CaCo-2 cells, can be used after only three days in culture and have low expression of drug transporter proteins. Brain microvessel endothelial (blood vessel) cells are used to evaluate permeability at the blood–brain barrier. Gentest has created engineered cells by introducing the gene for the CYP3A4 (cytochrome P450 3A4) enzyme into CaCo-2, MDCK, and porcine kidney cells. These modiÞed cells allow researchers to test cell permeability and intestinal metabolism in the same system. Cell-Free Assays. Cell-free approaches to assessing absorption include the use of immobilized artiÞcial membrane (IAM) chromatography or the parallel artiÞcial membrane permeation assay (PAMPA). Both of these techniques are relatively rapid compared to cell assays and are widely used to measure passive absorption across a cell membrane. In the IAM system, an artiÞcial membrane containing cell membrane phospholipids is bonded to a solid chromatographic support. In the PAMPA system, the artiÞcial cell membrane is simulated by phospholipid bilayers in microÞlter channels of a 96-well plate. To estimate a compound’s membrane permeability, researchers monitor drug transport in the system using high-performance liquid chromatography (HPLC) or LC/MS. Researchers have modiÞed the original PAMPA phospholipid system with a variety of natural lipids to make it more similar to particular tissues, such as intestinal or brain tissues. Predicting Drug Metabolism Metabolic stability is the most important factor that determines whether a lead compound progresses to clinical development. Drug metabolism may produce either pharmacologically active or inactive metabolites. If a drug’s metabolism to an inactive form is too rapid, the drug’s efÞcacy is reduced. If metabolism to an inactive form is too slow, the drug can accumulate over time, and this accumulation can

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lead to toxicity. Drug metabolism can also yield toxic metabolites that can damage organs and tissues. Finally, compounds may either induce or inhibit the activity of metabolizing enzymes, events that can affect the drug’s metabolism or result in undesirable drug–drug interactions (see “Predicting Drug–Drug Interactions”). Before beginning clinical trials, a detailed understanding of the metabolic pathways and kinetics of a drug’s metabolism is desirable. Various cell-based and biochemical screens are available to predict a drug’s in vivo metabolism. These screens enable researchers to rank compounds that have similar efÞcacy and determine which candidates should progress to clinical development. Assay systems for measuring metabolism include recombinant enzyme assays and cellular assays. Cellular assays may be based on isolated hepatocytes (liver cells), liver slices, or subcellular fractions. Cellular systems contain the full range of drug-metabolizing enzymes in their native conÞgurations. Having a full complement of enzymes in a single assay system is useful because different enzymes, such as Phase I and Phase II enzymes, may work together to increase the rate of a drug’s metabolism. Recombinant Enzyme Assays. Enzymes involved in various stages of metabolism can be used as components in assays to predict in vivo drug metabolism. Recombinant, puriÞed forms of many of these enzymes, including cytochrome P450 (P450) enzymes (see the sidebar “Cytochrome P450 Enzymes”), glutathione S-transferases (GSTs), sulfotransferases (SULTs), and uridine diphosphate-glucuronosyltransferase (UGT), are commercially available. Recombinant enzymes can help researchers determine a compound’s metabolic stability. Testing compounds against a panel of enzymes also provides information on selectivity and enables the identiÞcation of the particular enzymes responsible for a compound’s metabolism. CYTOCHROME P450 ENZYMES Cytochrome P450 (P450) enzymes are found in the endoplasmic reticulum of the liver, small intestine, lungs, kidneys, blood vessels, and almost all other cells. Members of this enzyme superfamily (referred to as the CYP family) catalyze the oxidative metabolism of structurally diverse chemicals. The CYP gene family produces more than 50 different P450 enzymes responsible for the metabolism of xenobiotics (foreign substances that include any drugs and other chemicals) that are ingested, inhaled, or absorbed through the skin. Six P450 enzymes account for the metabolism of nearly all clinically important drugs. Two, CYP3A4 and CYP2D6, are responsible for metabolizing about 80% of all drugs. CYP3A4 metabolizes antihistamines, antibiotics, statins, antihypertensives, protease inhibitors, and antifungals. CYP2D6 metabolizes selective serotonin reuptake inhibitors (SSRIs), analgesics, beta blockers, and others. An advantage of recombinant enzyme assays is that their results are more reproducible than those from assays performed using biologically derived material (e.g., cells). Because the results from recombinant assays help researchers

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identify which chemical structures on a compound are metabolized, they also suggest how compounds might be modiÞed to prevent unwanted metabolism, so recombinant assays are used frequently during lead optimization. The main disadvantage of these isolated systems is their inability to truly mimic in vivo complexity or take into account the effects of other interactions and processes. For example, isolated recombinant enzyme assays do not provide insight into the interactions between multiple metabolizing enzymes and excretion mechanisms. Cell-Based Assays. Hepatocytes are isolated from liver tissue and may be used as primary cells or as cell cultures in drug metabolism assays. Isolated hepatocytes maintain the integrity of multi-enzyme complexes and can be used to study both Phase I and Phase II metabolism. However, the expression of some drug transporter enzymes is downregulated after the hepatocytes are isolated from the liver tissue. The support matrices used in assay systems may also reduce normal enzyme activity in these cells. Alternatively, intact liver slices may be used in drug metabolism assays. These tissue slices maintain cellular integrity but cannot be frozen or easily stored. Subcellular fractions prepared from human liver tissue for use in metabolism assays include microsomes and S9 fractions. They are commercially available and contain most of the CYP enzymes involved in drug metabolism. These nontransformed primary cells have the advantage of more closely representing true metabolism than transformed, immortalized cell lines but are difÞcult to use and very expensive. Despite its utility for drug metabolism assays, human liver tissue is difÞcult to obtain. It must be harvested from cadavers and is only available sporadically. Donated livers are preferentially used for transplantation, so only those organs unsuitable for transplant can be used as sources for tissue. Furthermore, handling human tissue requires additional precautions because it may be a source of pathogens. Predicting Drug–Drug Interactions Predicting interactions between different drugs is an important task in ADMET assessment. Multiple-drug therapy is now the normal clinical situation, and speciÞc drug combinations can lead to interactions that could increase or decrease therapeutic efÞcacy or produce an adverse reaction. These interactions may affect a compound’s efÞcacy or toxicity by altering ADMET properties or by causing antagonistic or agonistic interactions at a drug receptor. Adverse reactions, including fatalities, due to drug–drug interactions have forced several drugs off the market. Therefore, understanding these interactions at the earliest stages of drug development is advantageous. Drug–drug interactions occur either by inhibition or induction of metabolism enzymes. Inhibition of a particular enzyme can reduce the rates of metabolism of other drugs that are metabolized by the same enzyme. Enzyme inhibition reactions can be rapidly reversible, slowly reversible, or irreversible. Inhibition of drug-metabolizing enzymes can also cause a drug to accumulate to toxic levels.

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Alternatively, inhibition may increase a drug’s bioavailability due to its extended half-life. In some cases, inhibition has no observable effects on drug metabolism. Enzyme induction is an adaptive response that protects organisms against chemical exposure and is the basis for a speciÞc class of drug–drug interactions. During induction, a compound induces the expression of (and thereby the concentration of) metabolizing enzymes, particularly the CYP enzymes. Higher CYP levels can accelerate the metabolism of other drugs. Induction can therefore reduce the efÞcacy of many drugs by increasing their metabolism and causing their concentrations to fall below therapeutic levels. Researchers use assays that evaluate the extent of metabolism by CYP enzymes to predict potential drug–drug interactions. Inhibitors of speciÞc P450 enzymes, antibodies speciÞc for different P450 isoforms (isozymes), and recombinant drugmetabolizing enzymes are useful tools in these evaluations. CYP induction assays can also be run in primary human or animal hepatocytes by measuring the expression of speciÞc CYP mRNAs. Researchers have also created engineered cell lines that express DNA transcription factors that are activated by certain types of compounds and regulate the expression of speciÞc CYP enzymes. These transcription factors include the pregnane X receptor (PXR), a nuclear receptor that is activated by many compounds (including glucocorticoids, antibiotics, and antifungals) and regulates CYP3A expression. Another nuclear receptor, the constitutive androstane receptor (CAR), is activated by various compounds and regulates the expression of CYP2B. (For more information on these receptors, see the report entitled “Orphan Nuclear Receptors” (2002).) High-throughput binding assays based on cell lines expressing PXR and CAR enable researchers to identify compounds that bind these receptors and that may be involved in undesirable drug–drug interactions. Puracyp (San Diego, California) has developed assays for CYP induction using human hepatoma cells that express a given CYP gene coupled to luciferase and the human PXR. Luciferase is a luminescent protein that can be easily detected using a luminometer. The assay is run in 96-well microtiter plates and can be used to rank compounds according to their ability to induce CYP isoforms. Predicting Specific Toxicities Toxicity can take many forms and can arise for many different reasons. Toxic effects may be either general or speciÞc to particular organs and tissues, such as the endocrine system, liver, nerves, immune cells, or kidney. Determining whether a lead compound’s toxicity is within acceptable limits is an important step in drug development. Cell-Based Assays. Automated cell-based assays of toxicity generally involve measurement of cell death, either by necrosis or apoptosis (programmed cell death). Assays for performing routine compound toxicity testing include MTT assays, lactate dehydrogenase (LDH) measurements, and cell viability tests. Several methods assess membrane integrity, a hallmark of active, viable cells. These

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methods include assays based on exclusion of the dye trypan blue, incorporation of radiolabeled nucleotides (tritiated thymidine or brominated uridine), and leakage of LDH from cells into extracellular matrix. Another method of assessing drug toxicity in cells is to measure changes in the ratio of the adenylate nucleotides adenosine diphosphate (ADP) and adenosine triphosphate (ATP). This approach can be used to detect and measure compromised mitochondrial function due to apoptosis, necrosis, and cell proliferation. When intracellular ATP concentrations fall to a level where the cell can no longer carry out basic metabolic functions, the cell will die. Measurement of the ADP/ATP ratio can be used to differentiate between the different types of cell death. In primary necrosis, ATP levels fall rapidly, while secondary necrosis occurs after induction of apoptosis and causes an intermediate drop in ATP levels. At the onset of apoptosis, intracellular ADP increases without an appreciable drop in ATP. Researchers can also use human bone marrow cells in assays to assess hematopoietic toxicities of drugs and help establish maximum tolerated dose levels for Phase I clinical trials. Limitations of this approach are access to fresh human bone marrow and variability among samples. Zebrafish. An emerging and innovative method for predicting the toxic effects of compounds involves the use of zebraÞsh (Danio rerio). These Þsh offer a potential method of high-throughput toxicity screening on whole organisms. Because they are vertebrates, zebraÞsh are more closely related to humans than are other commonly used model organisms (e.g., yeast, nematode worms, fruit ßies). Many zebraÞsh genes show a high degree of structural and functional similarity to their human homologues. Recently, researchers have characterized several features of zebraÞsh development, including early embryonic patterning, early development of the nervous system, and aspects of cell fate and lineage determination, and this understanding is proving useful in designing assays to evaluate the biological effects of test compounds. ZebraÞsh embryos are easily obtainable in large numbers, are transparent, and undergo rapid organogenesis. These features make zebraÞsh very amenable to compound screening in various toxicity assays. In situ hybridization techniques can be applied to monitor changes in the expression of zebraÞsh genes in response to drugs. Using subtractive techniques, researchers can identify genes and pathways involved in organ toxicity responses. Other zebraÞsh toxicity assays include assessments for lethality, embryo survival, behavior, and the microscopic examination of organ malformation. Phylonix has developed zebraÞsh-based assays for neurological, hepatic, reproductive, and developmental toxicity (see “Company ProÞles”). Stem Cells: An Emerging Tool for ADMET Screening Stem cells could provide a constant source of human primary cells that would be less expensive than those sourced from human donors. However, a method to

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produce hepatocytes reliably has not yet been achieved. Companies working on this problem include StemCells and VistaGen Therapeutics. The advantage of using hematopoietic progenitor or precursor cells is that every type of blood cell to which the precursors differentiate can be examined. Hematopoietic stem cells are derived from bone marrow or umbilical cord blood and are the least-differentiated cells. Stem cells differentiate into progenitor cells, then into precursor cells, and Þnally into mature cells. Stem cells, progenitor cells, and precursor cells are more susceptible to xenobiotic toxicity than terminally differentiated cells. Dose-limiting toxicity of drugs, especially anticancer drugs, is most commonly due to the susceptibility of bone marrow and epithelial mucosa of the gastrointestinal tract. Both of these tissues are continuously proliferating in adults and are therefore susceptible to destruction by drugs that target rapidly growing cells. Thus, identifying toxicity to these cells early in the discovery cycle is important. BUSINESS MODELS FOR HIGH-THROUGHPUT ADMET Growing demand for early-stage ADMET testing supports a burgeoning industry focused on developing and supplying tools and services for HTS. In this section, we describe several general business models that characterize the players in this Þeld. Several of these companies are also highlighted in the “Company ProÞles” section. AMDET-Focused Service Companies ADMET-focused companies offer fee-for-service screening or provide reagents or equipment for ADMET studies. Companies in this category are not pursuing internal drug discovery programs. Firms in this sector include Absorption Systems, Harvard Bioscience, Cyprotex, MB Research Laboratories, and HemoGenix. Experimental Model Companies Some companies are developing new experimental models that can be used for both ADMET and efÞcacy studies. Most of these companies offer these tools for sale as reagents or use them to perform screening services for clients. Among these Þrms are Solvo Biotechnology, Nimbus Biotechnology, MatTek, Phylonix, and Puracyp. Full-Service Discovery Services and Reagent Providers The advantages of offering a full range of services combined with economies of scale have encouraged many service and tool providers to consolidate by merging with or acquiring other vendors. Now, many companies that provide services and reagents for drug discovery and biomedical research also provide tools for some aspects of ADMET evaluation. These Þrms include Albany Molecular Research, Array BioPharma, BD Biosciences (BD Gentest), Cambrex, Cardinal Health, Cerep, MDSPharma Services, and Novascreen Biosciences.

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Service and Internal Discovery Companies Some companies with enabling technologies that can be used for ADMET screening as well as drug discovery are conducting internal discovery programs. These Þrms are also seeking to partner their drug discovery programs with pharmaceutical and biotechnology companies. Companies using this business model tend to have the most innovative technologies for ADMET prediction. Firms in this category include Amphioxus Cell Technologies and VistaGen Therapeutics. COMPANY PROFILES In the following sections, we proÞle select companies that are active in the Þeld of ADMET screening. These companies are representative of most of the major business models for companies involved in high-throughput ADMET. See Table 4 for a listing of these and other companies with ADMET technology. Absorption Systems Absorption Systems (Exton, Pennsylvania) is a private contract-research company that provides ADME assessment and formulation development services for pharmaceutical compounds. The company also provides consulting services to advise clients on approaches to ADME testing. ADME proÞling services offered by Absorption Systems include determinations of log D, solubility, chemical stability, permeability, cytotoxicity, metabolic stability, CYP inhibition and induction, metabolite identiÞcation, and blood–brain barrier transport. The company is also developing ex vivoassays for permeability with human intestinal epithelium cells obtained from cadavers. Amphioxus Cell Technologies Amphioxus Cell Technologies (Houston, Texas), a venture-backed company that specializes in liver metabolism, has developed the ACTIVTox cell-based assay system for high-throughput ADMET testing. Cell-based testing systems have several advantages over those based on primary hepatocytes. Cell lines are less expensive than primary liver cells, and cell-based systems are more convenient and yield results that are more consistent than those produced by primary liver cells. The ACTIVTox assay is based on an immortalized human liver cell line, C3A, which is derived from the widely used HepG2 cell line. C3A has a stable phenotype and properties of a normal human hepatocyte (including glucogenesis, albumin production, and drug metabolism). The C3A cell line expresses the genes for major CYP enzymes involved in drug metabolism: 1A2, 2B6, 2C9, 2C19, 2D6, 2E1, and 3A. Amphioxus claims that induction of these enzymes occurs in the cell line in response to known inducers. C3A cells also express P-glycoprotein, which is the product of the multidrug-resistance (MDR1 ) gene. In the ACTIVTox system, C3A cells are grown in microtiter plates and can be used to screen compound libraries for

TABLE 4. Selected Companies Active in ADMET Screening

Company Absorption Systems Albany Molecular Research Amphioxus Cell Technologies Array Biopharma BD Gentest (part of BD Biosciences) Cambrex Cardinal Health Cerep Cyprotex Harvard BioScience HemoGenix In Vitro Technologies

Web Site

ADMET-Related Technologies, Products, or Services

www.absorption.com www.albmolecular.com

ADME profiling assays Metabolic screening assays, P450 analysis

www.amphioxus.com

Cultured hepatocytes for toxicity and metabolism screening

www.arraybiopharma.com www.gentest.com

Drug metabolism assays Cell-based assays, engineered cell lines, recombinant drug-metabolizing enzymes

www.cambrex.com www.cardinal.com/pd www.cerep.fr www.cyprotex.co.uk www.harvardbioscience.com www.hemogenix.com www.invitrotech.com

Human and animal cells, including stem cells, for screening High-throughput assays for drug metabolism, permeability, solubility, and toxicology Assays for drug metabolism, solubility, and permeability High-throughput ADME screening, in silico prediction of pharmacokinetics Equipment, reagents, and kits for screening Hematopoietic stem-cell testing for hemotoxicity Human cells and cell lines, subcellular fractions, drug metabolism, drug interactions, absorption, and toxicity screening Tissue models for safety, drug delivery, and efficacy testing In vitro and ex vivotoxicology testing Drug metabolism assays Transil technology for physicochemical screening and functional protein assays for high-throughput screening Screening service for safety and efficacy testing, cell-based and biochemical high-throughput assays Gene expression analysis, cDNA microarrays, toxicogenomics databases

MatTek MB Research Laboratories MDS Pharma Services Nimbus Biotechnology

www.mattek.com www.mbresearch.com www.mdsps.com www.nimbus-biotech.com

Novascreen Biosciences

www.novascreen.com

Phase-1 Molecular Toxicology

www.phase1tox.com

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362 TABLE 4. (continued)

Company

Web Site

Phylonix Promega pION Puracyp

www.phylonix.com www.promega.com www.pion-inc.com www.puracyp.com

Regis Technologies

www.registech.com

Solvo Biotechnology StemCells Tecan Tissue Transformation Technologies VistaGen Therapeutics

www.solvobiotech.com www.stemcellsinc.com www.tecan.com http://www.tcubedinc.com/

XenoTech

www.xenotechllc.com

www.vistagen-inc.com

ADMET-Related Technologies, Products, or Services Zebrafish for toxicity and drug activity screening Assay reagents and kits. Tools for solubility, dissolution, permeability, and absorption screening Antisense probes of CYP cDNA, antibodies to human P450s, service to measure P450 induction by high-throughput screening Chromatography, including immobilized artificial-membrane chromatography columns for measuring drug–membrane permeability Whole-cell and membrane-based assays for ATP-coupled drug transporters Liver and neural stem cells. LabCD microfluidics platform for ADMET screening Human cells and subcellular fractions Pluripotent and tissue-specific stem cells for toxicity and efficacy screening; gene expression profiling Drug metabolism assays, drug interaction studies, early- enzyme-induction service, hepatic and subcellular fractions, hepatocytes, isoform-specific CYP antibodies, recombinant CYP enzymes, and branched DNA (bDNA) probes

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P450 induction and inhibition, liver metabolism, and toxicity. Amphioxus recommends multiparameter testing using four toxicity parameters: inhibition of cell proliferation, lactate dehydrogenase (LDH) release, ATP depletion, and caspase activation. In addition to providing its assay products and contract services to clients, Amphioxus is applying its expertise in liver biology and its proprietary cell line to internal discovery programs. Amphioxus is currently working with several pharmaceutical and biotechnology companies to provide early-stage structure–toxicity data for early-stage lead optimization and selection. Cyprotex Cyprotex (Manchester, United Kingdom) provides information and services for pharmacokinetic and ADME analysis and has detailed information on in vitro ADME and in vivo pharmacokinetics for drug discovery compounds. The company has established high-throughput ADME screening assays that it claims can screen up to 500,000 compounds per year. Cyprotex has also developed simulation software for predicting pharmacokinetics in humans and rats based on experimental data. Cyprotex has drug discovery support agreements with Astex, Roche, and AstraZeneca. HemoGenix HemoGenix (Columbia, South Carolina) is a privately held contract research Þrm that specializes in stem-cell hemotoxicity testing. The company has developed more than 200 assays and procedures for evaluating the effect of compounds on the blood-forming system. HemoGenix has developed the Hemotoxicity Assays via Luminescence Output (HALO), which is an in vitro, high-throughput hemotoxicity testing system that uses luminescence detection technology to determine the toxic effects of compounds on different cell types. The company’s OxyFlow system measures oxidative DNA damage in human and animal blood cells by ßow cytometry. In addition to its screening services, HemoGenix supplies cell and tissue culture products. In Vitro Technologies Privately held In Vitro Technologies (Baltimore, Maryland) offers products and services for ADMET screening. The company markets human and animal hepatocytes, tissue slices, subcellular fractions, and CaCo-2 cells, and it offers a variety of services, including metabolite analysis, intestinal absorption, drug metabolism, induction and inhibition, toxicity, skin absorption, and target organ toxicity. In Vitro has adapted various in vitro screening assays to higher-throughput formats. One assay the company has developed is a 96-well assay (coupled to LC/MS analysis) for metabolism screening in primary human hepatocytes. Other screens include assays for intestinal absorption in CaCo-2 cells and toxicity assays in human hepatocytes.

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MatTek MatTek (Ashland, Massachusetts) develops and manufactures human cell-based tissue models for testing the efÞcacy and safety of drugs and cosmetics. These three-dimensional models are structures that resemble human epithelial tissues, including skin, cornea, and respiratory epithelial tissue. MatTek’s EpiDerm human epidermal model consists of cultured normal, human-derived epidermal keratinocytes (NHEK) and can be used for transdermal drug delivery studies, skin irritancy testing, and toxicology testing. The EpiOcular model consists of cultured NHEK cells forming tissue similar to that found in the cornea and can be used in irritancy and cytotoxicity testing. EpiAirway consists of human tracheal or bronchial epithelial cells and is useful for inhaled drug delivery studies, as well as for inßammation, irritancy, and toxicity testing. MatTek is also developing an immunocompetent skin model, a vaginal epithelial model, and a buccal model. Nimbus Biotechnology Nimbus Biotechnology (Leipzig, Germany) is developing automated systems for conducting in vitro pharmacokinetic studies and transmembrane protein target screening. The underlying technology, Transil, involves the attachment of lipids and proteins onto beads. Transil beads can be distributed in microtiter plates (96 or 384 wells) and used in high-throughput assays for determining a compound’s lipophilicity (afÞnity to lipids, e.g., lipid membranes) or its tendency to bind to serum proteins (such as albumin). According to Nimbus, Transil technology offers a less expensive alternative to CaCo-2 cell-based testing for predicting intestinal absorption and uptake into target organs. In addition to physicochemical screening, Nimbus has developed Transil for functional high-throughput assays of membrane-bound proteins, such as ion channels and transporters. Phylonix Phylonix Pharmaceuticals (Cambridge, Massachusetts) is a contract research Þrm developing drug activity and toxicity screening assays using live zebraÞsh (see “Applications and Tools in High-Throughput ADMET Screening”). In addition to toxicity and pathology services, the company has cellular assays for detecting reactive oxygen, caspase activity, and cell cycle inhibition; behavioral studies for evaluating neural defects; and gene expression studies. Phylonix has also developed microtiter plate-based assays for compound screening, including a ßuorescent staining method to assess in vivo apoptosis and a screen for angiogenesis. pION pION (Woburn, Massachusetts) develops and markets instruments and provides services for predicting ADME properties, including solubility, dissolution, permeability, and charge state. The company offers high-throughput analyzers for

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permeability and solubility assessment, the PAMPA Explorer for entry-level permeability analysis, and pSOL for solubility assessment. The company also distributes a line of pKa , log P , and log D analyzers. StemCells StemCells (Palo Alto, California) is attempting to develop human stem cells for several therapeutic applications, including the treatment of central nervous system, liver, and pancreatic disorders. The company has identiÞed the human neural (brain) stem cell and a candidate population of human liver engrafting cells. StemCells has also developed a novel in vitro culture assay that allows these highly enriched populations of cells to grow and mature to albumin-producing hepatocytes. If they can be produced in sufÞcient quantities, these cells may eventually be useful in generating hepatocytes for metabolism and toxicity screening. Tecan Tecan (M¨annedorf, Switzerland) is leveraging its expertise and its penetration into the HTS market to develop and commercialize ADMET-speciÞc devices and applications. Tecan is developing the LabCD, a microßuidics platform (purchased from Gamera Bioscience) for performing sample separations and assays. Tests are performed in a microßuidics disc, a disposable CD-like platter that contains tiny channels, reactions chambers, and valves. Tecan’s system spins the disc, and centrifugal force and capillary action propel sample ßuids and reagents throughout the disc and achieve sample mixing and dilution. Other system components detect and process the results. Tecan has developed P450 inhibition and serum-protein-binding assays for the LabCD. The company initiated a technology early-access program for LabCD in December 2001 and plans to develop additional ADMET applications. In addition to the LabCD technology, Tecan has adapted its Genesis workstations for cell permeability and drug metabolism assays. Tecan also has assay systems that run on its Genesis HTS workstation for mutagenicity testing, automated cell culture, log P and log D determination, and toxicity testing. VistaGen Therapeutics VistaGen Therapeutics (Burlingame, California) is a drug discovery company developing products to treat epilepsy and other disorders of the central nervous system. The company is developing embryonic and nonembryonic murine stemcell lines for use in drug efÞcacy and toxicity assays and for functional genomics applications. The company uses differentiated pluripotent stem cells to produce test mixtures of different mammalian tissues (including blood, blood vessels, bone, muscle, and nerve tissues) in the same culture. VistaGen uses these tissues in screens to evaluate the efÞcacy and safety of drugs for a wide range of tissues and organ systems. The company’s screens can also be used to evaluate drug metabolism and to identify factors that affect neighboring tissues.

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VistaGen has formed collaborations with Novartis, PÞzer, and GlaxoSmithKline to evaluate its murine stem-cell-based Genesis Screen technology for use in the discovery-phase assessment of new-drug-candidate toxicity and efÞcacy. VistaGen also has a collaboration with chemistry and informatics company Tripos (St. Louis, Missouri) to develop algorithms to analyze data in VistaGen’s in vitro gene expression and biomarker screening assays. XenoTech XenoTech (Lenexa, Kansas) provides products and contract-research services for assessing various ADMET properties. The company’s products include hepatic and subcellular fractions, hepatocytes, antibodies against CYP enzymes, recombinant CYP enzymes, and branched DNA (bDNA) probes. XenoTech’s isoform-speciÞc antibodies and bDNA probes are useful for determining which enzyme isoforms are involved in metabolism. OUTLOOK Over the next several years, pharmaceutical and biotechnology companies will continue to develop high-throughput ADMET screening technologies and integrate them into the earlier stages of drug discovery and development. Although the ability to accurately predict a drug’s in vivo ADMET properties from in vitro screening is not yet possible, in vitro ADMET studies are frequently sufÞcient to determine major metabolic pathways, identify potential drug–drug interactions, and predict bioavailability. Similarly, although in vitro experiments are not completely predictive of clinical outcome, these tests can help guide clinical trials, especially in the testing of drug–drug interactions and different drug-metabolizing phenotypes. As we have discussed throughout this report, high-throughput in vitro ADMET screening technology is still in the early stages of development. Advances in many areas will be required before these tests can be widely applied and accurately predict human responses to drugs. The need to develop and improve these tests will provide many business opportunities for companies developing ADMET screening technology. Table 5 lists speciÞc areas of need. Most development efforts are currently focused on miniaturizing, automating, and reducing the cost of existing ADMET assays. Additional challenges include improving assay predictability, integrating data obtained from multiple ADMET and efÞcacy assays, increasing assay throughput, moving ADMET screening to earlier stages in the discovery process, developing predictive models of ADMET behavior, and mining historic ADMET data. One important and active area of development is the use of stem cells to generate hepatocytes. Isolated hepatocytes provide reasonably good models of hepatic metabolism, but these cells are expensive and not appropriate for largescale screening. Human hepatocytes can be obtained from stem cells in small quantities, but methods for reliably creating large quantities of hepatocytes for drug screening will probably require another two to three years of development.

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TABLE 5. Areas Requiring Additional Development for ADMET Screening Area of Development

Tasks and Technologies

Sources of cells and tissues for assays

Developing stem cells for producing human hepatocytes. Engineering cells that are easy to culture and that express the full array of drug-metabolizing enzymes. Engineered cells to simulate phenotypic and genotypic individual variations. Creating artificial miniature organs. Improving the ability to obtain human cells.

Assay development

Converting existing assays to high-throughput, low-volume, low-cost systems. Compressing assay times. Assays that account for multiple methods of drug uptake. Assay miniaturization and microfluidics technologies.

Data integration and prediction capabilities

Integrating data from ADMET studies, genomics, metabolomics, and chemistry. Correlating of ADMET assays with human clinical data. Predicting of inter individual variations in response.

An alternative to stem cells are engineered cells that express all of the drugmetabolizing enzymes and drug transporters that are normally found in human hepatocytes. Stem-cell and cell-engineering technologies will be probably be mature enough to dramatically improve the predictive ability of in vitro ADMET screening within three to Þve years. Quantifying the beneÞts of converting assays to high-throughput formats and conducting ADMET screening early in the drug discovery process are necessary to provide evidence that this approach saves development time. However, measuring the productivity of early ADMET testing is challenging. Metrics that validate early ADMET approaches, especially for toxicity testing, are difÞcult to obtain. The success of in vitro ADMET screening will be demonstrated only by reduced attrition in preclinical and clinical studies. Despite the absence of access to a deÞned metric, we predict that earlier ADMET screening will eventually improve the efÞciency of drug development. High-throughput in vitro ADMET testing promises to reduce compound attrition in preclinical and clinical development. In addition, this technology has the potential to reduce the cost and time required for clinical trials. This notion is supported by various FDA guidances to the industry stating that in vitro studies may be used to rule out the importance of a particular metabolic pathway and the drug-drug interactions related to the pathway. This information could reduce the amount of clinical testing that would usually be required at a later stage. For example, if in vitro experiments show that CYP2D6 is not involved

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in the metabolism of a drug, clinical studies of individuals with low levels of CYP2D6 and clinical drug–drug interaction studies of the investigational drug in combination with currently marketed drugs metabolized via CYP2D6 would not be needed. REFERENCES Guidance for industry: drug metabolism/drug interaction studies in the drug development process: studies in vitro. 1997. http://www.fda.gov/cder/guidance/clin3.pdf. Orphan nuclear receptors: promising targets for drug discovery. Spectrum, Drug Discovery and Design, Issue 10, 2002.

Emerging Concepts in GPCR Research and Their Implications for Drug Discovery

SUMMARY G-protein-coupled receptors (GPCRs) comprise the largest family of transmembrane signal receptors. GPCRs have long been targets for drug discovery research, although most successful drugs that target GPCRs were found serendipitously. The sequencing of the human genome, however, has revealed an abundance of new nonsensory receptors that could serve as therapeutic targets, and this Þnding has spurred researchers—who see potential for a wide variety of cardiovascular, neurological, immune and inßammatory, and gastrointestinal therapies—to reassess the direction of drug discovery research involving GPCRs. In this article, we explore the possibilities and the difÞculties facing future research in the area of GPCR-based drug discovery and highlight the activities of several companies focusing on the development of GPCR-based therapeutics and technologies. BUSINESS IMPLICATIONS •

•

G-protein-coupled receptors (GPCRs) present a great opportunity for new drug discovery and development. Researchers hope that targeted research on GPCRs, especially on orphan receptors, will yield many new drugs. Therapeutic areas under investigation include metabolic, immune and inßammatory, central nervous system, and cardiovascular. Researchers are revising classic concepts about GPCR function and are developing tools to study the molecular details of receptor binding events,

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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•

•

•

GPCR RESEARCH IMPLICATIONS FOR DRUG DISCOVERY

conformational changes, and dimerization. New and expensive instrumentation—such as mass spectrometry, positron emission tomography, and magnetic resonance imaging—is increasingly needed. A detailed understanding of the connections between receptor function and disease is not yet available for most GPCRs. Although animal models have been useful in determining drug effects on mammalian physiology, translating this knowledge to humans has been difÞcult. Advances in tissue-speciÞc mouse knockouts and phenotyping techniques will help to advance GPCRbased research. Interest in the potential of GPCR-based drug discovery is fueling the growth of biotechnology companies in this arena. Many of these companies are conducting internal discovery programs and are partnering with major pharmaceutical companies. Other biotech Þrms have developed valuable tools and technologies that promise to accelerate GPCR research and are offering their services and products to clients and partners. A potentially important area of drug development involving GPCRs that has so far not been investigated in detail is related to the concept of personalized medicine. The strong possibility that genetic variations in the GPCRs of different classes of patients might cause them to exhibit differential drug responses could open up new avenues of research.

INTRODUCTION Mammalian cells sense and respond to their environment through complex systems of intracellular signal transmissions. In these systems, an extracellular chemical signal contacts the cell surface and activates signal-speciÞc membranebound protein receptors, which then transfer the received signals into the interiors of the cells. G-protein-coupled receptors (GPCRs, also known as seventransmembrane receptors) comprise the largest family of these transmembrane receptors. Researchers have long recognized members of the GPCR family for their roles in mediating sensory transduction and cell homeostasis and, more recently, in regulating cell growth. The number of known receptors in this family, and their roles in cellular function, has expanded dramatically in recent years. Because of their ubiquity on cells, their involvement in many important cellular processes, and their “druggability” (meaning that their activity can be modulated by compounds in the extracellular environment), GPCRs are long-standing targets for drug discovery research. GPCRs present a great opportunity for new drug discovery and development. Currently marketed drugs address only approximately 500 targets. Cell membrane receptors—mostly GPCRs—represent about 45% of these targets, and more than 50% of currently marketed drugs target the GPCRs. Approximately 20% of the 100 top-selling pharmaceuticals are compounds that modulate GPCRs. These compounds account for approximately 8% of the global pharmaceutical market. Therapeutic areas targeted by current GPCR modulators include cardiovascular,

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central and peripheral nervous system, immune and inßammatory, and metabolic diseases and disorders. Although estimates vary, the sequencing of the human genome has revealed that approximately 720 genes encode GPCRs, approximately half of which researchers believe encode nonsensory receptors that could be therapeutic targets (sensory GPCRs, which are involved in sight, smell, and taste, are generally viewed to be unattractive as drug targets). Researchers have identiÞed the natural ligands for about 210 of these 360 GPCRs (Wise A, 2004). A pool of at least 150 GPCRs with unknown ligands or functions (termed “orphan” receptors) is therefore available that may furnish a rich source of drug targets. Historically, most successful drugs that target GPCRs were found serendipitously. Now, many experts assume that targeted research on GPCRs, especially on orphan receptors, will yield a large number of new drugs. Researchers are investigating the potential of GPCRs to serve as targets for treating obesity, diabetes, and cardiovascular, neurological, immune and inßammatory, and gastrointestinal diseases. Several recent developments, including the availability of human gene sequences, the introduction of molecular genetic techniques into pharmacology, and advances in understanding how drugs interact with membrane-bound receptors, are contributing to a growing body of knowledge about GPCRs. These developments are spurring researchers to reassess the direction that drug discovery research involving GPCRs is taking. This reassessment must confront certain boundaries that limit knowledge about GPCR functioning. For example, researchers have not yet elucidated a high-resolution, three-dimensional structure of any native human GPCR (although a research team has determined the structure of the bovine rhodopsin receptor in its inactive form (Palczewski K, 2000)). Also, the natural ligands and cellular functions of many GPCRs remain unknown, and the large-scale production of GPCRs (which is needed for structural studies and other experimentation) still lies in the future. Moreover, even in the few cases in which a GPCR’s sequence, ligand, and physiological function have been discovered, no practical advances in drug discovery based on this new knowledge have occurred. Despite these challenges, GPCRs remain popular targets for drug discovery. The historical success of drugs that modulate GPCRs, the development of new knowledge and experimental tools, and the sheer number of potential GPCR drug targets all serve to mitigate concerns that GPCR-based drug discovery might be difÞcult or unsuccessful. In this article, we explore the possibilities and the difÞculties facing future research in this area and highlight the activities of several companies focusing on the development of GPCR-based therapeutics and technologies. GPCR AND G-PROTEIN ACTIVATION Members of the GPCR family share a general structure that consists of seven hydrophobic transmembrane sections that span and loop in and out of the cell membrane (Figure 1). Depending on the receptor, agonist ligand binding sites are

372

GPCR RESEARCH IMPLICATIONS FOR DRUG DISCOVERY + H3N

Glycosylation sites

EC2

Extracellular

Cell membrane

EC3

EC1

I

II

III

IV

V

VI

VII

Intracellular

IC1

IC2 IC3

a

b

CO2

g G proteins

EC = Extracellular monophosphate (loop). IC = Intracellular monophosphate (loop).

FIGURE 1. GPCR structural motifs.

available either on the extracellular part of the receptor protein or at locations accessible from the extracellular surface. Ligand binding leads to the receptor’s interaction with an intracellular heterotrimeric G-protein, which consists of three subunits: G-alpha, G-beta, and G-gamma (Gα, Gβ, and Gγ ). At least 27 distinct genes in the human genome encode Gα subunits, and their expression levels vary widely. Gα subunits bind guanosine 5
-diphosphate (GDP) and guanosine 5
-triphosphate (GTP) tightly and speciÞcally and possess GTPase (an enzyme that removes a phosphate group from GTP) activity. Five and 14 distinct human genes encode the Gβ and Gγ subunits, respectively (Albert PR, 2002). Within a given cell, multiple GPCRs, G-protein subunits, and effectors (the targets of the G-protein subunits) are expressed. Researchers believe that at least some of the diverse cellular phenotypic effects signaled by GPCRs arise from the diversity of speciÞc combinations of the GPCRs, G-protein subunits, and effectors. SpeciÞc agonist binding to the receptor initiates GPCR signaling. Binding induces a conformational change in the GPCR that is transmitted through the membrane. In its inactivated state, the Gα subunits bind GDP. The conformational change in the GPCR induced by agonist binding catalyzes the exchange of the

DEVELOPMENTS IN CONCEPTUAL APPROACHES TO GPCRS

373

GDP with GTP, thus activating Gα and causing it to dissociate from the GβGγ subunit. The now-activated subunits initiate a complex signaling cascade that changes the cell’s phenotype and causes a speciÞc physiological response. After signal initiation, the GTPase activity of the Gα subunit converts the bound GTP to GDP, making the Gα subunit again inactive and allowing the subunits to recombine to form GαGβGγ in preparation for another cycle. DEVELOPMENTS IN CONCEPTUAL APPROACHES TO GPCRS As we have discussed, upon speciÞc binding with their extracellular ligands, GPCRs transmit the information about the binding event to the interior of the cells—presumably by subtle conformational changes in the receptor protein. The nature of both the binding events and the subsequent conformational changes are thought to be important factors that underlie the differences between agonism and inverse agonism—concepts that describe the results of particular receptor–ligand interactions. Another important concept in understanding GPCR signaling events is that of receptor dimerization. Researchers are therefore very interested in developing tools to study—especially in vivo —the molecular details of receptor binding events, conformational changes, and dimerization. Agonism and Inverse Agonism Until the 1990s, scientists described ligands and their receptor interactions in terms of the binding afÞnity of a ligand for its cognate receptor. According to this classic conception, high-binding-afÞnity ligands were classiÞed as either agonists or antagonists; agonist binding activated the receptor, while antagonist binding did not. Over the last twenty years, this classic formulation has gradually changed. Some ligands for GPCR that were previously thought to be antagonists have been shown to exert their actions not by blocking their cognate receptors but rather by reducing their activities. Such ligands are now termed “inverse” agonists. Nearly thirteen years ago, researchers proposed a model in which receptors exist in two states—active and inactive—that are in dynamic equilibrium with each other (Costa T, 1992). According to the proposal, agonists bound to and stabilized the active state, while inverse agonists bound and stabilized the inactive state. More-recent work indicates that the simple two-state model may need modiÞcation and that some inverse agonists may act not only by stabilizing the inactive state but also by initiating intracellular signaling cascades. The possibility of multiple states for GPCRs that can be differentially stabilized by speciÞc ligands has stimulated research using various tools for determining the conformations of the receptors upon ligand binding (see “Developments in Research Tools and Technologies”). Researchers have described the concept that GPCRs exist in multiple active conformations that can be differentially stabilized as agonist-directed trafÞcking of response (ADTR). This concept is of potentially great importance for future drug discovery. Current methods of identifying GPCR ligands often rely on in vitro screening to detect agonists that differ primarily in the strengths (efÞcacies)

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of the intracellular responses they elicit. However, studies on different conformational equilibrium effects elicited by binding of different ligands may show different ADTRs involving distinct conformations that could couple GPCRs to speciÞc G-proteins and intracellular signaling pathways. This approach could open the door to discovering drugs that affect only speciÞc signaling pathways and therefore have fewer unwanted side effects. For example, development of opioid ligands that induce analgesia but are nonaddictive might be possible. Dimerization A potentially very important and complicating factor in GPCR-based research is the phenomenon of receptor dimerization. While for many years the functional forms of GPCRs were thought to be monomers (i.e., they existed as a single receptor protein), some experimental evidence now suggests that many GPCRs may form dimers (complexes of two receptors). The concept of receptor dimerization has developed rapidly as techniques for studying protein–protein interactions have advanced. This aspect of GPCR research raises several important questions: • • •

•

Are dimers functional forms of GPCRs? Is dimerization kinetically stable, or is there an equilibrium between receptor monomers and dimers? What are the functional ramiÞcations of homodimerization (occurs when two of the same GPCRs form pairs) and heterodimerization (occurs when two different GPCRs form pairs) of GPCRs, and which of these forms predominate? Do GPCR-speciÞc ligands alter GPCR homodimer and heterodimer interactions?

The last question is probably the most important for drug discovery. If dimerization does prove to be physiologically signiÞcant, ligands capable of altering receptor dimer interactions may be pharmacologically important. Unfortunately, researchers have not deÞnitively answered any of these four questions. Although experimental evidence unequivocally shows that dimerization exists in some in vitro systems, the physiological signiÞcance of these Þndings is not known. Because of its potential importance to understanding biology and to drug discovery, GPCR dimerization is a key and active area of research. DEVELOPMENTS IN RESEARCH TOOLS AND TECHNOLOGIES Researchers use a variety of tools and technologies to elucidate the structure and function of GPCRs. Of particular interest are basic biophysical studies on GPCRs, such as analyses of ligand-activated conformational changes and studies of molecular modiÞcations, particularly post-translational modiÞcations to receptor proteins (e.g., phosphorylation and glycosylation). Other important areas of research include inquiry into the physiological sites of action of drugs affecting

DEVELOPMENTS IN RESEARCH TOOLS AND TECHNOLOGIES

375

GPCRs and the effects that various physiological conditions (e.g., stress, pain, inßammation) have on the intensity of action at the sites. Finally, continuing high-resolution studies of the sequences of human and mammalian genomes are producing results that will have an impact on drug discovery methodology and business decisions involving drugs targeting GPCRs. In the following sections, we describe some of the most important tools researchers are applying and developing for GPCR-based research. Fluorescence Technology Fluorescence technology is the basis for a group of related tools that allow researchers to study protein–protein and ligand–protein interactions in vitro and in vivo. The most important ßuorescence tools include total internal ßuorescence reßection for single-molecule imaging and ßuorescence energy transfer methods such as ßuorescence resonance energy transfer (FRET), bioluminescence resonance energy transfer (BRET), and ßuorescence lifetime imaging (FLIM). These ßuorescence technologies can allow researchers to analyze conformational changes during binding events on the necessary micro- to millisecond time scale. Mass Spectrometry Modern mass spectrometers can easily detect the addition or subtraction of a single phosphate or carbohydrate group on a GPCR. Because phosphorylation and glycosylation are the most frequent post-translational modiÞcations of proteins, various forms of mass spectrometry are the most appropriate and powerful tools available for studying these changes on GPCRs. Imaging Methods Positron emission tomography (PET) scanning is a powerful tool for locating a drug’s site of action as the drug binds to its receptor and becomes localized. Researchers have also recently developed magnetic resonance imaging (MRI) methods that allow the detection of receptors on single cells. Sophisticated imaging methods such as these are needed to understand the details of ligand–GPCR or drug–GPCR binding and signaling and their physiological effects. Genomics As the complete nucleotide sequences of human and other mammalian genomes have become available, researchers have identiÞed genetic variations among individuals called single nucleotide polymorphisms (SNPs). Some of these variations in gene sequences may affect the encoded protein’s functional activities or speciÞcities. Variations in protein drug targets among different patients could result in two patients having different responses (including adverse reactions) to the same drug. The notion that different people—based on their different genetic

376

GPCR RESEARCH IMPLICATIONS FOR DRUG DISCOVERY

makeup—will respond differently to the same drug has led to the concept of “personalized medicine,” in which a therapy is tailored for a speciÞc subset of the population. Because of the large number of potential GPCR targets, researchers may need to consider genetic variation when choosing which targets to pursue. Animal Models The development of better animal models should be a priority for GPCR-based drug discovery; however, precisely how to develop appropriate models is unclear. Preclinical drug development, especially drug-target validation, has traditionally relied heavily on animal models, particularly inbred strains of rodents. Although the use of homogeneous, inbred strains to test the effects of compounds provides the advantage of reproducibility, this advantage is offset by the frequent failure of these models to accurately predict a compound’s effect in humans. This failure results partly from the inability of inbred animals to adequately mirror human genetic diversity. Studying SNP patterns obtained from the sequences of important genes in noninbred strain rodents, and understanding the functional relevance of these variations, will be important steps toward developing more predictive animal models. Researchers have been rapidly developing new molecular genetic methods for manipulating gene expression in animals. The most popular genetic technique for GPCR research in animals is the production of “knockout” strains of rodents. In this approach, particular receptor-encoding genes are deleted, and the physiological effects of the deletion studied. Researchers have used knockout mice successfully to delineate the functions of several GPCRs. Another molecular genetic approach is to create transgenic mice containing human genes (“knockins”); this technique has also been applied to receptor research. However, both knockout and knockin animals are used mainly to conÞrm the function of a speciÞc GPCR rather than to discover new drugs. The best hopes in the near term for producing new animal models for GPCR-based drug discovery probably lie in the development of methods for tissue-speciÞc knockouts and knockins of individual GPCRs. In this approach, researchers are able to modulate the expression of a speciÞc receptor in speciÞc tissues. Another important area for development is that of phenotyping—that is, determining the physiological effects of a speciÞc genetic manipulation. Researchers widely believe that current phenotyping techniques are inadequate for monitoring subtle responses to GPCR activation and changes in response to stimuli, including test compounds. Improvements in phenotyping would therefore signiÞcantly advance the Þeld of GPCR-based research (see “Major Challenges in GPCR-Based Drug Discovery”). Reverse Pharmacology Reverse pharmacology is an important method for discovering the natural ligands of orphan receptors. The classical approach to the discovery of drugs that modulate a receptor’s function started with the identiÞcation of a biologically active

CHEMOKINE RECEPTORS AS DRUG TARGETS

377

Classical Pharmacology Functional Ligand purification activity

Ligand

Functional study

Receptor

Ligand “fishing”

Biological role

Receptor “fishing”

Receptor

Medicinal chemistry

Drug candidate

Functional study

Biological role

Medicinal chemistry

Drug candidate

Reverse Pharmacology Molecular genetics

Cloning

Ligand

FIGURE 2. Classical versus reverse pharmacology.

ligand. The ligand was then used to “Þsh” for the corresponding receptor. In the reverse pharmacological approach, an orphan receptor of interest is selected, cloned and expressed, and used to Þsh for a natural ligand (Figure 2). Once the receptor and ligand are in hand, researchers determine the biological role of the receptor and screen for synthetic ligands. Researchers at GlaxoSmithKline (GSK) demonstrated the utility of this strategy in 1998 by elucidating a novel neuropeptide system (Sakurai T, 1998). The researchers used an orphan GPCR in a cell-based reporter assay to screen tissue extracts for ligands. The team identiÞed two neuropeptides, Orexin A and Orexin B, that were expressed in the hypothalamus (the feeding center of the brain) and that bound selectively to the assayed receptor. Injection of Orexin A into a rat’s brain stimulated food intake, and fasting upregulated the expression of Orexin mRNA. These results suggest that this receptor (called the Orexin receptor) constitutes a candidate target for antiobesity drugs. CHEMOKINE RECEPTORS AS DRUG TARGETS Chemokines are members of a large family of low molecular-weight proteins. A category of the cytokines, the chemokines function mainly as chemoattractants for certain types of leukocytes and are important in recruiting them to regions of infection and inßammation. The biological effects of chemokines are mediated through binding to GPCRs present on the surfaces of various leukocytes. As with other GPCR ligands, the binding event activates the receptor and initiates an intracellular cascade of signals. The known chemokine system in humans comprises approximately 50 ligands and 20 receptors. Experimental evidence based on animal models has associated the expression of chemokines with the pathophysiology of many diseases, including bacterial and viral infections, allergy, psoriasis, rheumatoid arthritis, and atherosclerosis. The chemokine system is deÞnitively associated with human immunodeÞciency virus (HIV) infection: HIV utilizes the CCR5 and CXCR4 chemokine receptors to infect target cells. These Þndings have generated considerable excitement over the possibility that chemokine GPCRs could be useful targets for drug development, and researchers have developed nonpeptide antagonists that bind speciÞc

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GPCR RESEARCH IMPLICATIONS FOR DRUG DISCOVERY

chemokine receptors. Chemokines and their cognate receptors are attractive as drug targets in part because of their speciÞcity. Unlike the cytokines, many of which have pleiotropic effects, chemokines target speciÞc kinds of leukocytes. Therefore, drugs that modulate a chemokine ligand or receptor would be expected to produce limited side effects. The CCR1 chemokine receptor has received attention as a potential drug target from researchers. The receptor is found in many species, including humans, and sequences are available for some of these receptors. The human and rhesus protein sequences are 87% identical, and the rat and mouse proteins are 80% identical. Researchers have shown that the CCR1 receptor plays a role in the pathophysiology of multiple sclerosis and organ transplant rejection (Horuk R, 2001). Several pharmaceutical Þrms are developing small-molecule ligands that bind to CCR1 for indications such as multiple sclerosis, rheumatoid arthritis, inßammation, and transplant rejection. The CCR2 and CCR5 receptors have been well validated in various disease models and are under investigation for several indications. Other chemokine receptors under investigation as drug targets include CCR3, CXCR2, and CXCR4. In addition to the association of HIV with chemokine receptors, numerous herpesviruses and poxviruses are now known to encode chemokine mimics and are able to block chemokine action. SpeciÞc herpesviruses and lentiviruses can also exploit the immune system through chemokine mimicry, for example, to facilitate viral dissemination or, as in the case of HIV-1, to directly infect leukocyte target cells. The study of viral mimicry of chemokines and chemokine receptors is aiding researchers in developing important new concepts in viral immunopathogenesis as well as providing new anti-inßammatory drug targets, leads, and approaches for antiviral drug and vaccine development. MAJOR CHALLENGES IN GPCR-BASED DRUG DISCOVERY Certain technical obstacles, such as the expense of large-scale use of PET and MRI scanning and the inability to obtain accurate structural information on GPCRs, hinder the ability of researchers to conduct GPCR-based research. However, more important fundamental limitations that may be impossible to overcome in the foreseeable future present particularly formidable challenges. Chief among these presently intractable problems are the lack of good animal models and the lack of sensitive phenotyping techniques. Although the functions and natural ligands of many orphan receptors remain to be discovered in humans, researchers believe that many of these functions will affect the central nervous system (i.e., will be nonsensory receptors) and will be involved in difÞcultto-measure brain functions (e.g., feelings, mood, anxiety, stress, memory) and psychiatric disorders. Other than those for crude measures of pain, no animal models are currently available that adequately reßect any of these higher order human brain functions—and some might argue that such models will be impossible to develop.

CORPORATE ACTIVITIES

379

Researchers widely believe that the lack of accurate 3-D structures for GPCRs is holding back rapid advancement in understanding their modes of action, including activation by speciÞc ligands and interactions (such as dimerization) with other proteins. As discussed previously, only one high-resolution crystal GPCR structure—that of bovine rhodopsin in an inactive form—is known. The basic reason for the lack of progress in structure determination has been the difÞculty of obtaining the large quantities (i.e., milligram quantities) of puriÞed proteins that are required for either of the two major methods of structure determination: NMR spectroscopy and X-ray crystallography. Also, X-ray crystallography requires protein crystallization, a step that has also proved difÞcult for GPCRs. NMR spectroscopic structural determination of protein structures has advanced sufÞciently to allow reasonably accurate structures of proteins in the 40–70 kDa range of GPCRs, although these structures are determined in solution rather than in the native membrane environment. Recently, researchers have described the production of milligram quantities of one GPCR that is speciÞc for the peptide ligand neurotensin (White JF, 2004). To do so, these researchers overexpressed the GPCR as a fusion protein in bacteria and puriÞed the protein with neurotensin-afÞnity column chromatography. Whether the protein assumes a regular folded conformation in this approach remains to be shown. Another hurdle has developed simply because of the popularity of GPCR research in both industry and academia. The total number of publications related to GPCRs has increased by a factor of 100 during the past twenty years, and the number of patents related to GPCRs has increased more than 100 times during the past ten years. Unless companies can streamline solutions to various legal and intellectual property problems, the existence of broad patents and related licensing issues—particularly those for orphan receptors—will undoubtedly slow down research in this area. CORPORATE ACTIVITIES The most distinct trends for pharmaceutical and biotechnology companies engaged in GPCR-based drug discovery are (1) to conduct internal research programs aimed at discovering natural ligands for orphan receptors (“deorphanization”) and (2) to establish collaborations with other Þrms (usually small ones) that have developed proprietary techniques that are useful for GPCR research. For example, Millennium Pharmaceuticals is employing both of these strategies. Millennium has a large in-house program for identifying ligands for orphan GPCRs, but it also has formed alliances with smaller Þrms that have developed proprietary technologies that can accelerate identiÞcation of ligands. Smaller Þrms, academic groups, and consortia of academics are all active in developing technologies useful for GPCR-based research. Interest in the potential of GPCR-based drug discovery is fueling the growth of biotechnology companies in this arena. Many of these companies are conducting internal discovery programs and are partnering or seeking partnerships

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GPCR RESEARCH IMPLICATIONS FOR DRUG DISCOVERY

TABLE 1. Profiled Companies Active in G-Protein-Coupled Receptor (GPCR) Research or Technology Development/Services Company

Web Site

7TM Pharma

www.7tm.com

Actelion

www.actelion.com

Arena Pharmaceuticals

www.arenapharm.com

BioFocus

www.biofocus.com

Bio-Xtal

www.bioxtal.com

Euroscreen

www.euroscreendiscovery.com

Norak Biosciences

www.norakbio.com

Nura

www.nurainc.com

Septegen

www.septegen.com

GPCR-Related Technology/Focus Drug discovery for metabolic disorders and inflammation; structure-based analysis and screening tools. Drug development for endothelin, urotensin II, and orexin receptors; cheminformatics discovery platform. Constitutively activated receptor technology (CART); Melanophore screening technology. GPCR-targeted drug discovery and design services; compound libraries. X-ray crystallography, production of recombinant proteins. High-throughput assays; functional cell-based assays; mammalian cell lines and membrane preparations expressing recombinant GPCRs; custom cloning services. Discovery and development services; high-throughput screening; Transfluor bioassays; compound library. Drug discovery for neurological disorders; GPCR knockout mice; brain GPCR database. Yeast-based screening services.

with major pharmaceutical companies. Other biotech Þrms have developed valuable tools and technologies that promise to accelerate GPCR research and are offering their services and products to clients and partners. In this section, we highlight several biotechnology companies that are active in developing GPCR-based technologies and therapeutics (see Table 1 for a list of these companies). 7TM Pharma 7TM Pharma (Horsholm, Denmark) is a private company that was spun out from the University of Copenhagen in 2000. The company’s drug discovery programs are focused primarily on targeting GPCRs to develop treatments for metabolic disorders and inßammation. 7TM has preclinical and discovery-stage candidates for obesity, CNS disorders, inßammation, and angiogenesis.

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7TM uses its core technology, Site-Directed Drug Discovery, to deÞne key contact points and ligand binding structures on a receptor, and then identify other receptors with similar structures and known ligands. The company then uses this information in applying computational screening techniques and structural binding analysis (using target–receptor mutagenesis) to design small, GPCRspeciÞc compound libraries. 7TM then screens compounds in functional cellbased assays. The company has developed functional BRET assays, which can be used to screen GPCRs in living cells. The company is adapting these screens to medium- and high-throughput modes. 7TM is seeking to form strategic partnerships for speciÞc receptors or receptor families. The company intends to develop promising therapies for Phase II trials and then partner with a pharmaceutical company to complete clinical trials and bring the compound to market. In September 2004, 7TM formed a research collaboration with Synaptic (a wholly owned subsidiary of Lundbeck). In the collaboration, 7TM will apply its Site-Directed Drug Discovery to Synaptic targets. The companies also have a prior research agreement based on 7TM’s technology. In August 2004, 7TM formed a research collaboration with AstraZeneca in which 7TM will apply its technology to targets from AstraZeneca. Actelion Actelion (Allschwil, Switzerland) is a publicly traded company founded in 1997. Actelion specializes in endothelium-related research, including cardiovascular, central nervous system (CNS), and oncology as therapeutic areas. Actelion’s main source of funding is sales of bosentan (Tracleer), an orally administered, dual endothelin (ET) receptor antagonist to treat pulmonary arterial hypertension. Actelion is also expecting to generate revenue from sales of miglustat (Zavesca), an oral inhibitor of glucosylceramide synthase. In August 2003, the FDA approved miglustat for the treatment of type 1 Gaucher’s disease. Actelion is investigating several GPCR modulators for various indications. Tezosentan (Veletri), Actelion’s intravenous dual ET receptor antagonist, is under investigation as a treatment for hepatorenal syndrome and is in Phase III trials for acute heart failure. The company is evaluating clazosentan, a selective ETA antagonist, for the prevention of vasospasm following subarachnoid hemorrhage, and a urotensin II receptor antagonist for type II diabetes in patients with chronic renal failure. Actelion is also investigating orexin receptor antagonists for the acute or chronic treatment of sleep disorders and obesity. In 2003, Actelion acquired Axovan, a company that had developed a proprietary computational platform, Computer-Supported Accelerated Drug Discovery System (AXADDIS), aimed speciÞcally at GPCR drug discovery. This cheminformatics platform combines in-house databases and computational tools to improve selectivity and absorption, distribution, metabolism, excretion (ADME) properties of potential therapeutics. The platform enabled Axovan to generate a library of small chemical molecules that target GPCRs. The merger also brought

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clazosentan, which Axovan had inlicensed from Hoffmann-La Roche, and other compounds to Actelion. Arena Pharmaceuticals Founded in 1997, Arena Pharmaceuticals (San Diego, California) is a publicly traded biopharmaceutical company focused on GPCR-based drug discovery. Among the therapeutic areas Arena Pharmaceuticals is investigating are metabolic, cardiovascular, CNS, and inßammation disorders. Arena’s orphan GPCR screening technology, constitutively activated receptor technology (CART), allows investigators to identify compounds that modulate receptors of interest without having Þrst identiÞed the natural ligand. This technology serves as the core of Arena’s drug discovery efforts. Arena’s Melanophore technology, which the company acquired via its acquisition of receptor screening company Bunsen Rush Laboratories, is a functional assay that is effective for identifying modulators of GPCRs that couple to all major G-protein classes. In July 2004, Arena initiated a Phase Ib trial of its lead antiobesity drug, APD-356. In October 2002, Merck and Arena entered a multiyear research and licensing agreement to evaluate the potential of three GPCRs as drug targets. In February and August 2004, Arena announced the achievement of research milestones that triggered payments of $4 million and $3 million, respectively. Arena also has partnerships and/or licensing agreements with Eli Lilly and Company, Ferring Pharmaceutical, TaiGen Biotechnology, Taisho Pharmaceutical, Fujisawa Pharmaceutical, and others. BioFocus BioFocus (Chesterford Research Park, United Kingdom) is a publicly traded company that was founded in 1997. The company collaborates with biotechnology and pharmaceutical companies on projects for the development of therapies involving GPCRs, ion channels, and kinases and provides research support and services for drug discovery and design. BioFocus’s proprietary technology, Thematic Analysis, is part of its GPCR-targeted library for low-volume screening. The technology analyzes small-molecule activity and receptor protein sequences. The company’s SoftFocus GPCR-targeted libraries are made up of approximately 1000 high-purity compounds. Using these libraries, BioFocus is able to customize assays and high-throughput systems for GPCR-targeted drug discovery. BioFocus has numerous agreements with various pharmaceutical and biotechnology companies. In 2001, BioFocus formed a research collaboration with Biovitrum (Stockholm Sweden) to discover inhibitors of a GPCR thought to be related to weight reduction. In September 2004, the collaboration had reached the end of the lead generation phase and the companies announced that Biovitrum would acquire rights to antiobesity lead compounds from BioFocus. In February 2002, BioFocus announced it would partner with UCB Pharma to identify new therapeutics targeting GPCRs. BioFocus will provide lead discovery libraries for GPCRs that UCB Pharma is investigating.

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383

Bio-Xtal Founded in 1998, Bio-Xtal is a privately held company that offers proteomics services. These services include protein crystallographic structure resolution (via X-ray crystallography) and the production of recombinant proteins. The company is coordinating an international effort called the Membrane Protein Network (MePNet), to determine the structures of 100 GPCRs. The collaboration involves four European research institutes and is supported by more than 30 companies working together to develop high-throughput tools and crystallization methods to solve GPCR crystal structures. Euroscreen Founded in 1994 as a spinout from the University of Brussels, Euroscreen is focused on discovering ligands for orphan receptors and novel drugs that modulate them. Euroscreen provides HTS assays for GPCRs and is building an in-house portfolio of GPCR targets. The company’s core technology, AequoScreen, is a functional screening system in a high-throughput format. AequoScreen uses apoaequorin gene expression in cell-based assays to detect GPCR activation through luminescence; Euroscreen owns exclusive worldwide rights to the use of this gene. The company also provides various mammalian cell lines and membrane preparations expressing recombinant GPCRs. Custom cloning services for GPCRs of interest and in-house screening are also available. Revenue from corporate collaborations and the sale of GPCR-speciÞc drug discovery tools support Euroscreen’s research and operations. Norak Biosciences Norak Biosciences (Research Triangle Park, North Carolina) is a privately held company investigating compounds that modulate GPCR function in several therapeutic areas. Norak Biosciences develops high-throughput methods to screen potential GPCR-modulating compounds and provides discovery and development services to the pharmaceutical industry. Its proprietary technology, Transßuor bioassays, can identify ligands for GPCR targets (known or orphan). Transßuor uses green ßuorescent protein (GFP) as a reporter to signal GPCR conformational changes. The company has widely licensed the Transßuor technology. Norak expresses GPCRs of interest to other companies in HEK-293 cells and can customize this technology for high-throughput screening. The company has an internal library totaling 800,000 compounds to screen for GPCR modulation. Norak is planning to eventually develop GPCR-related therapeutics for a wide range of diseases. Nura Nura (Seattle, Washington), a start-up founded in 2003, is using a combination of localization knockout mice and behavioral studies to identify GPCRs in the brain that are involved in neurological and behavioral disorders. Nura has a map of GPCR expression throughout the body and within the brain and a database

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of more than 200 brain GPCRs. The company has developed mouse knockouts for brain GPCRs, and it is analyzing GPCRs that have drug-target characteristics. Nura intends to use this information to develop therapeutic compounds for neurodegenerative diseases and behavioral disorders such as Parkinson’s and Alzheimer’s diseases as well as various sleep, appetite, and other behavioral disorders. In September 2004, Nura completed its Þrst Þnancing, raising $9.5 million in a series A venture round. The company has also received a $1 million grant from the National Institutes of Health to study anxiety. In October 2004, Dr. Linda Buck of the Fred Hutchinson Cancer Research Center—and the founder of Nura and chair of its scientiÞc advisory board—shared the Nobel Prize in Medicine for her pioneering work in elucidating olfaction, which is mediated by GPCRs in the nose. Septegen Founded in February 2001 by researchers at the University of Warwick, Septegen (Coventry, United Kingdom) offers yeast-based screening services for GPCRrelated drug discovery and development. Septegen uses its proprietary Septecells technology to measure the activation of GPCRs. Researchers at Septegen engineer Þssion yeast to express human GPCRs and measure GPCR activation using a humanized Gα subunit. The measurements are based on a variety of readout methods, including beta-galactosidase, cell growth, and inhibition of growth. Septegen uses these cells in all aspects of its GPCR screening and proÞling process. Septegen is also developing a proprietary high-throughput screening method, Septescreen, to aid companies in their search for compounds that modulate GPCR activity. Using Septecells, Septescreen can be formatted to screen one GPCR against one or more compounds, or multiple GPCRs against multiple compounds. The company is developing a proprietary compound proÞling method, Septetox, to identify potential side effects of GPCR-modulating compounds. Septegen is also developing a proprietary GPCR orphan-characterizing technology called Septeorph. OUTLOOK Research into the genetics, molecular biology, and basic pharmacology of GPCRs has recently contributed to a rapid accumulation of knowledge about the roles these receptors play in human physiology. In the next Þve years, the ability to apply this knowledge successfully to drug discovery will prove critical to the development of novel GPCR-based therapeutics. Most scientists researching GPCR function assume that identifying the natural ligands for orphan receptors (deorphanization), detailing the conformational changes in the receptors upon ligand binding, and discovering associations between receptor activation and disease will lead to successful drug discovery. The rapid advances of the past ten to Þfteen years in developing concepts and technologies for GPCR research make a powerful case for optimism about this assumption.

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However, this optimism must be balanced against the fact that, so far, none of the marvelous technological developments in molecular and structural biology have produced any major advances in the discovery of new GPCR-based drugs. For example, researchers discovered the natural opioid ligands twenty-Þve years ago. That discovery was followed by the elucidation of the molecular biology of the natural opioids, by the sequencing of their GPCR, and then by the production of knockout mice that revealed much basic information on nociception (pain perception) mechanisms. Unfortunately, this large body of solid, in-depth, scientiÞc knowledge has led to no new drugs with any clinical advantage over morphine. Therefore, purely technological progress is unlikely to dramatically increase the number of successful GPCR-targeting drugs developed in the near future. A potentially important area of drug development involving GPCRs that has so far not been investigated in detail is related to the concept of personalized medicine. The strong possibility that genetic variations in the GPCRs of different classes of patients might cause them to exhibit differential drug responses could open up new avenues of research. Detailed knowledge about GPCR function and physiology may be useful in aiding researchers seeking to modify existing drugs to optimize efÞcacy and minimize the occurrence of adverse drug reactions, or to target speciÞc drugs to speciÞc classes of patients. Knowledge about the genetic variability for a particular receptor could also help guide the choice of novel GPCRs as targets for drug discovery. Furthermore, this information can help researchers design drugs that are effective against different variants of a target. The dominant concept of drug treatment (i.e., the time-honored idea of discovering a “magic bullet” drug that targets a single receptor to cure a single disease state) is changing. The change has been brought about by expanding research that emphasizes that many, and possibly most, diseases are complex states to which many factors contribute. Furthermore, all treatments for disease take place within complex cellular environments and depend on the existence of interactive molecular networks within cells. These concepts suggest that the way to treat many diseases may be far more complicated than that suggested by current approaches. Understanding the etiology of complex diseases and the cellular interactions that affect their treatment will guide the future course of GPCR research. REFERENCES Albert PR, Robillard L. Protein speciÞcity: trafÞc direction required. Cell Signal. 2002;14:407–418. Costa T, et al. Drug efÞcacy at guanine nucleotide-binding regulatory protein-linked receptors: thermodynamic interpretation of negative antagonism and of receptor activity in the absence of ligand. Molecular Pharmacology. 1992;41(3):549–560. Horuk R. Chemokine receptors. Cytokine & Growth Factor Reviews. 2001;12(4):313–335. Palczewski K, et al. Crystal structure of rhodopsin: a G protein-coupled receptor. Science. 2000;289:739–745. Sakurai T, et al. Orexins and orexin receptors: a family of hypothalamic neuropeptides and G protein-coupled receptors that regulate feeding and behavior. Cell. 1998;92:573–585.

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White JF, et al. Automated large-scale puriÞcation of a G protein-coupled receptor for neurotensin. Federation of European Biochemical Societies Letters. 2004;564(3):289–293. Wise A, et al. The identiÞcation of ligands at orphan G-protein coupled receptors. Annual Review of Pharmacology and Toxicology. 2004;44:43–66.

Optimizing Knowledge Management in Pharmaceutical Companies

SUMMARY The pharmaceutical and biotech industries are increasingly turning to knowledge management (KM) to help them succeed. KM can improve productivity in the face of an increasingly complex R&D process, allow the sharing of data and information between different parts of an organization, and enable the sharing of expertise with partnering companies in cases of licensing deals and acquisitions and mergers. In this article, we describe how KM is being used today, how the technologies for KM are evolving, and how to best exploit these new tools. BUSINESS IMPLICATIONS •

•

The growing interest in knowledge management (KM) within the pharmaceutical industry is fueled by the need to improve productivity in the face of an increasingly complex R&D process; share data and information between different parts of an organization, including drug discovery, development, manufacturing, and marketing; and identify and share expertise with partnering or merged companies. Even though KM initiatives in the pharmaceutical industry typically use some form of information technology (IT), KM is far more than an IT project. To be successful, those planning KM initiatives should focus Þrst on which projects can be most improved by an initiative, then on what goals are to be achieved, and Þnally on how it will be done—the step when IT often takes a leading role.

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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OPTIMIZING KNOWLEDGE MANAGEMENT IN PHARMACEUTICAL COMPANIES

Any process that would beneÞt from optimizing the capture, sharing, and application of information is a candidate for KM. Done correctly, such projects address an organization’s needs and goals. Pharmaceutical and biotechnology companies have some unique considerations that inßuence how KM projects are planned and executed, including having data and information as their primary asset, extreme sensitivity about data security, long-lived projects, a complex regulatory environment, and reliance on employees’ extensive personal knowledge. The tools for KM and ideas about its implementation have matured signiÞcantly–more than 60 companies now offer KM-related software and services. Careful integration of a variety of tools with the existing IT infrastructure is needed to facilitate sharing and application of complex and valuable data across many divisions.

INTRODUCTION In recent years, the pharmaceutical industry has experienced both a data explosion and a concurrent drop in productivity. It is not surprising that more and more pharmaceutical companies are turning to knowledge management (KM), a Þeld devoted to optimizing information capture, dissemination, and sharing. (See the sidebar, “What Is Knowledge Management?”) WHAT IS KNOWLEDGE MANAGEMENT? We deÞne knowledge management (KM) as the systematic capture, integration, distribution, and application of information used to advance the objectives of a company. (For more information, see “Knowledge management in drug discovery R&D,” a white paper from 3rd Millennium, available at http://ww.3rdmill.com; and Dweck R. “Knowledge management: is the hype justiÞed? Bio-ITWorld, http://www.bio-itworld.com/archive/011303/horizons km.html.) Knowledge begins with data, which are collected by instruments or by people. Often, the primary data have no intrinsic value, and distributing the data is not useful. Once the data have been analyzed, it is often insufÞcient to have only the resulting information aggregated in a database. Some users will want to understand how the data were analyzed and to track back to the original source if necessary. Being able to do so will help them determine if the information is credible and relevant, a very important condition if they hope to integrate information from multiple sources. This need for data transparency holds true when the information is related to qualities of a particular biological target being studied using microarrays or to the results of a clinical trial or marketing survey. As a process, KM has many parts and steps, but it has two main objectives: • •

To optimally manage the ßow of information within an organization. To monitor the progress of that information ßow and its impact on the organization’s goals.

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Properly implemented, KM will ultimately allow structured decision support—the ability to use the same relatively objective criteria when making similar decisions; in short, to answer similar questions in a standard, optimized way. Data, information, and knowledge are distinct. The raw material for knowledge creation is information. The emphasis of KM is on creating a process that accelerates the ßow and use of data and information. The appropriate strategy for a particular project depends on the type of knowledge being managed. Knowledge is difÞcult to deÞne and has many dimensions: • • • •

That known to the individual (e.g., the temperature on block 3 of the PCR machine ßuctuates randomly). That known to the collective (e.g., the PCR buffer and Taq polymerase are contained in the freezer by the sink). Explicit knowledge (e.g., the standard PCR conditions are X, Y, and Z). Tacit knowledge (e.g., to eliminate smears on the readout, do X, unless Y is true, in which case, do Z).

However, much knowledge may remain tacit, creating one of the key barriers to successful KM. One of the major challenges in KM is to render tacit knowledge explicit so that it can be managed. The Þeld of KM has now matured beyond the point where it is regarded mainly as a fuzzy, catch-all phrase that makes intuitive sense but has no substance. A large number of very useful tools and approaches are available or coming into use, and a growing number of successful case studies of KM in the pharmaceutical and biotech industries are described in the literature. (Two well-known examples of KM successes are PÞzer’s discovery of sildenaÞl citrate (Viagra) through mining clinical data and the discovery by Eric Lander’s group at the MIT/Whitehead Institute of a gene that causes a cytochrome c oxidase deÞciency. The latter discovery did not require any new information. It was made in part by integrating and mining multiple, existing genomic databases. It will lead directly to a diagnostic test for a rare and lethal genetic condition.) The potential uses for KM are extremely varied, and deciding why it should be implemented is just as important as determining how to do it. There are many examples (some we will discuss) in which successful KM projects have been carried out. However, if the wrong question is being addressed with KM or the process takes up as much time as it saves, the project is not worthwhile. Given the abundance of software tools for KM, many people have come to view KM as an IT project. However, the major challenge facing companies is Þrst identifying the projects that can beneÞt most from the application of a KM initiative. Once the best use for KM is determined, the challenge becomes Þnding the right mix of tools and knitting them into a system that suits the unique structure, culture, goals, and other systems within the company. Hence, KM must be regarded primarily as a management task, but one in which the optimal deployment of technology is essential. In this article, we describe how KM is being used today, how the technologies for KM are evolving, and how to best exploit these new tools.

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THE RISING DEMAND FOR KNOWLEDGE MANAGEMENT Although KM has only recently become popular in the pharmaceutical industry, there are several reasons for its rapidly growing acceptance in this Þeld. The difÞcult Þnancial situation in which pharmaceutical and biotech companies Þnd themselves today and the fact that KM is well suited to the particular challenges these industries face combine to make KM an increasingly attractive option. A Brewing Financial Crisis The pharmaceutical industry is under growing pressure to boost productivity. R&D spending has increased steadily while the rate of new-drug launches has signiÞcantly declined. IMS’s annual review of new active substances (NASs) showed there were just 36 NASs in 2002—the lowest number in twenty years. (See “Is the Pharma industry weathering the ‘perfect storm’?” March 18, 2003, at: http://www.ims-global.com/insight/news story/0303/news story 030317.htm.) Yet another indication of declining productivity is increasing development times. In its review, IMS found that among the 33 NASs it was able to review, the shortest development time was Þve and one-half years, while the longest was more than twenty-three years. The IMS study showed that biotechnology NASs moved faster than traditional ones: four biotech NASs required only an average of eight years from priority patent application to product launch. Although biotech products may be moving faster through development than many traditional products, the Tufts Center for the Study of Drug Development recently reported that, overall, biopharmaceuticals—drugs based on DNA, RNA, or protein—are moving slower than they once did. Tufts found that the clinical phase for new biopharmaceuticals increased by 137%—from 31.2 to 74.0 months—when it compared approval times for the years 1982–1989 with those for 2000–2002. Pharmaceutical companies often blame the FDA for part of the slowness of the drug approval process, but according to Tufts, the mean approval time for new biopharmaceuticals has actually fallen 21% between the time periods of 1982–1989 and 2000–2002. (See “Biopharmaceutical product approval times dropped 21% since the 1980s,” 3/11/2003, http://csdd.tufts.edu.) In other words, products are being reviewed more quickly but are taking longer, on average, to complete the clinical trial phase. Compounding this situation is the fact that the rate of pharmaceutical sales growth is slowing. According to IMS Health, worldwide pharmaceutical sales rose only approximately 10% from 2001 to 2002, from $364.2 billion to $400.6 billion, compared with nearly 15% growth over the prior year. (In 2000, worldwide sales totaled $317 billion.) Also, the bulk of sales relies heavily on blockbusters that will go off patent over the next few years. Spending on new drug discovery technologies, meanwhile, has been increasing, creating an untenable situation in which many, if not most, big pharma companies are struggling to get sufÞcient drugs to market to maintain Wall

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Street’s required growth. The Pharmaceutical Research and Manufacturers of America (PhRMA) estimates that pharmaceutical R&D spending grew from $11.5 billion in 1992 to $30.5 billion in 2001. So, are new technologies actually failing, or does this innovation gap represent just a phase shift—a period between their introduction and optimal use? Having invested heavily in technologies and seeing little progress, the pharmaceutical industry is now wary of technology’s often overblown promises. However, the industry continues to show a strong interest in KM. As with other technologies and services, “make or buy?” is the critical question. A Natural Fit for Knowledge Management? In the following sections, we discuss the factors prompting pharmaceutical companies to reexamine the assembly, sharing, and decision making involved in drug discovery and development. In this industry, decision making at the management level is usually individualized, in part because many managers are also scientists and individual experience usually plays an important role in making critical assessments in the Þeld of biology. Pharmaceutical companies, however, would like to be able to distinguish between decisions that are data-driven and those that are based on other factors, such as intuition. Knowledge Management in R&D. First, the pharmaceutical industry must rely heavily on data, and the number of data-generating technologies used by the industry continues to grow. Recent analyses of the industry recommend fundamental changes for the improvement of the systematic capture, integration, distribution, and application of information in the early stages of research and development (R&D). Figure 1 illustrates the many types of data, information, and knowledge that are gathered in the pharma industry, the steps in which they are gathered, and how the value of the information increases as you go forward. It is through the extensive capture of many types of data, analysis, tracking, and management that companies bring products to market in this industry: the drug discovery and development process is essentially a continuous ßow of information, punctuated by critical decisions. Bringing a single drug to market is a long-term project that usually takes Þve to Þfteen years. During this time, technology and staff changes take place, and knowledge about a drug target rapidly evolves, making it difÞcult to streamline the process. The situation is further complicated because the technology pipeline, unlike the approval process, is not linear. Researchers from the two key areas of chemistry and biology may not have access to each other’s data in an understandable way. As the number of genomic research technologies used by the industry increases, the collaboration problem worsens because, typically, competence in new technologies starts at a few physical locations that are separate from more mainstream research facilities. There is also a high degree of specialization among researchers, which leads to a silo effect and is a major barrier to data sharing, collaboration, and productivity.

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FIGURE 1. Data, Information, and knowledge.

Many pharmaceutical and biotechnology companies also have multiple research sites around the world. Thus, language, cultural, and geographic barriers may also impede sharing and collaboration. Adding to the need for KM in pharma R&D is the fact that the discovery process is changing. In the past, many discoveries were the result of serendipity or minor modiÞcations to an existing drug. There is growing consensus that the low-hanging fruit have all been picked and that it will require all the hightech tools R&D departments have, as well as truly systematic approaches, to generate the next wave of pharmaceutical products. Although serendipity and modiÞcations to current therapeutics will still play a role in this new era, they will no longer be driving factors but will themselves be driven by the systematic exploration of data. Finally, a huge investment goes into every pharmaceutical product. Tufts estimates that, including the cost of failures, it requires more than $850 million today to bring a drug successfully through to market. The most recent drugs launched by the ten largest pharmaceutical companies actually cost nearly $2 billion apiece to develop. Because the cost of failure is the key driver of cost per drug approved, minimizing failure is one of the major goals for the pharmaceutical industry going forward, and this goal requires very close attention to managing resources—most of which exist as information. As we discuss later KM can play a major role in addressing the challenges of pharmaceutical R&D. Knowledge Management in Business Development. Over the past thirteen years, nearly 20 major mergers or acquisitions have occurred in the pharmaceutical and biotechnology industries (Jurgen D, 2003). Licensing is also a very common occurrence in these industries. A key issue at these times is how

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decisions are made. Companies negotiating for rights to a particular product or seeking to merge will greatly beneÞt by having a clear understanding of how their prospective partners have been making decisions and where the evidence comes from to support those decisions. In particular, companies must clearly understand their partner’s intellectual assets. Once again, as we will discuss, KM can help companies address this challenge. KNOWLEDGE MANAGEMENT TOOLS: FINDING TECHNOLOGY THAT IS APPLICABLE KM tools come in a wide variety, and more than 60 companies offer KM software and services. A KM tool can be something as basic as a system that stores documents (i.e., document management systems, or repositories) or as complex as a data-mining algorithm. Most of this KM software is for data acquisition, which is a critical process, because it is likely to involve the greatest volume of data. It is also important that information be stored so that it is complementary to the organization’s domain model—that is, the key characteristics associated with the speciÞc functions of an organization. Therefore, data need to be stored in a way that allows easy analysis and access, both internally and by select partners. Generally, the more sophisticated the data capture or storage tools, the more customization it will require to Þt a particular company’s needs. Table 1 lists vendors of document management software. TABLE 1. Select Document Management Software Vendors Company

URL

Accelrys Acero

www.accelrys.com www.acero.com

Biosift Cadenza Cognigen Cognos Document Control Systems

www.biosift.com www.cadenzinc.com www.cognigencorp.com www.cognos.com www.mastercontrol.com

Documentum Entropia High Tower Software ID Business Solutions Imaginatic Ingenuity Systems

www.documentum.com www.entropica.com www.high-tower.com www.id-bs.com www.imaginatic.com www.ingenuity.com

LabBook Open Text Software Scientific Strategic INFO Thermo LabSystems

www.labbook.com www.opentext.com/pharmasolutions www.scisw.com www.strategicinfo.com www.thermolabsystems.com

Product SeqStore Genomics Knowledge Platform Scintilla KnowledgeLEAD Repository Cognos MASTERControl FDA Edition EDMS Entropia Quantum High Tower Software ActivityBase Idea Central Ingenuity Knowledge Management Solution eLabBook LiveLink CyberLAB ChemX eRecordManager

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TABLE 2. Select Data Mining and Visualization Software Vendors Company BioDiscovery Entrieva Genstruct LeadScope Lion Bioscience Omniviz Partek PubGene Recommind Silicon Genetics Spotfire SPSS Verity Virtual Genetics Xpogen

URL www.biodiscovery.com www.entrieva.com www.genstruct.com www.leadscope.com www.lionbioscience.com www.omniviz.com www.partek.com www.pubgene.com www.recommind.com www.silicongenetics.com www.spotfire.com www.spss.com www.verity.com www.vglab.com www.xpogen.com

Product Imagene Semio product line Genstruct Discovery System LeadScope SRS Omniviz Partek Pro PubGene MindServer GeneSpring DecisionSite LexiQuest Verity K2 Enterprise Virtual Adapt, Virtual Predict PathLinX

The next stage after data acquisition involves data analysis and visualization, which are used to draw information out of the data that have been collected. A huge variety of KM tools exist for this purpose, and different departments in the same company often use different analytical tools, even if they are doing the same task (e.g., analyzing microarray data). This software can be grouped under the general heading “data mining.” Table 2 lists vendors of data mining and visualization software. In pharmaceutical R&D, tools that can work with multiple data types are sorely needed, as is modeling software to present the data. After the data have been analyzed, decision support and groupware KM tools can be used to draw knowledge and make decisions about the information. KM directories can also be used to help keep track of tools, people, and access to data. Collaboration is obviously a key aspect of optimal KM, and, again, one of the most important factors is determining the domain model. How are things labeled? How are they related? If things are not indexed properly and consistently, collaborators cannot communicate efÞciently. Table 3 lists vendors of decision support and groupware tools. Another set of KM tools, including project and portfolio management applications, are aimed at project leaders and managers. Project management systems generate reports about the vital statistics of project-related tasks. They help managers plan, track, and assess projects. Portfolio management systems can be used to monitor projects and entire program portfolios. Table 4 lists vendors of project management and portfolio management software. Finally, tools such as portals can be used to set up an infrastructure for KM. One of the key goals of such projects is, after all, to provide comprehensive and yet selective access. A portal does not seem like an obvious KM tool at Þrst, but it can be very effective at distributing information and making it available to the decision makers who reside at different physical sites. Table 5 lists vendors of portal software.

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TABLE 3. Select Vendors of Decision Support and Groupware Tools Company

URL

ActiveStrategy

Product

www.activestrategy.com

ActiveStrategy enterprise Cadent Technologies www.cadent.com Cadent Secure Share Click2learn www.click2learn.com Aspen Suite Cognos www.cognos.com Cognos Documentum www.documentum.com eROOM Groove Networks www.groove.net Groove Workspace IBM www.ibm.com Lotus Notes iCohere www.icohere.com iCohere Workgroup Suite MedTower www.medtower.com OMNI Meridian Knowledge Solutions www.meridianksi.com Knowledge Centre Microsoft www.microsoft.com Exchange Niku www.niku.com Niku6 OpenText www.opentext.com/pharmasolutions LiveLink SiteScape www.sitescape.com Enterprise Forum Tacit www.tacit.com Tacit Knowledge Mail 3rd Millennium www.3rdmill.com Custom development and integration services Vertical*i www.verticali.com Holding LEA

TABLE 4. Select Vendors of Project Management and Portfolio Management Software Company Adis International Microsoft Niku PlanView

URL www.adis.com www.microsoft.com www.niku.com www.planview.com

Product Adis Zenith MS-Project Niku6 PlanView

Wyeth, for example, has implemented a portal that provides information about the progress of clinical trials. There is no conÞdential information about patients or the actual drug accessible through this portal so the security issues are less critical than they would be otherwise, but those with access to the portal can learn valuable information about the status of a particular trial. With the “click of a button,” investigators can Þnd out, for example, if enrollment is still open or closed, and managers can determine very quickly how trials are progressing, thereby making it easier to address manageable problems, such as slower than anticipated recruitment, early on. IMPLEMENTING A KM PROJECT In considering how to implement KM projects, it is useful to Þrst examine what makes the pharmaceutical and biotech industries different from most others.

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Uniqueness stems primarily from the industries’ data concerns, which are especially complex and fundamental. KM can be a key success factor in managing these data. In the pharmaceutical industry, there is not only the problem of getting the data organized but also the challenge of setting up a decision process fed by the data. Essentially all of the industry’s knowledge is built from research data, and there are multiple data-generating processes with varying reliability and reporting methods. The key problem is not the data volume. Managing the huge volume of data is simply a matter of planning and acquiring the right IT to scale up the IT infrastructure. Rather, the real problem lies in the complexity of the relationships that exist between the data that cloud its meaning and interpretation. Different organizations or subÞelds also make different simplifying assumptions of the underlying scientiÞc concepts when they generate data, making data integration difÞcult. Even similar data, such as gene expression measurements, can be generated by multiple means, and often the source of the data, all the way down to the name of the person who did the experiment, is relevant to its validity. KM can help integrate the different kinds of data. The pharmaceutical and biotech industries are also highly regulated, so there needs to be clear and consistent documentation and audit trails. Because of new concerns about genetic privacy and the effects of regulations such as the Health Insurance Portability and Accountability Act (HIPAA) and 21 CFR Part 11, this need will become more important and extend further downstream into discovery. There is a growing sense that having access to the stream of data that precedes any decision, from cellular assays through the approval of a drug and beyond, may be necessary in the future regulatory environment. Another important aspect of today’s pharmaceutical R&D process is that as a company moves upstream from the data (toward the market), two things happen: more types of data need to be integrated, and feedback loops need to be established at critical junctures to guide the data-gathering process—another potential role for KM.

TABLE 5. Select Vendors of Portal Software Company ActiveStrategy Autonomy IBM InfoStrength Paper Thin Perceptive Informatics Plumtree Quiver SAP

URL

Product

www.activestrategy.com www.autonomy.com www.ibm.com www.infostrength.com www.paperthin.com www.perceptive.com www.plumtree.com www.quiver.com Mysap.com

ActiveStrategy Enterprise Portal-in-a-Box Lotus K station InfoStrength Portal Common Spot Content Server Perceptive Portal Plumtree content servers Quiver knowledge Suite mySap.com

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In the next section, we describe the basic steps in organizing, or reorganizing, a KM process; the key factors that will make such an initiative successful; common hurdles; and how to evaluate KM projects. Taking a Process Perspective on the Flow and Use of Information For each step in developing a KM process, the right tools must be selected and properly implemented, and there must be a clear means of measuring a project’s success. These steps and considerations seem simple enough, but it is easy to derail a project by not paying enough attention to them. InÞnity Pharmaceuticals offers an example of a successful KM process. The company has placed major emphasis on KM, speciÞcally, bridging the barrier between chemistry and biology to accelerate drug discovery. After examining how these two groups work, InÞnity designed elegant systems to connect them and to enhance both inter- and intradivision sharing. One goal of InÞnity’s KM system was to link all the biological information associated with a compound to its chemical information. The company kept in mind that the two groups have very different work practices: chemists tend to think in terms of reactions, while biologists deal with more unstructured information and need more open and ßexible templates. The core of InÞnity’s knowledge-sharing strategy is having a standard vocabulary that both groups understand and an easy-to-use Web services system with “dashboards” developed using Microsoft.NET and delivered through a standard portal framework. The dashboards are designed so that scientists can access tools and data speciÞc to their areas of expertise. For instance, a chemistry Web services dashboard provides access to a reaction database, while a biology dashboard displays the results from many assays. They also have a “partner” dashboard, which lets partnering companies collaborate with InÞnity over the Web. InÞnity can control which partners have access to certain applications and data. Another goal at InÞnity was to “capture” as much data as possible. Some of the most common types of data never captured in pharmaceutical R&D are also some of the most basic—details of how experiments are performed and conclusions or inferences drawn about the results. At InÞnity, in addition to storing information about the basic structure of compounds, all experimental results are captured, including the synthetic reactions, purity of the product, and experimental conditions. Results are captured differently at InÞnity than at other pharmaceutical Þrms. Typically, companies record experimental results in paper-based laboratory notebooks. InÞnity, however, is using electronic notebooks (the “E-Notebook”) from CambridgeSoft. The E-Notebook is designed for chemists, but its design is ßexible enough to provide InÞnity biologists with similar collaboration and analytical beneÞts. InÞnity scientists have collaborated with CambridgeSoft on enhancements to this product. When a problem has been well deÞned, this kind of collaboration between vendors or system integrators and pharmaceutical or biotech companies is one of the most fruitful ways of developing optimal tools. For

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example, Millennium Pharmaceuticals has built its KM system in collaboration with Ingenuity Systems, and Genome Therapeutics Corporation has built its KM system in collaboration with 3rd Millennium. Bayer has multiple collaborations, with companies such as Lion Bioscience, SciTegic, and Acero. Meanwhile, Tripos, Accenture, and Bristol-Myers Squibb (BMS) worked together to develop BMS’s SMART-IDEA (Structure Modeling and Analysis Research Tool—Integrated Data for Experimental Analysis) drug discovery system. SMART-IDEA uses Accenture’s ICE3 (Integrated Computing Environment for High-Performance Discovery) and software from Tripos. The goal of the project, which serves 2000 drug discovery scientists at BMS, is to achieve a 10–15% reduction in cycle times and a 10–15% increase in success rates. Keys to Success As noted earlier, KM can be used to address a wide range of problems. Probably the key challenge in the pharmaceutical and biotech industries today has been the increase in the complexity and, correspondingly, in the need for interpretation of data. Many KM projects address this particular challenge, although it need not be the major focus of such a project. Clearly Defined Goals. The Þrst step to success in any KM project must be to clearly deÞne the goals of the project and to do so within the context of the organization. This task requires identifying the problem and determining what kind of change will address it. To decide on a “path to change,” it is important to think through the potential solutions and see how each one would help or hurt the status quo. Because no company can make all necessary changes, managers must determine which changes will bring the greatest return on investment. Often, a remarkably simple solution will deliver relatively more beneÞts than a complex one. Table 6 lists the steps to identifying the problem. It is also best if the new solution does not require new processes. KM is about optimization and using technology to improve collaboration and integration, not about creating new projects for already overtaxed staffers. Table 7 lists the steps to thinking through potential solutions. To make these choices, companies must consider the input of advisers and look at the options without having any particular bias. Sometimes, those kinds of advisers can be found within a company’s own ranks; at other times, it is necessary to hire external advisers. However the decision is reached, multiple options should be considered, weighed, and compared to ensure that the company is addressing the right problem in the right way. The Right Technologies. It is usually best to “start small and local” and build the KM project up over time, especially for organizations with little prior experience with KM. Therefore, choosing the right technologies to use is another important key to success. It is critical to make sure the technologies either Þt with existing systems or can be made to Þt relatively easily. It is also useful to run a pilot project on a small scale and then quickly elicit feedback. One of

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TABLE 6. Identifying the Right Problem An organization’s KM needs can by analyzed using a map of the key processes guiding the targeted group’s work. 1. Find where information can be used more effectively. The following areas often require improvement: • Communication between individuals and/or groups, particularly those with different functions. • Time-consuming processes with the sole purpose of acquiring or transforming data to support downstream analyses. (For example, before SMART-IDEA was implemented, Bristol-Myers Squibb said that its scientists spent almost as much of their time reformatting and moving information between analysis packages as they spent analyzing it.) • Redundant data being captured in incompatible information systems. • Procedural inconsistencies across identical functions. • Ad hoc information gathering solely to provide status updates for management. • Barriers to cross-functional information. • Knowledge that is generated without being captured. 2. Determine if solving the problem will make a measurable impact. • From an organizational perspective, ask the following questions: How is the value chain affected? What feature will ‘‘sell’’ management on the initiative? Does it speed processes? Does it improve quality? Does it provide a competitive advantage? • From the employees’ perspectives, ask the following questions: Where is the greatest frustration and pain? What does the initiative do for them?

the primary issues regarding any technology choice is the question whether to make, buy, or both. All of these factors will have a great deal of inßuence on the outcome of the project. Careful Deployment. Finally, the success of a KM project will also be shaped by how well it is deployed. Successful deployment is a matter not only of having a plan and making sure that it is adhered to properly but also of creating an environment that focuses on user adoption and promotes the use of the system by creating incentives or interest around the initiative. Table 8 lists the steps for successfully deploying a KM project. Common Hurdles In implementing pharma/biotech KM, companies may encounter some common problems. One such problem is the difÞculty of clearly deÞning the likely return on investment and its beneÞt to the company. This task is particularly difÞcult in an industry with such long product development times. Millennium Pharmaceuticals and CuraGen, for example, are KM pioneers that have clearly achieved what they set out for, in terms of acquiring many novel targets. However, that particular achievement has not yet translated into commercial success. Both companies

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TABLE 7. Thinking Through Potential Solutions 1. Identify a leader for the initiative. 2. Gather information. • Have a guided discussion. • Identify all the important players. • Examine how the information flow can be improved. • How is the information currently flowing or not flowing? 3. Evaluate potential solutions. • How can the process be optimized? • Can an old process be augmented? • If the process needs to be redesigned, what is the least disruptive way to do so? 4. Focus on the user. 5. Consider the constraints carefully.

have generated “many” targets and clinical candidates, and CuraGen’s system has even allowed the company to Þle its Þrst investigational new drug application entirely electronically, but it will be awhile before it is clear that KM has helped them commercially. (This commercial issue is a major one, and we discuss it in more detail in a later section.) Another potential problem is legacy systems and the difÞculty in building on top of or around them. Most KM projects involve IT on some level, and IT involves legacy systems. This hurdle is a tough one, particularly for large companies with many sites. Other potential obstacles are as follows: •

As noted earlier, many pharmaceutical companies have business silos that do not communicate well with each other and do not understand each other’s work.

TABLE 8. Planning for Successful Deployment • Focus on user adoption. Load the system with relevant information before giving it to users. Train and mentor users. Develop a mechanism to ensure questions are answered. Encourage feedback and rapidly incorporate it. • Reward information sharing. • Promote the benefits and impact of the project. • Create incentives to encourage use of the tools. • Lead by example. • Measure the outcome and impact of the project. • Measure the right thing. • Learn from the result.

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•

•

• •

401

There is a great deal of tacit knowledge in the pharmaceutical industry, and much of it is extremely difÞcult to convert to explicit knowledge, a job that is one of KM’s fundamental goals. Data integration, particularly in the age of data overload from genomics, high-throughput screening, and combinatorial chemistry, is extremely challenging. Staff concerns that a KM system will reduce their value as employees: “If I put everything I know into the system, the company will not need me.” Staff and management concerns that a KM system will remove their ability to make independent decisions or that it will highlight the bad decisions they make. For example, according to a manager of a core microarray facility at a major biotech company, many scientists continue to pursue targets that do not look promising based on microarray study results.

Potential Solutions Each of these hurdles can sometimes require a speciÞc solution, but there are general strategies for addressing several of these problems as well. Focus on Behavior. The one consistent message from experts who have successfully implemented KM projects is to focus on behavior as much as, or more than, on technology implementation. For example, Claire Hogikyan, worldwide head of information management for PÞzer Global Research and Development (GRD), reports that PÞzer GRD’s KM portal solution focuses 20% on technology and 80% on “behavior changes.” She emphasizes that KM is a “business solution, not an IT solution.” (See “PÞzer: knowledge management—more than half the battle is behavioral change.” www.eyeforpharma.com/index.asp?news=27940.) Doing a lot of work to understand how people work and what they will expect from the system is essential. Both PÞzer and Ilex Oncology addressed this issue by conducting workshops with employees. Ilex was implementing a new system to manage all the information and data (including Þnancial information) related to clinical trials. In addition to meeting regularly with the staff, the group setting up the system also met regularly with managers. If problems started recycling in the staff meeting (i.e., coming up, being addressed, coming up again) the managers were simply told to make a decision, once and for all. Hire Staff Skilled in KM. Many people still believe that a software system can do all the work for a KM project. Experience shows that this is not the case, particularly in an established organization where new habits must be employed. As a result, at larger companies, large-scale KM also requires dedicated staff with specialized skills. Millennium Pharmaceuticals and AstraZeneca, for example, have both found that to capture the maximum information and to do so in a consistent manner, they need KM staff, who either attend meetings and make sure the results are put into a database or who write up information and have it reviewed by experts for

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inclusion in the database. It is not reasonable, at least not at the start of a KM project, to expect the entire scientiÞc staff to begin feeding the system exactly the way you want it fed. Establish and Enforce Standard Nomenclature. CuraGen has staff in charge of enforcing the company’s nomenclature. This standardized nomenclature is the linchpin of its KM system, allowing rapid retrieval of all information pertinent to a CuraGen project. However, as science evolves, so does nomenclature, and scientists continually try to use their own, favored terms. Without enforcement, the nomenclature would quickly be corrupted. Nomenclatures are key to most pharmaceutical and biotech KM systems. Bayer and Roche, for example, are initiating KM systems simply by establishing standardized terms for identifying genes and then linking all information related to a gene. Bayer has already demonstrated the power of standardization; simply by using common terms, two major data centers (one in Kyoto and one in England) were able to share data. Companies like Acero have built object models that provide a context for linking scientiÞc terms. Also, some established public nomenclatures exist that can be useful starting points. Create Knowledge Directories. It is also important to recognize that a lot of knowledge cannot be reduced to something that can be captured. Doing so would require so much work that it is much more efÞcient simply to point people to the correct source. For example, rather than capture the methodology for every single biological assay performed, it would be useful simply to have a directory that can guide people to colleagues who have worked on similar problems. For example, InÞnity, at one point, tried to “distill” chemists’ decisionmaking process in selecting compounds for further development. However, the information InÞnity gleaned from this exercise was hard to generalize or act upon; each chemist seemed to have his or her own decision-making approach. Get Management Support. Finally, it is essential to have the support of management and for management to demonstrate its conÞdence in the system. At Vertex Pharmaceuticals, this approach played a key role in making the KM system successful. As employees were made aware that management was using the system and getting results from it, they became more enthusiastic about using it. Judging Success Naturally, in these days of budget scrutiny, management is always the most interested in knowing whether a project is worth the investment and what the beneÞts will be of a speciÞc initiative. The value of KM projects can be difÞcult to measure because sometimes investment returns come in the form of simply making it easier for people to do their jobs. If productivity is being curtailed by other factors, such as inexperienced staff, inadequate resources, or just plain

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unsolvable research problems, then the KM solution may seem ineffective. Ideally, a project’s success should be evaluated in several ways, in which case it will be clear if the KM project has solved the intended problem and if solving that problem has had the intended effect. In the early stages of a project, the measures of success are often extremely qualitative (e.g., people say that it is easier to do their work). The goal should be to move toward parameters that can be measured and that accurately reßect the utility of the project. For example, if the goal is to increase connectivity and the solution is a directory, the company can track the number of times employees access the directory and their impression of its utility. However, it would be useless to measure a project’s success by counting the number of entries in the database. After all, if it contains incorrect names and nobody uses it, the database is essentially worthless. Unfortunately, such counting is often how companies evaluate such projects. The Þrst, and most critical, milestone in any KM project is staff acceptance. Unless staff are using the system, it cannot be effective. PÞzer’s Claire Hogikyan purposefully started off the company’s KM portal project with a broad generic content that would appeal to a wide range of users. The PGRD gPortal is essentially a workspace on the intranet where divisions can easily communicate, collaborate, and share. The aim was Þrst to show staff that the portal could help staffers, even in a small way. In the long term, PÞzer hopes to make this portal not just a groupspace but also an individual workspace where employees can access speciÞc tool sets depending on their needs. However, because each business group has different priorities and concerns, PÞzer is undertaking a more individualized approach in which it focuses, business unit by business unit, on each unit’s speciÞc needs. At this point, the project is largely about behavior and change rather than about technology. Millennium Pharmaceuticals also keeps track of the number of employees that access its KM system, but the company is far enough along that it is starting to look for more tangible results. Its KM system pulls together a huge amount of internal information and links it to popular external sources. One of the tools at the center of the system is its Pathway Resource and Information System (PARIS), the company’s pathways analysis and visualization software. Millennium recently reported that one of its investigators had used the system to identify potential molecular mechanisms underlying resistance to Velcade (bortezomib), the company’s novel, prospective cancer drug, which is being evaluated by the FDA for the treatment of multiple myeloma. By comparing the gene expression pattern of a patient who did not respond to the drug with the gene expression patterns of the many patients who did and then examining which genes were differentially expressed, the scientists identiÞed some reasonable molecular mechanisms that could explain the difference in response. Going forward, such examples (particularly well-validated ones) will be necessary to keep pharmaceutical and biotechnology companies interested in KM tool development. Ultimately, these companies will be looking for increases in

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productivity and proÞtability. As a result, managers of KM products should establish, in advance, key milestones and parameters—such as the number of regular users, the money saved, or the time saved doing a particular step in a typical process–that can be used to deÞne success. OUTLOOK KM has received mixed reviews in other industries, most likely because it is a complex process that is widely misunderstood and often misapplied. The pharmaceutical and biotech industries should not be discouraged from pursuing KM. Because of their data-rich processes, long project timelines, and complex organizational structures, these industries stand to beneÞt signiÞcantly if they implement KM projects properly. Indeed, we believe KM will be a deÞning feature of successful pharmaceutical and biotech companies in the future. The explosion of data these industries are experiencing cannot be handled any other way. Both small and large companies, including Bayer, Boehringer Ingelheim, InÞnity Pharmaceuticals, Millennium Pharmaceuticals, PÞzer, and Roche, are pioneering KM projects, and we anticipate that many more companies will join their ranks over the next Þve years. REFERENCE Jurgen D. Strategic trends in the drug industry. Drug Discovery Today. 2003;8(9):411–420.

The EU Clinical Trials Directive: Impact in Europe and Beyond

SUMMARY The European Union (EU) Clinical Trials Directive (CTD), implemented on May 1, 2004, has caused considerable controversy—particularly among the noncommercial research community—because of the additional cost and administrative burden that it has created. The ultimate beneÞts of the legislation remain to be seen: Will it make the EU more or less attractive for clinical trials? This article examines the background and aims of the CTD as well as the ways in which it is changing the roles of national regulatory agencies and ethics committees. It analyzes pharmaceutical companies’ experience with the new Eudract and Eudravigilance databases as well as the GCP and GMP requirements. It concludes with an assessment of the CTD’s potential for achieving its goal of standardization across the EU, including the new accession countries of Eastern Europe. BUSINESS IMPLICATIONS •

The new European Union (EU) Clinical Trials Directive (CTD) aims to promote patient safety and standardize trial application procedures to competent authorities (CAs) and ethics committees in each EU member state. The commercial intention of the CTD is to speed regulatory submissions and recruitment rates in the EU, thereby increasing the region’s competitiveness globally. Standardization of application procedures would clearly beneÞt large pharmaceutical companies running multicountry trials. However, it also entails additional bureaucracy and cost. This additional burden affects large pharmaceutical companies, but it is more onerous for smaller companies and biotechs.

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

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The CTD makes no distinction between healthy volunteers enrolled in Phase I trials and patients in later-stage trials, so Phase I trials must now be registered with both the national CAs and ethics committees. Although this requirement may prompt some large pharmaceutical companies to move their trials out of the EU in the short term, most of these companies will still need to conduct later-stage trials in the EU if they want to launch their drugs there and build acceptance of them. The EU is in a transition stage, with many countries yet to implement the CTD. Despite the short-term disruption, many trial sponsors believe that the CTD will not damage the EU’s attractiveness as a trial location and may even enhance it in the longer term. We believe that the beneÞts of standardized application procedures will at least balance, or even slightly outweigh, the additional paperwork and cost. Thus, the overall impact of the CTD on pharmaceutical companies should be neutral or slightly positive once the transition phase is complete.

INTRODUCTION The European Union (EU) Clinical Trials Directive (CTD) was implemented on May 1, 2004. Prior to May 2004, no European legislation governed the conduct of clinical trials, and the trial requirements of member states differed signiÞcantly. (The International Conference on Harmonization of Technical Requirements for Registration of Pharmaceuticals for Human Use (ICH)—though widely adhered to—is only a voluntary, rather than a legal, code of practice.) The CTD is the Þrst EU attempt to legally protect patients participating in clinical trials. In addition, the CTD is intended to standardize application procedures for clinical trials, a goal that (if achieved) should simplify the registration process for pharmaceutical companies conducting multicountry trials across the EU. Despite the potential beneÞts of this new directive, the CTD has caused considerable controversy since it was implemented—particularly among the noncommercial research community—because of the additional cost and administrative burden that it has created. We begin by examining the background and aims of the CTD and the ways in which it is changing the roles of national regulatory agencies and ethics committees. We then analyze pharmaceutical companies’ experience with the new Eudract and Eudravigilance databases as well as the GCP and GMP requirements. We conclude with an assessment of the CTD’s potential for achieving its goal of standardization across the EU, including the new accession countries of Eastern Europe, and whether its net effect will be to make the EU more or less attractive for clinical trials. OVERVIEW OF THE CTD The European Commission’s (EC’s) CTD—Directive 2001/20/EC—took many years to appear, as shown in Figure 1. The EC recognized from the outset that

OVERVIEW OF THE CTD

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FIGURE 1. EU clinical trials directive: a gradual evolution.

differences would remain—hence the use of the term approximation rather than harmonization in the CTD. A directive is, by deÞnition, less restrictive than a regulation, as each member state must transpose it into its national law but is free to choose the manner in which it does so. The scope of the CTD is limited: it excludes medical devices but does apply to the medicine component of a combination product. Noninterventional trials, which involve products with a marketing authorization that are prescribed in the usual manner, are also excluded. The CTD covers only studies that are undertaken to ascertain the efÞcacy or safety of a medicine in human subjects. In the following sections, we review the goals and potential concerns associated with the two objectives of the CTD: (1) to guarantee patient rights and protection in clinical trials and (2) to provide greater standardization of national regulatory procedures.

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Patients’ Rights Requirement Patients’ rights are now guaranteed in a legal framework, such as the privacy and protection of their personal data, their right to receive full information about their trial, and the obligation of sponsors to include certain information in trial advertisements. Concerns have arisen, however, that the CTD will slow patient access to new drugs, given the risk that the extra paperwork involved will slow the registration process and the actual trial. This criticism of slower patient access may, however, turn out to be short-lived, at least for commercial trials—once the industry has understood the new application procedures and the EC’s systems are Þrmly embedded, delays should be minimized. Noncommercial trials, which are often less well resourced, may continue to struggle with delays. Another potentially longer-lasting concern is that these complex patient registration procedures may mean that certain types of trials—for example, trials of new combinations of existing drugs and extra indications that would not result in commercial gain—may not be conducted, limiting practical, useful research. This scenario could lead to more off-label prescribing than before, using combinations that have not been ofÞcially evaluated in clinical trials. The CTD also outlines the need for patient consent (or, for an incapacitated adult, consent from a legal representative). This requirement could complicate trials using emergency treatments, where this consent may be difÞcult to obtain quickly enough. Another concern raised by many are the implications of Article 5, clause (i) of the CTD, which states that trials using incapacitated adults may be conducted only if “there are grounds for expecting that administering the medicinal product to be tested will produce a beneÞt to the patient outweighing the risks or produce no risk at all.” This focus on the individual patient could be problematic for researchers because by deÞnition the question of beneÞt cannot be answered prior to the trial. Standardization of Regulatory Procedures Standardization of application procedures across the EU, the other main aim of the CTD, would clearly beneÞt large pharmaceutical companies running multicountry trials and also smaller companies conducting trials in rare diseases, for which trials must take place in several member states to recruit enough patients. The commercial intent of the CTD is to speed regulatory submissions and recruitment rates in the EU, thereby increasing the region’s competitiveness globally. One of the most potentially beneÞcial areas of standardization is the CTD’s requirement for all ethics committees across the EU to review trial applications within 60 days—previously, these reviews could take several months, delaying patient recruitment and causing frustration for companies coordinating multicountry studies. Doubts have been raised, however, as to how well the CTD will succeed in standardizing trial applications, especially given that member states’ freedom

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to implement an EU directive however they choose will guarantee considerable variation. Also, the standardization that does occur will be of little beneÞt to the numerous small, single-country trials, which include most noncommercial studies. A curious feature of the CTD is that its regulatory requirements are similar but not identical to the guidelines set by the ICH, which since 1997 has aimed to facilitate the mutual acceptance of clinical data by regulatory authorities in the EU, the United States, and Japan. The CTD now overrides the ICH in the EU, a change that many in the pharmaceutical industry Þnd irritating, citing the fact that the CTD shares most of the ICH’s principles but requires new sets of paperwork. It remains unclear why the EU did not adopt the ICH’s principles in full, given that the ICH enjoys the approval of the United States, the EU, Japan, and the World Health Organization (WHO). The CTD’s differences from the well-known ICH mean that everyone affected now must change the practices and paperwork that support their ICH-sanctioned approach, thereby slowing and perhaps impeding some of the standardization that the CTD hopes to achieve. NOTABLE FEATURES OF THE CTD Several notable features of the CTD represent substantial changes in the way clinical researchers plan, organize, and conduct their trials. These features include the changing roles of national competent authorities (CAs) and ethics committees, the introduction of the Eudract and Eudravigilance databases, and the CTD’s requirements for GCP and GMP. Competent Authorities The CTD stipulates that every EU member state must have a national regulatory authority, known as a competent authority (CA), to assess every trial application. Trial sponsors must submit an array of paperwork. For many member states, this paperwork requirement is signiÞcantly larger than prior to the CTD, placing a strain on the limited resources of both the CAs and trial sponsors, especially small companies and noncommercial researchers. On the positive side, the paperwork is now standardized across the EU, so companies submitting applications for the same trial in several member states need only have their documents translated rather than struggling to meet each member state’s different requirements. The impact of the CTD on trial sponsors will depend in part on how well equipped the CAs are to cope with the new processes and extra paperwork; for example, any “substantial” amendment to a trial’s procedure now must be approved by both the CA and the ethics committee. Researchers are taking care to word their trial protocols carefully at the outset, building a broad trial design and scope in order to avoid having to submit amendments for reapproval. The general consensus, according to Steve Smith, CEO of Origin Pharmaceutical Services and president of the Clinical Contract Research Association, is that the United Kingdom’s CA, the Medicines and Healthcare Products Regulatory

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Agency (MHRA), has been very efÞcient at managing the transition to the CTD and has maintained its speed and ßexibility, adjusting its fee categories. Not all EU CAs may be able to cope with the added resource pressure, however, at least in the short term. Ethics Committees The CTD aims to standardize the administrative functioning of ethics committees across the EU, making the trial approval process more transparent and consistent for multinational trials and aiming in particular to reduce delays. The main differences introduced are as follows: • •

A single opinion from both local and national ethics committees. The establishment of a 60-day timeline.

Sponsors now must submit documentation to both local and national ethics committees, after which the ethics committees must process trial applications and provide a single opinion within 60 days. Local ethics committees could previously reject a trial at speciÞc sites, which would delay the start of the trial until consensus could be reached. Trial sponsors must submit a wide array of paperwork to ethics committees. The CTD has created a greater administrative burden for ethics committees, just as it has done for CAs, and it is unclear whether all member states will be able to supply their ethics committees with the extra resources required to cope with the new workload to meet the 60-day time limit for each trial, which applies regardless of how many proposals a committee has received that week. Additionally, several member states are in the process of signiÞcantly reducing their number of ethics committees to enable each committee member to review approximately 20 applications per year, the minimum considered necessary for members to acquire and retain their evaluation skills. One organization tackling this issue is the European Forum for Good Clinical Practice (EFGCP), a nonproÞt that is trying to help member states bring their ethics committees in line with the CTD by encouraging committee members to communicate and share practices across national boundaries, in the belief that the issuing of EU directives alone will not achieve harmonization. Considerable differences among national ethics committees continue to exist, however, and it is very unlikely that the CTD will achieve full standardization of them in the foreseeable future. In The Netherlands and Italy, for example, ethics committees have signiÞcant scientiÞc responsibilities. Dutch ethics committees have ofÞcial governmental status and even receive copies of investigational medicinal product (IMP) dossiers, while the Dutch CA barely considers GCP issues. Similarly, in Italy, ethics committees have ofÞcial status in local government and assess both the scientiÞc and ethical aspects of trials. In the United Kingdom and Germany, on the other hand, the CAs conduct the scientiÞc review and the ethics committees are conÞned to ethics. In Germany, the role of the

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ethics committee is solely to examine the objectives and mechanics of the trial, with a clear distinction between scientiÞc and ethical review. These ongoing differences mean that companies must continue to manage varying ethical review procedures in multinational clinical trials. For ethics committees, another consequence of the CTD is that they are now legally recognized bodies under the authority of each member state’s Ministry of Health and can, therefore, be sued by a patient maltreated in an approved trial or by a sponsor if they ßout the time limit for approval. Many committee members are now concerned that they will need higher levels of training and administrative support so that they can do a sufÞciently professional job. Eudract The European Clinical Trials Database, known as Eudract, is a new online database established as part of the CTD. Sponsors apply for a study number through its Web site (http://eudract.emea.eu.int/) and then input the required data. It contains details of trial approval applications and any subsequent amendments, the ethics committee review, GCP and GMP inspection results, and all further trial data through trial declaration. The registration process has been found to work relatively well and to be straightforward for sponsors to use. The EC hopes that Eudract will provide a means of avoiding past problems that occurred when pharmaceutical companies chose not to publish unfavorable trial results, thus leaving national regulatory bodies to make recommendations based on incomplete sets of results. All trials must be registered in the Eudract database so that national CAs will be able to access all trial data. Eudract should also prevent trial investigators from changing the stated purpose of the trial as it progresses so that acceptable Þndings, rather than inconvenient results, can be published. A few minor issues have arisen around Eudract. Sponsors are concerned that, while they have to provide data for input into Eudract, they do not have access to the actual database. They fear that errors might occur during electronic data entry that cannot later be identiÞed and corrected. Questions have been raised about how to enact quality control to ensure accuracy and integrity at both sponsor and CA level. And who in the wider research community should have access to Eudract’s data? At present, only the national CAs, ethics committees, and the European Medicines Agency (EMEA) have access. Those conducting commercial trials are unwilling to relinquish their sensitive trial protocol data to anyone else, while noncommercial researchers and patient groups, such as Richard Sullivan at Cancer Research UK, are lobbying for wider access to enable thorough peer reviews of all results and to avoid trial duplication. It seems unlikely that the industry will make many concessions in this area. A Þnal concern is that a single multicenter trial can appear to the EC to be several different trials on Eudract because each national sponsor is registering their element of the trial with a different Eudract number. Such multiple entries are likely to distort the recruitment numbers, the validity of the data, and the

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number of separate trials conducted with a particular product. However, now that this problem has been identiÞed, the EC will likely Þnd a means to resolve it. Eudravigilance The Eudravigilance database (www.eudravigilance.org/human/index.asp) is being established to enable the electronic reporting of all suspected unexpected serious adverse reactions (SUSARs) encountered during trials. The EC has concluded that safety data must be reported and analyzed more efÞciently than is currently the case, to prevent drugs from reaching the market with insufÞcient safety records. On the plus side, Eudravigilance should enable the EC to remove an unsafe product from the market after just three days of real-time data analysis rather than three months of paper reports, as previously. The beneÞt for pharmaceutical companies, in theory, is substantial savings in terms of avoided corporate exposure and trial expenditure. Also, the data required for Eudravigilance are similar to the data required by ethics committees, so sponsors should not Þnd provision of the data too demanding. However, on the downside, the research community points to the cost and complexity of Eudravigilance and says those factors may discourage smaller trials. The signiÞcant IT requirements are noted as well; for instance, Steve Smith at Origin Pharmaceutical Services believes small companies will Þnd it difÞcult and expensive to connect with Eudravigilance. Also, although the concept of one-stop reporting of SUSARs to CAs is clearly a good one, some member states may choose to impose additional reporting requirements not covered by Eudravigilance and thus create more work for trial sponsors. The main controversy regarding Eudravigilance, as with Eudract, is the issue of who will have access to it. At present, only the CAs, EMA, and the EC have access. In contrast, in the United States, the government, in response to growing concerns about the adverse effects of unpublished clinical trials, has set up a database of all ongoing trials that is accessible to the general public (www.clinicaltrials.gov). Some executives in the U.S. pharmaceutical industry hope to improve their recruitment rates through this Web site, but European industry executives remain concerned about protecting their commercially sensitive trial data. GCP Among the CTD’s requirements for good clinical practice (GCP) are the need to assess and inspect the suitability of trial investigators, their supporting staff, and the quality of their facilities. Principal investigators must submit their qualiÞcations and GCP training or experience to local ethics committees. The beneÞt to patients is clear, but for trial sponsors the outcome is a considerable increase in cost and work. The EC has supplemented the CTD with a subsidiary GCP directive. This GCP directive has pleased noncommercial researchers because, unlike the original CTD, it explicitly acknowledges the beneÞt of noncommercial trials and also

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allows for a “variety of GCP arrangements,” giving noncommercial researchers more ßexibility in meeting GCP standards. One negative aspect of this GCP directive for researchers, however, is its assertion that no “unnecessary” trials should take place. Although trials may have undeniable clinical utility and be conducted to the highest standards, it is impossible to predict which trials will produce useful results and answer key clinical questions. Also, the sheer diversity of cultural and political views within Europe and the United States is likely to make full GCP compliance difÞcult for most trial sponsors. Furthermore, the CTD requirements will pose complications for EU trials of products approved outside the EU because, per the directive, such products’ original trials must have been conducted according to GCP standards. Some CAs may accept public-domain prescribing information, but it could be difÞcult for CAs to address safety issues fully if the sponsor does not have access to the manufacturer’s original documentation. Despite these minor issues, investigators do not claim to have encountered any problems implementing the CTD’s GCP requirements. Steve Smith’s response is typical: “GCP has not caused any signiÞcant problems for us, as we had always conducted trials to the very similar ICH GCP standards anyway. The only change has been that GCP has changed from being a voluntary code of practice to law.” GMP The good manufacturing practice (GMP) element of the CTD requires that the manufacturing site of clinical trials supplies be licensed and that a “qualiÞed person” (QP) be on hand to release each batch of clinical trials supplies. The QP thereby certiÞes that each production batch has been manufactured to standards similar to the FDA’s GMP regulations, which require that manufacturers take steps to ensure that their products are safe, pure, and effective. The investigational product will not have to undergo further testing if imported from one EU member state to another as long as the QP signs batch release certiÞcations. GMP is not seen as a major obstacle by EU-based pharmaceutical companies and is of beneÞt overall, given that it prevents researchers from mixing their own drugs, which could be extremely toxic. Some member states, such as Germany, have had a similar system for many years, but it is new to others, such as the United Kingdom. The main issue now affecting pharmaceutical manufacturers is the need for a QP. The shortage of QPs in several member states means that contract manufacturing organizations (CMOs) with their own QPs have enjoyed a surge in business since the introduction of the CTD. Chris Higgins, CEO of Penn Pharmaceutical Services, now employs four QPs, all of whom are kept fully booked auditing and releasing manufacturing sites for contract research organizations (CROs) or pharmaceutical companies. Higgins notes, “All this extra work for QPs has meant that the directive has a strong positive impact on companies like us.” GMP has created more complications for non-EU-based companies exporting products to the EU, as manufacturing sites outside the EU have to be audited.

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Products manufactured in the EU require only a copy of the QP’s original manufacturing authorization as part of the clinical trial application dossier, but manufacturing sites outside the EU require a new authorization to certify that they operate in accordance with standards equivalent to EU GMP. Exporters to the EU may encounter difÞculties as they seek to impose EU GMP standards on, for example, FDA-registered facilities, creating a substantial disincentive for them to conduct clinical studies in the EU. The EU’s GMP import regulations have also created signiÞcant Þnancial burdens on the industry that could displace Phase I and II studies in particular. North American companies manufacturing and exporting clinical trial products to Europe will need to employ EU-based QPs who know the differences between U.S. GMP and EU GMP, which, although based on essentially the same principles, differ in some details and paperwork. The FDA requires full GMP compliance only at Phase III; in the EU, full GMP is required from Phase I. The discrepancies between U.S. GMP and EU GMP raise the issue of mutual recognition of the two systems, which, according to Chris Higgins at Penn Pharmaceutical and unfortunately for industry, is likely to take at least another ten years. It is much closer to happening between the EU and Japan, which already have a mutual recognition agreement for commercial products, although it could still take several years for them to recognize each other’s trial products. The United States and the EU are still far from recognizing one another’s commercial products and are unlikely to recognize one another’s trial products in the foreseeable future. Mutual recognition agreements have also been made by individual member states for commercial products; for example, Germany has entered into such an agreement with Japan, and each member state can similarly set its own criteria for importing trial products. Five member states in the EU, including Germany and Denmark, require a complete retest program for trial products from outside the EU, which means that it is easier to import these products into the United Kingdom, which is considerably less strict, than to move them around freely within the EU. This issue of different member states’ retesting requirements is one of the main areas of difference in GMP across the EU. AREAS OF NEGATIVE IMPACT In two areas, the CTD has a particularly negative impact on commercial and noncommercial researchers: the additional overall bureaucracy and cost created by the CTD’s range of requirements (outlined in the previous section) and Phase I trials, which are now regulated under the CTD with the result that some EU members are losing their previous global competitive advantage for these early trials. Additional Bureaucracy and Cost For EU-based pharmaceutical trials, the CTD poses difÞculty in terms of the additional bureaucracy and cost that it entails; trial sponsors must devote time

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and resources to master the new application processes and the Eudract and Eudravigilance databases and to ensure that they comply with the new GCP and GMP requirements. This additional burden is less signiÞcant for large pharmaceutical companies, which are more likely to have substantial in-house expertise and resources for regulatory affairs, than it is for smaller companies and biotechs, some of which are deciding to conduct trials through a CRO in the United States instead. One adversely affected company, Oxxon Pharmaccines, intends to Þle an IND in the United States for its next vaccine candidate and then register it in only a very few EU countries, citing the perceived complexity, cost, and length of the EU’s application procedure. Pharmaceutical companies conducting research into rare diseases to develop orphan drugs are particularly concerned about the CTD because the burden of paperwork is threatening to deter the already overworked hospital specialists from conducting individual, noncommercial trials with tiny subsets of patients—trials that produce the initial results upon which companies can build. However, although the CTD may prompt some large pharmaceutical companies to move their trials out of the EU in the next two to three years, most of these companies will still need to conduct later-stage trials in the EU if they want to launch their drugs there and build acceptance of them. The EU may, therefore, lose some early-stage trials, and concern remains about how long the current loss of Phase I trials will continue, but it is unlikely to lose many later-stage ones. A related issue of particular signiÞcance to the noncommercial research community has been the CTD’s deÞnition of the sponsor of a trial, which implies a single individual or company. This concern led to a vigorous campaign in the United Kingdom in 2003 and early 2004, led by academics and charities, that resulted in a change of the U.K. legislation to allow for group sponsorship and, therefore, liabilities. Two or more individuals can now be jointly liable for a trial and different people can be sponsors of a particular domain of a trial, whether the CTA certiÞcate, pharmacovigilance, GCP, or trial management. Elsewhere in the EU, the deÞnition of sponsorship is a source of much confusion, making comparisons difÞcult at present. Some member states (e.g., Belgium, France) are tending loosely toward the United Kingdom’s multisponsor model; others (e.g., Spain), toward a single-sponsor model; and still others toward the idea of a single sponsor for the whole of the EU. Phase I Trials The main negative impact of the CTD on both the pharmaceutical industry and noncommercial research has been on Phase I trials. The United Kingdom, Belgium, and Holland used to have less strict requirements for Phase I trials than for later trials—only ethics committee approval, not approval by the CA, was required for the healthy volunteers involved. Thus, Phase I trials were easy and quick to conduct in these member states, making them attractive trial locales to multinational pharmaceutical companies and also encouraging an environment where academics carried out many such trials and often generated commercial interest in their results.

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The CTD makes no distinction between healthy volunteers and patients, deÞning a clinical trial as “any investigation in human subjects.” Consequently, all Phase I trials must now be approved by both an ethics committee and a CA, just as later trials must be. This requirement is likely to reduce the attractiveness of these member states to large pharmaceutical companies, hinder academics’ small-scale trials, and increase the workload of ethics committees. The United Kingdom has been leading the rest of the EU in its implementation of the CTD and its CA, the MHRA, is trying to minimize the loss of attractiveness of the United Kingdom as a location for Phase I trials. One action the MHRA has taken is to commit to assess Phase I study applications in a mean time of 14 days and a maximum of 21 days, rather than the 60-day time frame given by the CTD. One concern for European CROs and CMOs is that, if Phase I trials leave the EU, pharmaceutical companies may prefer to place whole development programs overseas. Another concern is that many national CAs will not be able to match the response time necessary for the efÞcient study scheduling, bed occupation, and staff utilization of a viable Phase I unit, especially given that many CAs already lack sufÞcient resources for inspections. The CTD is already having a substantial negative effect on many CROs involved in Phase I trials in member states where the CTD has been implemented. Others have, however, held on to their business and attribute their success to the following factors: • • • •

CROs’ ability to explain the CTD to current and potential clients and how to overcome any obstacles that it might present. CROs’ degree of globalization. CROs’ direct contacts at their national CAs; such relationships can halve trial approval times. Registering trials in member states, such as The Netherlands and Belgium, that have not fully implemented the CTD yet.

APPROXIMATION AND ITS CHANCES FOR SUCCESS To offset the main negative aspects of the CTD for the pharmaceutical industry—namely, the increased bureaucracy and cost and loss of some Phase I trials—the CTD needs to achieve, at least in part, its aim of approximation of registration procedures for clinical trials across the diverse and newly expanded EU. This section examines the current variations in procedures across Western Europe and the new accession countries of Eastern Europe and assesses the CTD’s potential to achieve a satisfactory degree of standardization. (In this article, “Western” Europe refers to the traditional members of the EU, before the new “accession” countries joined on May 1, 2004. The accession countries consist of the Czech Republic, Cyprus, Estonia, Hungary, Latvia, Lithuania, Malta, Poland, Slovakia, and Slovenia.)

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Help or Hindrance? Will the extent of the approximation and standardization of clinical research procedures brought about by the CTD bring sufÞcient beneÞts to trial sponsors to offset the extra paperwork and cost? The 60-day time limit for CAs and ethics committees to process trial applications across the EU should, if adhered to, help large pharmaceutical companies plan multicenter trials across several member states, although doubts have been raised as to whether all of the CAs and ethics committees have sufÞcient resources to meet that time limit. There is also still widespread confusion on how to implement the CTD across the EU, and each member state is taking a different approach. The Directorate General Enterprise at the EC has even admitted that the CTD has “failed to simplify and harmonize,” and there is much concern over member states’ failure, or unwillingness, to comply with the CTD. Questions are being asked about whether the EC will have sufÞcient strength or motivation to take legal action against noncompliant members. A more optimistic view from the pharmaceutical industry is that the CTD, even if it achieves a lesser degree of standardization than was originally sought, should make it slightly easier for pharmaceutical companies to conduct multicenter trials across Europe. By deÞnition, a directive will be implemented in different ways, and the CTD merely provides minimal standard criteria to be met. Given another 6–12 months, most of the member states should have implemented the CTD more fully, and greater standardization should be in evidence. Variations Across ‘‘Western’’ Europe By July 2004, only 11 of the 25 member states of Europe had implemented the CTD, despite the deadline of May 1, 2004. Other member states are implementing the CTD week by week, putting systems such as centralized ethics committees into place, but considerable variations remain. Among those that have not put their legislation through parliament yet are France and Germany, which are still using their old easier and quicker systems. One French company, therefore, recently decided not to conduct its trials in the United Kingdom but to keep them in France. The U.K. Clinical Contract Research Association wrote to the U.K. Ministry of Health to complain that France should be penalized for its delay, but no action in response is likely. France. France has transposed the CTD into a draft law that is still being Þnalized. The Assembl´ee Nationale and the Senate voted on August 9, 2004, to transpose the CTD into legislation. This vote approved the CTD in principle, but several areas require further clariÞcation through decrees. These areas, which cannot be implemented before the decrees are issued, include validating the proposed forms to be used for clinical trial applications and setting a minimum guarantee for trial sponsors’ insurance. France’s implementation delays have, according to a spokesperson at its CA, the Agence Franc¸aise de S´ecurit´e Sanitaire des Produits de Sant´e (AFSSAPS), been due to “lengthy parliamentary

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debates on issues such as trial ßight out of Europe, delays in trial approval, and the single-sponsor deÞnition, which could damage noncommercial trials.” France feels threatened by competition from the United Kingdom, with its quicker trial approval process, and so is trying to Þnd ways to reduce its delays. Germany. Similarly, the CTD still has no presence in German law. The German parliament recently failed to pass an amendment to its Medicines Law to transpose the CTD into law because of concerns that it would be anticompetitive and present too many obstacles to medical research. The federal and regional governments have been making conßicting demands on how to implement the CTD, while the inßuential local doctors’ boards are opposed to many of its principles. The Netherlands. The Netherlands still has several anomalies with regard to the CTD. Dossiers are still reviewed by individual ethics committees, not by a single centralized ethics committee as the CTD stipulates. The Netherlands has not yet made many changes to bring its regulations into line with the directive; for example, contrary to the CTD’s requirements, the role of CA is performed for most products by Commissie Mensgebonden Onderzoek (CCMO), the country’s central ethics committee, instead of by a separate body. United Kingdom. The United Kingdom is the member state that has most thoroughly and successfully implemented the CTD. The MHRA has worked very hard to achieve a smooth transition for all affected. In United Kingdom-based commercial trials, the main impact of the CTD has been on Phase I trials; there has been little impact on Phase II, III, and IV trials. Belgium. Belgium is an EU leader in pharmaceutical research. There, powerful groups are lobbying the government to ensure that Phase I trial units do not lose business as a consequence of the CTD, and the government has agreed to process Phase I trial applications in only 15 days. Italy. Italy was the Þrst EU member state to implement the CTD, doing so on January 1, 2004, although it has not followed the CTD to the letter—it still has no national drug agency. The role of CA is, therefore, usually delegated to the general manager of the hospital hosting the trial, and the ethics committee usually takes responsibility for allowing the trial to begin and checking the sponsor’s indemnity insurance. Spain. Spain is one of many member states trying to minimize bureaucratic delays as it implements the CTD. It requires only a single ethics committee opinion for multicenter studies, although the documentation must be sent to all relevant local ethics committees for consultation. Sponsors can also make submissions in parallel to the ethics committee and the Spanish Medicines Agency.

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Greece. Greece is being assisted by the EFGCP to make the many changes required before it can claim to have fully implemented the CTD. For example, the national CA and ethics committee currently consist of one single body, contrary to the CTD’s requirements. Accession Member States With the inclusion of the new accession member states, clinical trial review systems across the EU are even more varied. The EFGCP is helping several accession member states to set up appropriate legislation to implement the CTD; for example, it held a seminar in May 2004 in Bulgaria, which is working toward EU membership, to look at CAs, ethical committees, GCP, and GMP and to devise training and education programs to address any shortcomings identiÞed. Ethical review is the area requiring the most attention in the accession states, with legislation either substandard or entirely missing. Opinions differ on what sort of information committees should receive and how they should be constituted. Competence of members is often insufÞcient; funding is not available; the job lacks the prestige to provide the necessary motivation; inconsistent or conßicting opinions often emerge; waiting times are long; standard operating procedures are often lacking. Some member states, such as the Czech Republic, Slovakia, and Hungary, have had scientiÞcally rigorous ethics committees for many years, created out of the former Communist drug committees, but still lack the administrative infrastructure to support a centralized system that satisÞes the CTD’s requirements. Of the leading Central and Eastern European countries, all except Bulgaria are now EU members. The accession member states still host relatively few trials, almost all of which are pharma-sponsored. Once they have established CTD-compliant review systems, they are likely to attract more trials, given that trials are far less expensive to conduct there than in Western Europe; also, the accession member states feature more treatment-na¨õve patients. Poland in particular has recently become extremely attractive for clinical trials: its medical professionals are very well trained, and the large number of treatment-na¨õve patients yields extremely high recruitment rates. It is likely, therefore, to set CTD implementation as a priority. Issues for it to address include its numerous but badly organized ethics committees; its lack of a single national ethics committee; and its different application formats, insurance requirements, and timelines for decisions. OUTLOOK Many concerns were raised in 2003 and early 2004, mainly by the noncommercial research community, that the CTD would cause some trials to be cancelled, others to be delayed, and others to ßee the EU to be conducted in a region with less complex regulations. Here, we examine each of these issues, then conclude by outlining the challenges ahead for the CTD and assessing its overall impact on the global pharmaceutical industry.

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Trials Cancelled? The concern that the CTD would lead to the cancellation of some trials has resulted in the creation of a number of lobbying groups. One such group, Save European Research, has stated “this directive. . .will effectively end all clinical research except for those trials that are commercially inspired and drugsponsored” (www.saveeuropeanresearch.org). These lobbying groups have, however, now lost much of their momentum and voice. This downturn implies that they are at least partially satisÞed with the Þnal version of the CTD as it appeared in May 2004 and have moved on to issues such as broadening access to the data in Eudract and Eudravigilance. In member states that have implemented the CTD, the directive has prevented some small trials, most of which were using already approved products. Some of these trials have been stopped by hospital R&D departments, which now examine trial proposals more carefully than before because of the extra work of having to register them with the national CA. This change is, in part, a positive development, given that some of these trials were too small and not well enough administered to yield meaningful results. On the other hand, this trend could have a harmful effect on innovation, preventing some of the small trials that experiment with new combinations of existing drugs and can lead to unexpected breakthroughs. According to Dr. Barry Peters, principal investigator for HIV and genitourinary medicine (GUM) trials, Guy’s and St. Thomas’ Hospital, London, “The CTD will reduce the number of smaller, poorly funded studies, particularly those done by individual academics or researchers.” Noncommercial research centers do not have the resources for in-house regulatory staff to cope with the additional paperwork. Cancer Research UK claims, however, that anyone with a genuinely good idea should still be able to set up their trial, commercially or noncommercially, and not be prevented from doing so by the CTD. Overall, while trial cancellation has been a contentious issue, the hope remains that only those trials not well enough designed will be cancelled, mainly by hospital R&D departments unwilling to complete the registration paperwork for substandard trial proposals, and that any genuinely good trial idea will proceed. Trials Delayed? While no trials stopped on May 1, 2004, when the CTD ofÞcially took effect, a spokesperson from Cancer Research UK says that, in her experience, “the CTD legislation does appear to be slowing down the registration of new trials, due to the large amount of paperwork involved in setting up contracts between the participating centers, sponsors, and funders.” (No data are yet available on the extent of these delays.) While the extra cost involved is generally not proving to be prohibitive, some good trials could be delayed. Trial sponsors can be helped considerably by national advisory networks, which offer standard advice and guidance through the paperwork. The National Cancer Research Network in the United Kingdom is one such organization; it works with sponsors and ethics committees.

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Jane Barrett, a practicing physician within the pharmaceutical industry, had hoped that it might be quicker to start trials under the CTD than before because of the ethics committees’ strict time constraints, but says now that the goal is “looking unlikely as ethics committees have considerable extra paperwork and are allowed to stop their 60-day clock once to ask the sponsor for further details.” Furthermore, the United Kingdom’s efÞciency prior to the CTD will make it difÞcult for it to achieve much further improvement, while other countries, such as France and Spain, are struggling to Þnd ways to keep the CTD’s extra administrative burden from lengthening their approval times any further. Trial Flight? Many concerns were raised in 2003 and early 2004 that the CTD would discourage multinational sponsors from conducting their trials in the EU due to rising costs, the threat of delays, and the fact that the United Kingdom and Belgium no longer have simple registration procedures for Phase I trials. Earlier fears of the CTD—for example, that it could cause a global decline in clinical research by eliminating some trials completely—are now waning as researchers realize that, while the early months of the CTD will be accompanied by some disruption and confusion, the EU remains an extremely important market and attractive location for later-stage trials, although some Phase I trials may be lost. The CTD is unlikely to deter multinational companies from conducting later-stage trials in the EU in the long term—they need locally sourced data to present to CAs, and they need to give local physicians a chance to gain experience with and accept their drugs before launch. The outlook may not be so bright for the short term, however: evidence suggests that some non-EU-based companies are not coming to the EU to conduct trials because they fear the changes being rendered by the CTD and prefer to wait until things have settled down. Phase I trial units in particular have lost business across the EU, simply because of the novelty of the CTD. A Canadian CRO, Anapharm, which has a large Phase I unit, has tried to capitalize on this fear by publishing advertisements saying, “Concerned about the clinical trials directive? If so, consider Canada.” Steve Smith at Origin Pharmaceutical Services concludes from his discussions with the U.K. CRO community that the United States and Canada are attracting greater numbers of Phase I trials, some of which would previously have been conducted in Europe. Some EU-based CMOs claim that, thanks to good advertising campaigns designed to reassure clients, they have not lost business as a result of the CTD. Indeed, the CTD may have created opportunity for them: some non-EU-based pharmaceutical companies have understood and adapted to the CTD less well than others and have moved non-GMP-compliant products to the EU for trials and then have had to be rescued by CMOs. This trend implies that many EU governments and CMOs have not conveyed a sufÞciently strong and accurate message to would-be trial sponsors from outside the EU. However, although many non-EU based companies are wary of the CTD at the moment, in the long term these sponsors will become familiar with its processes.

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The accession member states may stand to gain the most from the CTD—with their inexpensive labor and as they become better regulated, they will attract more trials (another success factor will be the diminishment of the language barrier as individuals in these countries make the effort to learn English). Even if Western Europe loses a few trials outside the EU as a result of the CTD, the likely increase in trials in the accession states (sometimes trials displaced from Western Europe as a result of the lower costs possible in the accession states) suggests that the number of trials conducted in the EU as a whole will not decrease. Challenges Ahead for the CTD One challenge for the CTD moving forward is whether the EC will have the power to enforce the CTD, compelling those member states that have still not implemented it to do so and resolving the persisting anomalies in member states that claim that they have implemented it. The greater the EC’s power to enforce it, the more successful the CTD will be in an area of great beneÞt to pharmaceutical companies conducting multinational trials in the EU: the standardization of application procedures to CAs and ethics committees. Without that standardization, the CTD runs the risk of presenting more obstacles to pharmaceutical companies, in the form of increased paperwork and costs, than beneÞts, thereby reducing the EU’s overall attractiveness for trials. The EC has the power to enforce the CTD through legal action and may well do so once the current period of grace is considered to be over, although lengthy delays have occurred with other directives. This period of grace is likely to last for perhaps one and a half to two years after the implementation date of May 1, 2004. Standardization will be a gradual process, as member states work toward CTD compliance and the EC threatens or succeeds in imposing Þnes or taking persistently offending member states to the European Court of Justice. Another challenge for the CTD is how it will Þt in with ICH standards and the increasing push toward international harmonization of regulations to improve the quality control of clinical trials worldwide. The EC will need to cooperate further with the United States to work toward the mutual recognition of trial data, and also with other parts of the world such as Asia, where considerable effort is being invested to make less developed countries, with their inexpensive labor and large treatment-na¨õve populations, into viable locations for trials. The EU will need to balance helping them to raise their standards with managing the competition that these new countries could present. Quantitative data on the impact of the CTD is as yet hard to come by. But qualitatively speaking, the hope is that the Eudract and Eudravigilance databases will lead to more credible clinical trial data and CA access to complete trial records, rather than just the favorable ones that pharmaceutical companies choose to publish, thus providing a more accurate basis for drug recommendations. This change should silence the criticisms leveled at pharmaceutical companies for withholding trial data, but it may give companies less control over how their data are presented to regulatory authorities considering whether

OUTLOOK

133

to approve a new drug and include it in a national formulary. The impact of the CTD on trial costs is currently being assessed by the National Cancer Research Network (NCRN) and Cancer Research UK, which compiled baseline data on U.K. cancer trial costs before May 1, 2004, and are now measuring costs from mid-2004 to the end of 2005 to determine which costs have been changed by the CTD. While it is still too early to judge the full impact of the CTD, pessimism about it is much less prevalent than it was in 2003, and we anticipate even less negativity once the CTD’s systems are properly in place. The CTD could be of long-term economic beneÞt to the EU, further bolstering its reputation for high-quality research. Once the short-term disruption has been worked through and sponsors have invested the necessary resources in, for example, employing QPs, the EU may even attract more pharmaceutical companies because the standardization of application procedures should make it at least slightly easier to conduct trials in the EU than before. Some poorly funded trials are, however, likely to be prevented, which may stop some useful research from being conducted. In conclusion, we believe the CTD will not inßict signiÞcant damage on the EU’s commercial research effort in the long run because its impact on most pharma-sponsored studies will be minimal, with the slight beneÞts of the approximation of member states’ application procedures balancing, or perhaps just outweighing, the greater administrative burden and the extra approval required for Phase I trials. The CTD has mainly just put into law the GCP and GMP practices that most trial researchers adhered to in any case. It also affords patients proper legal protection for the Þrst time. Noncommercial trials are suffering more from the CTD because the guidelines do not as yet go far enough to protect them, and many of them are ill-prepared to cope with the extra administration and costs. The noncommercial trial community is continuing to lobby national governments and the EC for more favorable conditions and any of their future successes will clearly be of beneÞt to pharmaceutical companies, encouraging the early-phase innovation on which much of the industry depends.

Therapeutic Area Trends in R&D of the Top 20 Pharmaceutical Companies

SUMMARY This article illuminates trends in the overall therapeutic direction of leading pharmaceutical companies’ R&D since 1998. For companies seeking partners for their intellectual property, it provides practical, up-to-date insight into the therapeutic areas that each of the top 20 companies covers, particularly highlighting the areas these companies are entering and exiting. Also, this article provides an overall, industry-wide insight into therapeutic area trends for the top 20 pharmaceutical companies. Further more, it identiÞes weaknesses in the overall R&D effort by the pharmaceutical industry and reveals possible areas of unexploited opportunity. We contrast our current Þndings with those in a similar but broader analysis we conducted earlier. BUSINESS IMPLICATIONS •

•

•

The top 20 pharmaceutical companies are broadening their therapeutic horizons. The largest companies are now active in practically all of the halfdozen most popular therapeutic areas. Central nervous system (CNS) disorders has recently become the most popular therapeutic area in R&D and is currently the only one in which all the top 20 companies are active. Gastroenterology is undergoing the most marked renaissance. No less than six companies are newly active in gastroenterology—more than any other therapeutic area. The proportion of the top 20 companies both active and successful in progressing their R&D in the cancer, anti-infectives, and respiratory/allergy

Wiley Handbook of Current and Emerging Drug Therapies, Volumes 1–4 Copyright © 2007 Decision Resources, Inc. Published by John Wiley & Sons, Inc.

48

HOW WE CLASSIFIED PROJECTS BY THERAPY AREA

• •

•

49

Þelds is in decline. Companies are having particular difÞculty in making clinical headway in the respiratory/allergy area. The top 20 companies are particularly attracted to the therapeutic areas of rheumatology, dermatology, and ophthalmology. The largest companies among the top 20 continue to be most active in pursuing the least popular therapeutic areas—vaccines, transplantation, and wound healing. Because the strength of competition in different therapeutic areas is less transparent in R&D than it is in the market, companies are not necessarily succeeding in identifying less obvious product opportunities; we assess there to be considerable scope for seeking out these opportunities.

INTRODUCTION Leading pharmaceutical companies do not follow any particular rule for determining the range of their R&D efforts, and no obvious correlation exists between the size of a company and the breadth of its therapeutic portfolio. We do observe, however, that the more obvious dearth of new products in the last Þve years has led the top 20 pharmaceutical companies to become much more prepared to broaden their therapeutic horizons. The largest companies are now active in practically all of the half-dozen most popular therapeutic areas. In this article, we discuss trends in the overall therapeutic direction of leading pharmaceutical companies’ R&D since 1998. First, our aim is to provide practical, up-to-date insight into the therapeutic areas that each of the top 20 companies covers, particularly highlighting the areas these companies are entering and exiting. Second, we provide an overall, industry-wide overview of therapeutic area trends for the top 20 pharmaceutical companies, and third, we identify weaknesses in the overall R&D effort by the pharmaceutical industry and reveal possible areas of unexploited opportunity. HOW WE CLASSIFIED PROJECTS BY THERAPY AREA Pharmaceutical companies apply varying therapeutic area terminology to their R&D pipelines. Therefore, we devised a uniform therapeutic classiÞcation system that takes into account these variations to enable this analysis. (The classiÞcation system we use is shown in Table 1.) The aim of the therapeutic classiÞcation system we devised was to reßect in a pragmatic manner terminology that the industry uses. Therefore, wherever necessary, we have reinterpreted the terminology that companies themselves employ, according to the uniform classiÞcation system we devised, to enable this analysis. As shown in Table 1, the terminology that results uses anatomical (e.g., gastroenterology, ophthalmology) and therapeutic (e.g., anesthesia, cancer) categories. In this article, we include only projects that are in development for therapeutic purposes and are potential prescription products; thus, we exclude diagnostics

50

THERAPEUTIC AREA TRENDS IN R&D OF THE TOP 20 PHARMACEUTICAL COMPANIES

TABLE 1. Classification System for Therapeutic Areas Therapeutic Area CNS Cardiovascular Metabolic

Cancer Anti-infectives

Respiratory/allergy Rheumatology Gastroenterology Dermatology Ophthalmology Vaccines

Transplantation Wound healing Anesthesia

Comments on Classification Includes pain (of either central or peripheral origin) but not when associated with rheumatological disorders. Includes stroke, renal disorders associated with the cardiovascular system, blood and clotting disorders. Includes diabetes, obesity, urological disorders, and hormones; male erectile dysfunction, baldness, and disorders of the prostate; gynecological disorders, osteoporosis, and contraception. Excludes infections and cancer. Excludes antiemesis associated with cancer chemotherapy. Includes antibacterials, antivirals, antifungals, and other anti-infectives; excludes vaccines and ophthalmological infections. Includes allergies associated with respiratory disorders; excludes allergy associated with dermatological disorders. Includes pain associated with rheumatological disorders. Includes all antiemesis compounds, including those specifically in use with cancer chemotherapy. Includes allergies associated with dermatological disorders. Includes ophthalmological infections. All vaccines. (Although a vaccine may fall under another therapeutic area (e.g., a smallpox vaccine could also be considered an anti-infective), to avoid double counting, we classify all vaccines in this area.) Transplantation of any organ. Wounds of all types. Including inhalational, intravenous, and local.

(e.g., contrast media) and over-the-counter products. We have restricted inclusion to projects that have reached Phase II clinical trials but have not yet been granted registration or reached the immediate postlaunch phase. A company’s declaration of involvement in a therapeutic area is insufÞcient in itself for that area to be included; it must also have made progress with at least one project beyond the Phase I clinical trial stage. The Phase II cut-off reßects a company’s true commitment to a particular therapeutic area as well as some tangible progress in that area. That progress, however, may have been made outside the company, such that, for example, inlicensing of a rheumatological project would still qualify a company as active in that therapeutic area. To investigate therapeutic trends in more detail, we also apply a more stringent cut-off, whereby the bar is raised to projects at Phase III and beyond. When an originating company is still active in the rollout of a product in major markets, it can still be considered as partially engaged in the R&D phase—it is now the rule rather than the exception for major additional indications to still be in development for a new product currently in the launch phase. Moreover, companies are increasingly designating additional indications as new Phase I, II, or III projects rather than as postlaunch Phase IV projects. For example, in December 2003, GlaxoSmithKline (GSK) stated that of 147 projects in clinical

THERAPEUTIC AREA R&D ACTIVITY OF THE TOP 20 COMPANIES

51

trials, 45 (31%) were product license extensions. GSK’s product license extension projects are in various clinical phases back to Phase I, but these projects are skewed toward the later clinical phases—product license extensions—because they are based on already marketed products and are not subject to trials intended to support a new chemical entity. We Þnd the proportion of GSK’s projects that are product license extensions, and the skewing to later phases, not atypical for major pharmaceutical companies. THE TOP 20 COMPANIES Table 2 shows the therapeutic areas in which the top 20 companies in 2003 were currently active, namely those with at least one project in development that reached Phase II clinical trials or beyond. In 1999, we looked at a larger group—the top 50 companies. This group included companies outside the current top 20 such as Boehringer Ingelheim, Schering AG, and Amgen, which have risen in the rankings to be included in the 2003 top 20. Largely as a result of mergers, the composition of the top 20 has changed considerably since our 1999 study, which was based on data from the end of 1998. When reporting on trends, we ensure that we are comparing like with like by including all constituent companies of merged companies that were separate entities in 1998. For example, for PÞzer we included the following constituent companies: Pharmacia & Upjohn, Warner-Lambert, and Monsanto (Searle). Those separate entities, however, had to be big enough to rank in the top 50 for the latest rankings available in that year. Any smaller companies outside the top 50 at that time are not included. For example, Alza, which was acquired by Johnson & Johnson in 2001, is not included. THERAPEUTIC AREA R&D ACTIVITY OF THE TOP 20 COMPANIES On average, the top 20 companies are involved in just under 8 (7.8) therapeutic areas. The largest company, PÞzer, is involved in 8. But one company, Wyeth, is involved in no less than 12 areas, with two others, Novartis and Roche, being involved in 11, and one company, Abbott, in 10. As Table 2 shows, the companies active in an above-average number of therapeutic areas are all in the middle third of the top 20 rankings. And although there appears to be some correlation between the size of a company and the number of therapeutic areas in which it is active, this correlation is not particularly strong. For example, the third-largest company, Merck & Co., is involved in only seven therapeutic areas, whereas Schering-Plough, ranked 17th, is involved in eight. We conclude that there is no general consensus among the leading pharmaceutical companies on breadth of R&D portfolio; nor is there a general rule of thumb linking size of company to breadth of therapeutic portfolio. Levels of Activity by Therapeutic Area In 2003, every top 20 company was active in the CNS area. According to our earlier surveys, cardiovascular was the most popular area, but CNS was never

a Company rankings as of 2003.

• • 19

• • • • • • • • • • • • • • • • • • 18

• 15

• 14

• 14

16

• •

• • • • •

•

• • • • • • •

• •

•

13

9

Wound Healing

•

• • •

• •

•

Transplantation

•

• • • •

•

• • • •

•

Vaccines

• • • •

•

• •

• • •

Ophthalmology

• • • • • • • • • • •

•

Dermatology

• • • • • • • • • •

• • • • • • •

Gastroenterology

• •

Rheumatology

• •

Total

• • • • • • • • • • • • • • • • •

Anesthesia

• • • • • • • • • • • • • • • • • • • • 20

Respiratory/Allergy

Pfizer GlaxoSmithKline Merck & Co. Johnson & Johnson Aventis AstraZeneca Novartis Bristol-Myers Squibb Wyeth Eli Lilly Abbott Roche ´ Sanofi-Synthelabo Boehringer Ingelheim Amgen Takeda Schering-Plough Schering AG Bayer Sankyo

Anti-infectives

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 Total

Cancer

Company

Metabolic

Ranka

Cardiovascular

Therapeutic Areas

CNS

52

TABLE 2. Therapeutic Areas in Which the Top 20 Pharmaceutical Companies Are Active—Phase II or Beyond

0

8 9 7 9 7 7 11 8 12 5 10 11 8 9 7 4 8 4 4 7 155

• • •

• •

•

• • •

• 7

4

• • •

• •

4

2

THERAPEUTIC AREA R&D ACTIVITY OF THE TOP 20 COMPANIES

53

too far behind. CNS is undoubtedly a therapeutic area that will continue to hold a leading activity level among the top 20 companies. The gap between CNS and cardiovascular is very small. Cardiovascular R&D is still being actively pursued by 19 out of the 20 companies—it is only the recent withdrawal of Schering AG from the cardiovascular area that has allowed CNS to take the lead. (Table 3 shows the top 20 companies that have exited the leading therapeutic areas since 1998.) In recent years, the metabolic area has gained a level of importance almost equal to that of the cardiovascular area: 18 of the top 20 companies have projects in it. Oncology follows the cardiovascular and metabolic areas with 16 companies active in it. Its ranking in fourth place is a little surprising, because it is the therapeutic area accounting for by far the greatest number of R&D projects in development, taking into account developers of any size. Thus, we see that the level of popularity in R&D of therapeutic areas within the top 20 pharmaceutical companies does not necessarily apply to the activity of smaller companies (including start-ups). Moreover, if we compare like with like, current oncology activity shows a marked reduction from 1998, when the equivalent of all 20 companies were active—a rather sudden decline in the popularity of oncology. The principal explanation for this trend is that a few of the top companies do not qualify for inclusion this time because they have not yet reached Phase II with any of their cancer projects. This is the case, for example, for Sankyo. This company considers cancer to be what it terms a challenge therapeutic area, in which it has yet to make clinical progress; the most advanced of its cancer projects is still at the preclinical phase. The next most popular therapeutic area is the anti-infectives, in which we Þnd 15 companies active. As with oncology, interest in anti-infectives declined considerably among the top 20 since 1998, when the equivalent of 18 companies were active. This decline in interest in anti-infectives has been ongoing for more than a decade. Although the antiviral area is still popular, leading companies have particularly lost interest in antibiotics. Fourteen companies are involved in the respiratory/allergy area, a considerable decline in activity from the 18 equivalent companies involved with this therapeutic area in 1998. This decline may reßect the relatively modest opportunity for improvement in this therapeutic area and consequent lesser interest in developing new approaches to treating respiratory diseases. Rheumatology is a second therapeutic area with 14 companies active. The level of top 20 company interest is similar to that in our report from 1999. One notable difference in this therapeutic area is that antibody products now contribute signiÞcantly. The top 20 companies have increased their interest in the gastroenterology therapeutic area. In 1998, just 8 companies had active projects in this therapeutic area; this number has now grown to 13 companies—the greatest increase observed among the therapeutic areas tracked. During the early 1990s, gastroenterology

54 TABLE 3. Companies That Have Exited Therapeutic Areas Since 1998

E

E E E

E

E

E E E E E E E E

E E

3

E 4

E

E

E E 1

E 2

E 2

1

1

5

1

1

1

3

Total

Anesthesia

Wound Healing

Transplantation

Ophthalmology

Dermatology

Gastroenterology

E

Rheumatology

Anti-infectives

E

Respiratory/Allergy

Cancer

GlaxoSmithKline Merck & Co. Aventis AstraZeneca Novartis Bristol-Myers Squibb Abbott Roche ´ Sanofi-Synthelabo Boehringer Ingelheim Amgen Takeda Schering-Plough Schering AG Bayer Total

Metabolic

Company

Cardiovascular

Therapeutic Areasa

2 3 2 2 1 1 1 1 1 1 1 4 2 2 2 25

a E = Exit from R&D in a therapeutic area. This table indicates those companies not qualifying for therapeutic areas in 2004 in which they qualified as active in

1998. Excluded from the table are companies that have exited no therapeutic areas, and therapeutic areas that no company has exited.

THERAPEUTIC AREA R&D ACTIVITY OF THE TOP 20 COMPANIES

55

became unpopular with companies because of a widely perceived but now clearly erroneous perception that there was little room for improvement after the establishment of the successive blockbuster antiulcer products Tagamet (cimetidine), Zantac (ranitidine), and Prilosec/Losec (omeprazole). Thereafter, many companies withdrew from R&D in this area, but now 13 companies are involved. The recent revival in company interest stems particularly from gastroenterological developments in monoclonal antibodies. (For more information on this topic, see the section “The Trend Toward Greater Therapeutic Diversity.”) Monoclonal antibodies have played an important role in raising company interest in inßammatory bowel disease, which was formerly considered a rather esoteric area of gastroenterology with only modest commercial potential. 5-HT antagonists, such as alosetron (GlaxoSmithKline’s Lotronex) and tegaserod (Novartis’s Zelnorm), have helped to resuscitate company interest in irritable bowel syndrome, another badly served gastrointestinal indication, although these products have yet to make much of an impact on the market. The next most popular therapeutic area is dermatology, which, like rheumatology, has also beneÞted from antibody developments. Beyond the leading therapeutic areas we have listed so far, company interest among the top 20 in the remaining Þve therapeutic areas is much more limited. Seven companies are active in ophthalmology, an increase of two over 1998. Four are active in vaccines and in transplantation, and two in wound healing. However, numbers for these therapeutic areas are too small to consider the trends to be meaningful. The equivalent of four companies that were active in anesthesia in 1998 have all withdrawn from this area. One of these companies was AstraZeneca—and given that both of its constituent companies, Astra and Zeneca, were active in anesthesia before they merged, this withdrawal is even more of a loss than is apparent from our Þgures. This decline in interest in anesthesia is universal: practically no R&D projects are ongoing in this Þeld from companies of any type. Patterns of Therapeutic Area Popularity As we have noted, participation for the ten most popular therapeutic areas does not vary obviously with company size. But thereafter, a pattern is evident. As Table 2 shows, those companies that are active in the three least popular areas—vaccines, transplantation, and wound healing (we exclude the now dormant area of anesthetics)—all appear in the Þrst dozen of the top 20. Our earlier Þndings were similar. One might expect that such therapeutic areas would have less commercial attractiveness for a big pharma company and would be more suited to the lowerranking companies in the top 20, for which the commercial impact might still be signiÞcant. However, the lower-ranked companies are not attracted to these therapeutic areas. This fact indicates that smaller companies are more limited in the number of therapeutic areas in which they can afford to be active than are larger companies. Table 2 shows that the 16th, 18th, and 19th ranked companies

56

THERAPEUTIC AREA TRENDS IN R&D OF THE TOP 20 PHARMACEUTICAL COMPANIES

in the top 20 (Takeda, Schering AG, and Bayer) are each active in only four therapeutic areas—the smallest number among the top 20. Companies of this size may wish to be more adventurous in the areas they support. But if they were to pursue any of the more esoteric therapeutic areas, they would probably need to opt out of some of the most popular therapeutic areas, which contain the majority of the largest markets. APPLYING A MORE STRINGENT CUT-OFF: LIMITING INCLUSION TO PHASE III AND BEYOND Although the focus of this article is on tracking the presence of companies in therapeutic areas from Phase II onwards, we have also—for the Þrst time—applied a more stringent cut-off: activity in Phase III and beyond. This information serves to differentiate companies in each therapeutic area that have made more progress in R&D from those that have only a foothold. Table 4 shows therapeutic areas in which companies have projects that have advanced at least as far as Phase III. With this more stringent restriction, on average, the top 20 companies are each involved in slightly more than six (6.3) therapeutic areas. We Þnd that the Þrst six companies—those with some of the largest R&D portfolios—are largely unaffected by exclusion of Phase II projects. But as Table 4 shows, the two most affected companies, Roche and Boehringer Ingelheim, each loses four therapeutic areas when Phase II projects are excluded. Boehringer Ingelheim is a company that has grown rapidly in recent years, and it will take time for it to develop projects beyond Phase II. But for the remaining companies, the data serve to show how limited their therapeutic coverage is when only advanced projects are taken into account. Impact by Therapeutic Area As Table 4 shows, for three therapeutic areas—cancer, anti-infectives, and vaccines—the same number of companies were considered active when classiÞed as having projects at Phase III and beyond as for projects at Phase II and beyond. The most impressive aspect of this Þnding is the newfound strength of vaccine R&D. Our Þndings show that although this area is making impressive progress, it still has a long way to go in attracting the broad interest of major pharmaceutical companies. Rheumatology maintains a high level of company commitment when we consider projects that have reached Phase III and beyond—11 of the 14 companies are still counted. This fact shows that the renaissance in rheumatology—a long time dormant therapeutic area for new product development—is well under way. Several of the other therapeutic areas do not maintain their level of representation among the top 20 companies when projects at Phase II are excluded. Only 6 of the 13 gastroenterology companies remain—less than half the number as when Phase II is included. This outcome is likely due to the fact that renewed top 20 company interest in gastroenterology is fairly recent—more recent than for rheumatology.

TABLE 4. Therapeutic Areas in Which Top 20 Pharmaceutical Companies Are Active—Phase III or Beyond

57

a Company rankings as of 2003.

• • • • • • • • • 15

• • • 18

• • • • • • • • • • • • 17

• • • • 16

•

• • • • • • •

•

•

• •

• 15

• 8

• 11

•

•

•

•

• •

• •

•

• •

•

•

6

7

• •

•

• • •

5

•

4

2

1

0

Total

•

Anesthesia

• • •

•

Wound Healing

• • • • • • • • • • •

•

Transplantation

• • • • • • • • • •

•

Vaccines

• •

Ophthalmology

• •

Rheumatology

• •

Dermatology

• • • • •

Gastroenterology

• • • • • • • • • • • • • • •

Respiratory/Allergy

• • • • • •

Anti-infectives

Pfizer GlaxoSmithKline Merck & Co. Johnson & Johnson Aventis AstraZeneca Novartis Bristol-Myers Squibb Wyeth Eli Lilly Abbott Roche ´ Sanofi-Synthelabo Boehringer Ingelheim Amgen Takeda Schering-Plough Schering AG Bayer Sankyo

Cancer

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 Total

Metabolic

Company

Cardiovascular

Ranka

CNS

Therapeutic Areas

8 9 5 9 7 6 8 5 9 5 9 7 6 5 5 3 7 2 4 6 125

58

THERAPEUTIC AREA TRENDS IN R&D OF THE TOP 20 PHARMACEUTICAL COMPANIES

In the respiratory/allergy area, the number drops to 8 companies from 14. This decline likely reßects the difÞculty in progressing products through clinical development that prove to have qualities superior to existing products. The CNS area had a reduction of 5 companies—down to 15 from 20—when we excluded projects that had not progressed at least as far as Phase III. This reßects the fact that CNS is a therapeutic area with particularly high attrition rates during clinical development. THERAPEUTIC AREA TRENDS OF THE TOP 20 COMPANY COHORT We now review therapeutic area trends of the top 20 company cohort in toto. Companies Entering Therapeutic Areas Since 1998 Table 5 shows the therapeutic areas in which the top 20 companies are newly active since 1998. We revert here to our base criterion of companies with projects at least in Phase II clinical trials. We include new areas entered through acquisition (e.g., PÞzer has entered ophthalmology as a result of its acquisition of Pharmacia). We determined that the top 20 companies entered 27 new areas (an average of 1.35 per company). At one extreme, two companies (Bristol-Myers Squibb and Boehringer Ingelheim) entered three areas, whereas at the other extreme, Þve companies entered no new therapeutic areas. Sankyo is another company that for some time has been focusing on the cancer area but whose efforts have yet to result in at least a Phase II project. Companies Exiting Therapeutic Areas Since 1998 As shown in Table 3, Takeda exited four therapeutic areas since 1998—the most exits among the top 20. Merck & Co. exited three areas, and GlaxoSmithKline, Aventis, AstraZeneca, Schering AG, and Bayer exited two each. Companies exiting therapeutic areas are not necessarily doing so voluntarily—an exit may simply indicate that companies do not currently have any qualifying project that has reached Phase II. The most commonly exited therapeutic areas are dermatology and antiinfectives, with Þve and four companies exiting, respectively. Surprisingly, three companies have exited cancer. Also, the only three active in anesthesia have exited this area (four in reality, because both Astra and Zeneca have exited this Þeld). Two companies also exited the respiratory/allergy and metabolic areas. We can see from these analyses of exiting and entry of therapeutic areas that dermatology is the area most in a state of ßux. Therapeutic Areas in Which Companies Did Not Succeed In the second half of the 1990s, GlaxoSmithKline attempted to establish itself in rheumatology, but currently, its most advanced projects have advanced no further than Phase I. Schering AG attempted to enter the cardiovascular area, but withdrew in 2004. Bayer tried and failed in the late 1990s to enter the respiratory area.

TABLE 5. Therapeutic Areas in Which Companies Are Newly Active—Phase II or Beyond

• • • • • • • • • • • • • • • • • N • N 20

• • • • • • • • • • • • • • • • • • • 19

• • • • • • • • • • • • • • • • • • 18

• 15

• 14

• 14

16

a Company rankings as of 2003. b New areas (N) include those entered as a result of acquisitions (e.g., Pfizer of Pharmacia).

Anesthesia

•

• • •

Wound Healing

•

• • • N

•

Transplantation

• • • •

•

• N

Vaccines

• • • N • • • • • N •

•

Ophthalmology

• • • • • • • • • •

• • • • • • •

Dermatology

• •

Rheumatology

N •

Gastroenterology

Respiratory/Allergy

Anti-infectives

Pfizer GlaxoSmithKline Merck & Co. Johnson & Johnson Aventis AstraZeneca Novartis Bristol-Myers Squibb Wyeth Eli Lilly Abbott Roche ´ Sanofi-Synthelabo Boehringer Ingelheim Amgen Takeda Schering-Plough Schering AG Bayer Sankyo

Metabolic

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 Total

Cardiovascular

Company

Cancer

59

Ranka

CNS

Therapeutic Areasb

N • N •

N • N •

• •

N •

• N • N N

•

•

• •

• • N N •

N N

N N

•

13

9

• • N

•

• N N

• 7

4

• N • •

N

4

2

0

60

THERAPEUTIC AREA TRENDS IN R&D OF THE TOP 20 PHARMACEUTICAL COMPANIES

Therapeutic Areas That Companies Exited and Later Reentered After producing the global best-selling antiulcer product Zantac (ranitidine), Glaxo subsequently made a conscious decision to exit gastroenterology, maintaining just a limited gastroenterology research program. As GlaxoSmithKline, it decided to reenter this market in the late 1990s. Merck & Co. was very much involved in gastroenterology, notably with its highly successful antiulcer product Pepcid/Pepdul (famotidine) licensed from Yamanouchi and subsequently, until rights returned to its originator Astra, with Prilosec (omeprazole). However, by 1998, the company did not have a sufÞcient presence in gastroenterology R&D to qualify in this therapeutic area. But Merck & Co. is represented once again, with antiemetic compounds at Phase II. Its novel antiemetic, Emend (apretinant), was launched in 2003 in its Þrst market, for chemotherapy-induced nausea and vomiting. This product was originally in development for CNS indications; emesis was only a subsidiary indication, but the product did not succeed in the former indication, and gastroenterology became its lead area. Boehringer Ingelheim has a heritage in gastroenterology with products such as the antispasmodic Buscopan (hyoscine butylbromide) but had exited the area by the late 1990s. However, the company is active again in a different area of gastroenterology: liver Þbrosis. This development stems from the company’s involvement with Genentech’s Immunikin (interferon gamma 1b), which is at Phase II. Heritage Areas Abandoned Companies are generally reluctant to abandon therapeutic areas in which they have many years of experience. However, on occasion companies abandon these heritage areas for a variety of reasons. The most notable recent example of a heritage area being exited was the decision of Schering AG in June 2004 to abandon dermatology, an area the company had a presence in for more than forty years. Schering AG decided to focus its resources on other areas it considered more promising. Another example is AstraZeneca’s withdrawal from anesthesia. The constituent companies, Astra and Zeneca, had both been active in the Þeld for approximately sixty years. Astra had developed the pioneering product lidocaine (Xylocaine). Presumably, AstraZeneca regarded anesthesia as too small a Þeld to warrant inclusion in the R&D pipeline of the merged company. Companies may consciously exit areas if they no longer see the commercial sense in maintaining a presence in it. Or the R&D pipeline in that area may simply, despite best efforts, dry up. Finally, even when companies formally drop heritage areas from their R&D portfolios, they have a tendency to later reinstate them. For GlaxoSmithKline and previous Glaxo companies, dermatology and gastroenterology have been two such areas. Conclusions for Partnering Were a start-up company considering development of a novel anesthetic agent, it would be faced with a bleak situation in that that there are no longer any top

SMALLER COMPANIES

61

20 pharmaceutical companies active in developing anesthetic agents. In contrast, in 1998 Abbott, Astra, and Zeneca were all involved in this therapeutic area. Is the anesthesia area so well served with products that it can afford to do without the R&D effort of any top 20 pharmaceutical companies? While this is an extreme example, it does serve to show that the workings of the market may be imperfect. The industry may, through an incomplete understanding of competitor portfolios, be unaware that such development vacuums are being created. These misunderstandings are capable—rightly or wrongly—of rendering a therapeutic area inactive. This particular case may present opportunities for a medium-sized pharmaceutical company with the capability to develop and market products to Þll such a market vacuum. While opportunities in the anesthesia market may not be huge, they could well be sufÞciently large to be of interest to a company of this size. As mentioned, anesthesia is an extreme example. But we suspect that the market is operating inefÞciently, with some therapeutic areas being oversubscribed in terms of company participation, while others are being neglected. This fact means that companies short of new product ideas should consider neglected areas, which may be particularly open to partnering opportunities. The Demise of Core Therapeutic Area Strategy The factors discussed previously ultimately rendered obsolete the fashionable “core” therapeutic area philosophy that, when it was at its peak in the mid1990s, was espoused by two-thirds of the leading companies. The trend then was to focus down to fewer of these core therapeutic areas. This trend was already abating by the late 1990s, and the concept of limited core therapeutic areas has now gone so completely out of fashion that the word core is now seldom used by companies in describing their strategies. Consequently, supposed economies from focusing down have largely proved not to be possible—one factor, no doubt, in pushing up the cost of R&D. SMALLER COMPANIES The growing disparity in size between the largest pharmaceutical companies and even those in the lower reaches of the top 20, let alone beyond, has rendered the concept of selectivity, with focus on core therapeutic areas, largely obsolete as far as the major companies are concerned. For as the size of major pharmaceutical companies increases, in particular through mergers, they increasingly cannot afford to neglect any therapeutic area that might contain a major product. However, smaller companies cannot possibly afford to cover more than a few therapeutic areas—so now it is these companies that more readily declare their commitment to a limited number of therapeutic areas. Therefore, although the main focus of this article is the top 20 companies, it is worthwhile to review smaller companies that have recently implemented changes in their therapeutic area focus. Table 6 shows the therapeutic area interests of several smaller companies.

62

THERAPEUTIC AREA TRENDS IN R&D OF THE TOP 20 PHARMACEUTICAL COMPANIES

TABLE 6. Therapeutic Area Strategies of Select Smaller Companies Company

Therapeutic Area Strategya

Pharmaceutical company Akzo/Nobel (Organon) Entered vaccines and biopharmaceuticals in 2003. Almirall Now focuses on therapies for asthma, bronchitis, allergies, atopic dermatitis, and gastrointestinal conditions. Kowa Aims to concentrate on lifestyle diseases (including diabetes), allergies, and immune disorders. Leo Aims to become world leader in dermatology by 2010. Leo believes that dermatology is often too small an area for large pharma companies but that it suits Leo well. The company has been aiming to enter the cancer area, but its projects so far have advanced only as far as Phase I. Meiji Seika Though continuing to concentrate resources on expanding its mainstay anti-infectives business, it is looking to build up activities in the central nervous system (CNS) area over the next few years. Also, vaccines are seen as another growth opportunity. The company additionally has compounds in the respiratory area. Mochida The in-house R&D effort is now concentrated in CNS, allergy, metabolic, circulatory, and biotechnology areas. Novo Nordisk Stated an interest in the cancer area in 2003. Novo Nordisk is also trying to apply its expertise in protein research outside its main metabolic area. Orion Stated in 2004 that it intended to diversify its product pipeline, which at that time focused on the CNS, cardiology, critical care, hormone therapy, and urology areas. Pliva The company plans to develop proprietary molecules in six therapeutic areas: antifungals, inflammatory bowel disease, oncology, wound healing, anti-infectives, and asthma. It also mentions already being involved in diabetes. Shionogi Seeks to diversify from its main antibiotic franchise into arthritis, antiobesity, and spinocerebellar ataxia. Solvay Stated in 2004 that its current focus is on four key therapeutic areas: gastroenterology, gynecology/andrology, cardiology, and mental health. Start-up company Biogen Idec Formed through a merger in 2003, the new company considers itself strong in oncology, immunology, multiple sclerosis, and Crohn’s disease. Shire Shire is exiting its cancer and anti-infectives research programs. The company is also in the process of divesting its vaccines business. The company will instead focus on the CNS, gastrointestinal, and renal disease areas. Vernalis Concentrates, following its recent merger with British Biotech, on CNS disorders and oncology, though it also has late-stage programs in sexual dysfunction, thrombotic disorders, and community-acquired pneumonia. a We give summaries of therapeutic area policies, including changes, that have been announced since the

beginning of 2003 by select smaller companies. Because it was not our intention to carry out a comparative analysis of these companies’ pipelines, we have not attempted to restate the therapeutic areas in which they are active in terms of our classification system.

PARTNERING FOR MULTIPLE INDICATIONS

63

THE TREND TOWARD GREATER THERAPEUTIC DIVERSITY The more obvious dearth of new products now than in 1998 has meant that most companies have become much more prepared to broaden their therapeutic horizons to improve their chances of success in developing new products. Consequently, the biggest companies are active in practically all of the most popular therapeutic areas. Companies now appear to be more prepared to develop compounds in unusual new indications. GlaxoSmithKline’s ReQuip (ropinirole) launched initially for Parkinson’s disease and was also investigated for other CNS disorders but is now being developed for restless legs syndrome, a disorder for which there is currently no approved product. The dearth of new products has also forced companies to more fully exploit products that they do succeed in developing. A current example of broad exploitation of a product is Johnson & Johnson’s Topamax (topiramate). This antiepileptic (AED) agent has multiple mechanisms of action. It has been developed for epilepsy and is in development for migraine, post-traumatic stress disorder, postherpetic neuralgia, essential tremor, and diabetes. It has also been investigated but found inactive in pain associated with diabetic neuropathy and in bipolar disorder. A number of different classes of product with application across different antiinßammatory areas have helped this trend. Monoclonal antibodies, which began to reach the market in 1995, have become the most important of these products. Monoclonals have already been launched in Þve different therapeutic areas, and there are few areas in which their application has not been investigated. Table 7 shows select antibody products that are being developed in two or more therapeutic areas. Many more, which do not appear in Table 7, have been or are being developed in just one therapeutic area. This fact may be a function of inherently more speciÞc properties of some monoclonal antibodies or may reßect the considerable potential that exists within a single area. Cancer is the best example. If the scope exists to develop treatments for a wide variety of cancer indications, why go outside the cancer area? It might also be that, for commercial or other reasons, companies prefer to develop different antibodies for different therapeutic markets. But there is not very much evidence for this so far with antibodies. We believe that, with the growing costs of getting a new chemical entity to market, the duplication in costs involved is diminishing the use of this strategy. We suspect that there is still a considerable amount of therapeutic underexploitation of antibodies, which reßects some companies’ reluctance to break through into new areas. PARTNERING FOR MULTIPLE INDICATIONS Can a company wishing to Þnd a partner for a multiple-indication project ensure that its product will be fully exploited commercially? It is essential to look at the track records of candidate partners.

64 TABLE 7. Development of Select Antibody Products Across Therapeutic Areas

Siplizumab Ntalizumab CDP-571 Declizumabb Eculizumab

X

L

Medimmune — Antegren Humicade Zenapax

Biogen Idec/Elan Celltech Protein Design Labs/Roche Alexion

L D L

X L D

L D D D

D

X D

D D

D

D X

D X X D

L L D

D X D

Protein Design Labs

Transplantation

D

X

X D

L

— Fontolizumab

D

D

D D

Dermatology

Gastroenterology

Rheumatology

Johnson & Johnson Genentech/Xoma/Roche Abbott Johnson & Johnson Roche

Respiratory/Allergy

Remicade Raptiva Humira Reopro Rituxan/Mab Thera

Anti-infectives

Infliximab Efalizumab Adalimumab Abciximab Rituximab

Cancer

Main Company (Marketing or Developing)

Metabolic

Product Name

Cardiovascular

Generic Name

CNS

Therapeutic Areasa

D

D

— a L, launched in therapeutic area; D, in development for therapeutic area; X, investigated in therapeutic area, but development discontinued or no longer progressing. b Daclizumab is also being developed in ophthalmological indications.

AREAS OF MISMATCH IN PRODUCT SUPPLY AND DEMAND

65

For example, Table 7 includes three major pharmaceutical companies, Johnson & Johnson, Abbott, and Roche, that already have a track record in broad exploitation of their monoclonal products. The track records of these companies thus provide, for those seeking partners for multiarea projects, evidence of their ability to succeed in exploiting antibody products broadly. Imagine a start-up company that has so far been developing an anti-inßammatory (whether an antibody or not) in three different therapeutic areas: transplantation rejection, rheumatology, and gastroenterology. It is likely to want to choose a partner that will be committed to developing the compound in all three of these areas. Table 2 shows that transplantation is the most critical area—because only four top 20 companies are active in this area. All four of them are, as it happens, active in one of the other two required therapeutic areas, rheumatology, and three are active in the other, gastroenterology. But that is too limited a list of candidate partners. The company may well have to cast its net much wider to include candidates outside the top 20. But not many of these candidates will qualify for all three areas; they will be smaller companies active in fewer therapeutic areas and therefore less likely than top 20 companies to cover precisely the three therapeutic areas required. This is a fairly extreme example where no ideal match may exist. But only by going through an exercise such as this can a company properly gauge its chances of gaining a satisfactory partner that will fully exploit the product and avoid later disappointments through underexploitation of some of the indications. AREAS OF MISMATCH IN PRODUCT SUPPLY AND DEMAND Pharmaceutical companies now look to inlicense a greater proportion of their R&D pipeline than they formerly did. Thus, the nature of what exactly is available for potential inlicensing is now playing a greater role in deciding where, therapeutically, R&D investments are made. If there is little available from third parties in a particular therapeutic area, then it falls upon the company itself to generate compounds internally if it really intends to be active in that area. On the other hand, what start-up companies and other smaller companies are focusing upon may not necessarily match what leading pharmaceutical companies are interested in inlicensing. We detect several such areas of mismatch. Cancer is by far the most intensively researched therapeutic area in terms of the number of projects, accounting for well over 20% of total pharmaceutical R&D projects. Even with aboveaverage attrition rates, many cancer products should reach the market over the next ten years. Although cancer is certainly among the most popular areas for leading pharmaceutical companies, it is by no means the leading area. It is ranked fourth in terms of top 20 company activity, with four companies no longer qualifying by our standard of having cancer projects in Phase II development or beyond. Fortunately, at least two of the companies we term inactive, Merck & Co. and

66

THERAPEUTIC AREA TRENDS IN R&D OF THE TOP 20 PHARMACEUTICAL COMPANIES

Sankyo, are known to be keen to become more successful in cancer. And so this apparent mismatch between therapeutic area popularity overall versus within the top 20 may not be the end of the world for start-up companies in the cancer area. We suspect, however, that compounds that do come through R&D with less than impressive proÞles (perhaps a majority) will not easily attract interest from potential marketing partners. They will be crowded out by more impressive products. A more obvious area of mismatch is vaccines. There are, for example, well over 50 prophylactic vaccine projects at Phase II or beyond, and approximately 20 therapeutic vaccines are also in development. Well under half of these projects are associated with the top 20 pharmaceutical companies. But only four of these top 20 companies are actively involved in vaccine R&D. This must surely be an opportunity for pharmaceutical companies short of promising new products—if only they will abandon their reluctance to be involved with vaccines. Similarly, some three dozen immunosuppressants have reached Phase II or beyond, the majority of which are being developed to prevent post-transplantation rejection. Only about a quarter of these projects are associated with the top 20 companies—not surprisingly, given that only four of the top companies are active in this Þeld, according to our deÞnition. Again, is reluctance to be involved with the transplantation Þeld preventing companies from taking up potentially promising new products? Thus, in several areas the supply of relatively advanced compounds from all sources appears likely to exceed demand by major pharmaceutical companies. In light of the general shortage of new products, major companies could do well to reconsider whether they should be more ßexible in reviewing opportunities from the less popular therapeutic areas. Naturally, pharmaceutical companies outside the top 20 can also consider opportunities in these less popular areas. Some of the opportunities may prove of insufÞcient commercial potential for the top 20 companies but may be adequate for companies at the next level down. Where the latter companies do decide to become involved, they stand to beneÞt from the fact that they will face less intensive competition in the market than they would with products in the more popular therapeutic areas. Our analysis thus suggests that because the strength of competition in different therapeutic areas is less transparent in R&D than it is in the market, companies are not necessarily succeeding in identifying some good product opportunities; we therefore believe there is considerable scope for seeking these out.